Skip to main content
Download PDF

Patient centered guidelines for the laboratory diagnosis of Gaucher disease type 1

Orphanet Journal of Rare Diseases volume 17, Article number: 442 (2022) Cite this article

Abstract

Gaucher disease (GD) is an autosomal recessive lysosomal storage disorder due to the deficient activity of the acid beta-glucosidase (GCase) enzyme, resulting in the progressive lysosomal accumulation of glucosylceramide (GlcCer) and its deacylated derivate, glucosylsphingosine (GlcSph). GCase is encoded by the GBA1 gene, located on chromosome 1q21 16 kb upstream from a highly homologous pseudogene. To date, more than 400 GBA1 pathogenic variants have been reported, many of them derived from recombination events between the gene and the pseudogene. In the last years, the increased access to new technologies has led to an exponential growth in the number of diagnostic laboratories offering GD testing. However, both biochemical and genetic diagnosis of GD are challenging and to date no specific evidence-based guidelines for the laboratory diagnosis of GD have been published. The objective of the guidelines presented here is to provide evidence-based recommendations for the technical implementation and interpretation of biochemical and genetic testing for the diagnosis of GD to ensure a timely and accurate diagnosis for patients with GD worldwide. The guidelines have been developed by members of the Diagnostic Working group of the International Working Group of Gaucher Disease (IWGGD), a non-profit network established to promote clinical and basic research into GD for the ultimate purpose of improving the lives of patients with this disease. One of the goals of the IWGGD is to support equitable access to diagnosis of GD and to standardize procedures to ensure an accurate diagnosis. Therefore, a guideline development group consisting of biochemists and geneticists working in the field of GD diagnosis was established and a list of topics to be discussed was selected. In these guidelines, twenty recommendations are provided based on information gathered through a systematic review of the literature and two different diagnostic algorithms are presented, considering the geographical differences in the access to diagnostic services. Besides, several gaps in the current diagnostic workflow were identified and actions to fulfill them were taken within the IWGGD. We believe that the implementation of recommendations provided in these guidelines will promote an equitable, timely and accurate diagnosis for patients with GD worldwide.

Background

Gaucher disease (GD- OMIM #230800) is an autosomal recessive lysosomal storage disorder due to the deficient activity of the lysosomal hydrolase, acid beta-glucosidase (GCase; EC 3.2.1.45). The enzyme is present in the lysosomes of all nucleated cells and cleaves the beta-glucosidic linkage of glucosylceramide (GlcCer) yielding glucose and ceramide. Therefore, the deficiency of GCase leads to the progressive lysosomal accumulation of GlcCer and its deacylated derivate, glucosylsphingosine (GlcSph) mainly in the monocyte/macrophage system, resulting in multiorgan dysfunction [1].

The disease presents as a continuum of phenotypes, ranging from severe forms presenting at birth to very mild phenotypes. However, in the words of Knudson, broadly speaking, three main forms of the disease can be recognized: Type 1, non-cerebral; Type 2, cerebral acute; Type 3 cerebral chronic [2].

Type 1 GD (MIM No. 230800), the most common phenotype, is characterized by enlargement and dysfunction of the liver and spleen, displacement of normal bone marrow by storage cells and bone damage leading to infarctions and fractures. Although type 1 GD is considered a non-neuronopathic form, there is increasing evidence of neurological involvement in these patients (ie Parkinson syndrome, and Lewy body dementia [3,4,5,6,7]. Type 2 GD (MIM No. 230900) is a rare phenotype associated with an acute neurodegenerative course and death at a very early age; while type 3, the chronic neuronopathic GD (MIM No. 231000), comprises an extremely heterogeneous group of patients who present with either attenuated or severe systemic disease associated with neurological involvement originating in childhood to early adulthood [8, 9].

The human GCase is encoded by the GBA1 gene (GRCh37/hg19 Chromosome 1: 155,204,239 to 155,214,653), located on chromosome 1q21. The GBA1 gene is approximately 7.5-kb long and contains 11 exons. A highly homologous 5.5 kb-pseudogene (GBAP; MIM No. 606463; GenBank accession no. J03060.1) has been located 16 kb downstream from the active gene [10].

GCase protein is synthesized on polyribosomes as a 55-kDa peptide, which is then translocated into the endoplasmic reticulum (ER), where it is modified by the addition of high mannose oligosaccharides and transported to the trans-Golgi network from where it is trafficked to the lysosomes [11]. GCase protein is targeted to the lysosomal compartment through a mannose 6-phosphate-independent receptor, the lysosomal integral membrane protein type 2 (LIMP-2) [12], a trans-membrane protein mainly found in the lysosomes and late endosomes [13, 14]. At the acidic lysosomal pH, LIMP-2 dissociates from GCase enabling enzymatic activity facilitated by the co-factor Saposin C (Sap C) [15,16,17,18].

The diagnosis of GD is based on the demonstration of deficient GCase activity in cells and the identification of pathogenic variants in the GBA1 gene.

Latterly, the development of new technologies has improved the diagnostic capacity of expert laboratories. At the same time, increased access to these technologies has led to an exponential growth in the number of diagnostic laboratories that offer GD testing. However, both biochemical and genetic diagnosis of GD are challenging and to date no specific, evidence-based guidelines for the laboratory diagnosis of GD have been published.

The objective of the guidelines presented here is to provide evidence-based recommendations for the technical implementation and interpretation of biochemical and genetic testing for the diagnosis of GD to ensure a timely and accurate diagnosis for patients with GD worldwide.

Methods

The guidelines have been developed by members of the Diagnostic Working group of the International Working Group of Gaucher Disease (IWGGD), a non-profit network established to promote clinical and basic research into GD for the ultimate purpose of improving the lives of patients with this disease.

One of the goals of the IWGGD is to support the provision of equitable access to diagnostic testing and the introduction of standardized procedures that ensure patients with GD can readily obtain an accurate diagnosis.

A guideline development group (GDG) consisting of biochemists and geneticists working in the field of GD diagnosis was therefore established and a list of guideline topics were selected for development.

A systematic literature review on GD biomarkers, biochemical diagnosis, GCase activity, molecular diagnosis and GBA1 mutations was carried out using Medline and the Cochrane Library. The literature search on molecular diagnosis and GBA1 mutations was limited to the last 20 years. The following search terms were used: “Gaucher” and “biomarkers” or “chitotriosidase or CCL18 or PARC or glucosylsphingosine, lysoGL1 or lysoGb1 or ACE or angiotensin converting enzyme or tartrate resistant acid phosphatase or TRAP or tartrate-resistant acid phosphatase”; “Gaucher” and “activity” and “fibroblasts or leukocytes” and “sensitivity or specificity or predictive value or analytical range”; “Gaucher” and “dry blood spot or dried blood spot or DBS”; “Gaucher” and “NGS”; “GBA or GBA1” and “NGS or Sanger”; “Lysosomal storage disorders” and “NGS”; “Gaucher” and “frequency and mutation”; “Gaucher” and “genotype” and “registry”. Searches were limited to English language publications only.

One hundred eighty-six papers were selected as relevant.

References related to a single topic (i.e., biomarkers, enzyme activity, genetic testing) were pulled together and the GDG was divided into subgroups to critically revise references, grade them, write a draft summarizing evidence and formulate recommendations.

The group met three times virtually (December 10th, 2020; July 12th, 2021; December 21, 2021) and corresponded by email regularly for the duration of the guideline development.

All GDG members discussed the draft documents. Evidence levels were classified in accordance with the method proposed by Burns et al. [19] (Tables 1, 2).

Table 1 Level Type of evidence
Full size table
Table 2 Grade of recommendation & criteria
Full size table

These guidelines will be revised every 2 years to update the recommendations in light of the development and validation of novel diagnostic methods.

Topics:

  1. 1.

    Biomarkers of GD assisting in diagnosis

    1. A.

      Biomarkers described in GD:

      • Chitotriosidase activity

      • PARC/CCL18 (pulmonary and activation-regulated chemokine)

      • Glucosylsphingosine (GlcSph, lysoGL1, lysoGb1).

      • ACE (angiotensin-converting enzyme)

      • TRAP (tartrate-resistant acid phosphatase)

      • gpNMB (glycoprotein nonmetastatic melanoma protein B)

    2. B.

      Biological materials and methods used to assess recommended biomarkers

  2. 2.

    Enzyme activity

    1. A.

      In what samples glucocerebrosidase (GCase) activity can be measured?

    2. B.

      How GCase activity can be measured?

    3. C.

      What is the role of enzymatic activity in GD diagnosis?

    4. D.

      How to validate GCase assay in the laboratory?

  3. 3.

    Genetic testing

    1. A.

      What is the role of genetic testing in the diagnosis of GD?

    2. B.

      How should molecular testing be performed?

    3. C.

      Conditions with a biochemical profile suggestive of GD and no pathogenetic variants in GBA1 gene

  4. 4.

    Use of Dried Blood Spot (DBS) samples for diagnosis in external laboratories

  5. 5.

    Final conclusions and algorithms

  6. 6.

    Future challenges

Biomarkers of Gaucher disease assisting in diagnosis

Biomarkers are in general chemical entities, ranging from simple metabolites to complex proteins, which indicate the presence of a biological process linked to the clinical manifestations and outcome of a particular disease. They are the focus of much research and, when available, they play a critical role in the diagnosis, monitoring of disease progression as well as the assessment of therapeutic interventions.

An ideal biomarker should fulfill a number of criteria.

For diagnostic purposes it should be significantly elevated in the disease with no overlap in the values obtained in untreated patients and quantification in healthy subjects. The analyte should not be influenced by factors that are unrelated to the disease. It should change in response to specific treatment. Finally, reliable, fast, and cheap methods should be available for its estimation in easily accessible biological materials (For FDA view on Biomarkers see: https://www.fda.gov/about-fda/innovation-fda/fda-facts-biomarkers-and-surrogate-endpoints).

Distinct biomarkers of GD can be recognized. The first category is associated with the presence of Gaucher cells (e.g. Chitotriosidase and CCL18/PARC) while the second includes the lipid, glucosylsphingosine (GlcSph)- sometimes termed ‘lysoGL1’ or ‘lysoGb1’ in literature supported by different companies- which accumulates as a result of the deficiency of GCase activity in cells.

Biomarkers described in Gaucher diseases

Chitotriosidase Chitotriosidase is the human analogue of chitinases from lower organisms; the enzyme is released from pathological macrophages in Gaucher disease.

Sensitivity: In terms of diagnosing GD, assaying plasma chitotriosidase activity is commonly employed in many centers as a first line screening test. The activity of chitotriosidase in plasma is elevated up to 1000-fold above the mean values in a healthy reference population. In the initial studies of chitotriosidase, plasma activity was found to be elevated on average 641-fold (median control plasma, 20 nmol/mL/h; range, 4–76 nmol/mL/h; median GD plasma, 12 824 nmol/mL/h; range, 3122–65 349 nmol/mL/h) [20]. Several subsequent reports have confirmed these findings [21,22,23,24]. Generally, higher plasma chitotriosidase activity is observed in type 1 patients than patients with types 2 and 3. Increased activity has also been reported in asymptomatic/ pre-symptomatic patients identified through the screening of family members of an index case [25].

The interpretation of plasma chitotriosidase activity is complicated by the occurrence of an intragenic 24-base pair (bp) duplication in the chitotriosidase gene CHIT1, which prevents the formation of chitotriosidase protein. This effectively null allele is frequent in most populations, and among GD patients, where one in every three individuals is a heterozygous carrier and about one in every 20 individuals is homozygous for the mutation [26]. Several other mutations which affect chitotriosidase activity, have been described [27,28,29,30,31].

Specificity: Increased plasma chitotriosidase activity is not unique to GD patients. Modest elevation of activity is also found in many different lysosomal and non-lysosomal diseases such as Niemann-Pickdisease type C, Acid sphingomyelinase deficiency, Alagille syndrome, Amyotropic lateral sclerosis, hydrops fetalis due to congenital herpes virus infection, neonatal systemic candidiasis, sarcoidosis, leprosy, arthritis, multiple sclerosis, thalassemia, chronic obstructive pulmonary disease (COPD), malaria, and atherosclerosis. Generally, although the levels of activity detected in these disorders may be within the range observed in GD (especially those patients receiving specific therapy), the values are lower than those found in GD patients. Indeed, in the absence of the intragenic duplication in CHIT1, a marked elevation of chitotriosidase activity in plasma appears to be characteristic of and diagnostic for GD [20, 32,33,34,35,36,37,38,39,40,41]. Individuals who are homozygous for this CHIT1 allele have effectively no or near-absent chitotriosidase activity.

PARC/CCL18: pulmonary and activation-regulated chemokine (PARC, systematic name CCL18), a member of the C–C chemokine family which like chitotriosidase, accumulates in the alternatively activated macrophages that accumulate in GD “Gaucher cells” [42] but appears to be actively released.

Sensitivity: A 10- to 50- fold increase in the abundance of PARC/CCL18 has been reported in plasma and serum of symptomatic GD patients compared with healthy individuals [23, 24, 43]. Increased PARC/CCL18 polypeptides release s has been reported in the asymptomatic identical twin of a patient with severe disease which were substantially lower than in the symptomatic patient. PARC/CCL18 is stable upon storage and multiple freeze thaw cycles.

Specificity: Increased concentrations that can overlap those found in GD have been described in patients with a-mannosidosis and Niemann-Pick disease type A and B [23, 34]. Non-lysosomal storage diseases with increased PARC/CCL18 levels include atherosclerosis, rheumatoid arthritis, beta-thalassemia, sarcoidosis [36, 44,45,46]. So far, no genetic variations that significantly alter the concentrations of PARC/CCL18 have been described. Of note, PARC/CCL18 chemokine is not expressed in mice.

Glucosylsphingosine (a.k.a. lysoGL1, lysoGb1)

Sensitivity: an average 180—fold increase in the concentration of GlcSph has been reported in plasma and serum of symptomatic type 1 GD patients compared with healthy individuals [47, 48]. A similar abnormality is noted in mice and zebrafish with deficient GCase [49,50,51]. This characteristic abnormality has been confirmed by numerous laboratories worldwide (e.g. [52, 53]; recently reviewed in [54, 55]).

Specificity: More modestly increased levels of plasma GlcSph have also been noted in patients suffering from Action Myoclonus Renal Failure syndrome with a defective LIMP-2 [56], patients with Sap C deficiency [57] and in some patients with Niemann-Pick disease type C [58].

ACE (angiotensin-converting enzyme): A 2–tenfold increase in ACE has been described in serum/plasma of GD patients apparently originating from storage cells [23, 59,60,61,62,63,64,65]. Increased serum/plasma ACE has been reported in other disorders involving activation of the monocyte/macrophage lineage and sarcoidosis is the most frequent and the better studied [66]. Increased activity is not observed in all GD patients [60] up to fivefold variation in blood ACE across a population can be observed and several mutations/polymorphisms in the ACE gene have been described which result in increased ACE blood levels [67, 68]. ACE activity can be repressed in patients who are take ACE inhibitors [69].

TRAP (tartrate-resistant acid phosphatase): TRAP was the first biomarker to be assayed in the diagnosis of GD [70]. TRAP is not specific for GD and the observed increase in the serum is modest. It is unstable in the blood and shows marked analytical variability [23, 62]. In interpreting TRAP serum levels, its increased activity in children as compared to adults should be taken into consideration together with its thermo-instability [71].

gpNMB (glycoprotein nonmetastatic melanoma protein B): gpNMB has been identified by proteomics analysis of laser dissected Gaucher cells from GD spleens [42, 65, 72]. It is selectively overexpressed by Gaucher cells that release a soluble fragment into plasma that can be conveniently detected by ELISA. The soluble fragment of gpNMB is found to be elevated over 50-fold in plasma of patients with type 1 GD [72] and was also found to be elevated in human NPC plasma samples [65]. A recent investigation confirms the value of soluble gpNMB as a plasma marker of Gaucher cells and substantiates its diagnostic potential [73]. However further studies are needed before its role as a diagnostic biomarker is established.

  • Recommendation #1: Based on the data available to date, it is recommended that chitotriosidase activity, PARC/CCL18 or GlcSph concentrations can be used as a first line test when the diagnosis of GD is suspected.

  • If chitotriosidase activity is the only assessed biomarker and the result is normal, the presence of the 24 bp duplication in the CHIT1should be excluded. In these cases, measurement of the PARC/CCL18 and/or GlcSph is recommended.

  • However, a suspected diagnosis of GD needs to be established by assay of GCase activity (in peripheral blood leukocytes or extracts of cultured fibroblasts), preferably supported by molecular analysis of the GBA1 gene or by the identification of biallelic pathogenetic variants in the GBA1 gene

  • Level of evidence: II (cohort studies /case series with consistent results/ research articles)

  • Grade: B (Recommendation)

Biological materials and methods used to assess recommended biomarkers

Assaying chitotriosidase activity: The biological material to be used is serum and/or plasma. The enzyme in plasma is stable upon storage and multiple freeze thaw cycles, (storage: stable at room temperature for 24 h; storage at -30 after 8 months recovery 95.3–102%, data presented by Aerts et al. GD Biomarker Qualification Workshop, September 2010, FDA Campus). Although the use of DBS in the diagnosis of lysosomal storage disorders has become increasingly popular mainly due to its convenience, at present extensive studies documenting sensitivity and specificity of assaying chitotriosidase activity in this type of biological material are not yet available [74,75,76,77]. The activity of chitotriosidase in plasma/serum can be determined using the fluorogenic substrate 4- methylumbelliferyl-β-D-N,N′,N″-triacetyl-chitotrioside (4MU-C3).However, the assay is complicated by the ability of chitotriosidase to transglycosylate as well as hydrolyze this substrate and thus the reaction has nonlinear kinetics with respect to time shows non-Michaelis–Menten behaviour [78]. Therefore, it is essential that special care is taken to ensure that the enzyme activity is truly proportional to the amount of chitotriosidase protein and there is an urgent need to standardize the assay across laboratories.. Alternatively, a far more convenient, sensitive, and accurate detection can be achieved by measuring the activity of chitotriosidase toward the fluorogenic substrate 4-methylumbelliferyl-deoxychitobiose (4MU-dC2). Chitotriosidase shows normal Michaelis–Menten kinetics with this substrate, allowing the use of saturating substrate concentrations. Thus, a more accurate and robust assay is now available [78, 79].

Measurement of the levels of PARC/CCL18: The biological material to be used is serum and/or plasma. PARC/CCL18 is stable upon storage and multiple freeze thaw cycles (storage: stable at room temperature for 48 h; storage at-30; 8 month recovery 107–109%, data presented by Aerts et al. GD Biomarker Qualification Workshop, September 2010, FDA Campus). Its levels cannot be reliably estimated using SELDI-TOF but enzyme-linked immunosorbent assay (ELISA) and dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) can be used for reliable estimation [24, 43, 80].

Measurement of the levels of GlcSph: Different techniques can be used for detection of which LC–MS/MS is presently the most sensitive. Its levels cannot be reliably estimated using SELDI-TOF. Reliable determination of absolute concentrations of GlcSph by mass spectrometry requires use of an appropriate internal standard. The concentration of GlcSph can be measured in either plasma or serum. GlcSph can be quantified in previously frozen serum or plasma samples.

GlcSph has been reported to be also increased in DBS of GD patients [74, 81, 82]. However, the outcome of extensive studies documenting specificity, stability and the impact of sample storage and shipping conditions on sensitivity of this biomarker in DBS is not yet available.

  • Recommendation #2: The biological material to be used for assessment of recommended biomarkers is serum and/or plasma. Monocentric studies report good sensitivity of DBS GlcSph assessment in identifying GD patients. However, the outcome of extensive studies documenting specificity, stability and the impact of sample storage and shipping conditions on sensitivity of this biomarker in DBS is not yet available.

  • Recommendation #3: Chitotriosidase can be measured using fluorogenic substrates: 4-methylumbelliferyl-β-D-N,N′,N″-triacetyl-chitotrioside (4MU-C3) or 4-methylumbelliferyl-deoxychitobiose (4MU-dC2), which allows a more convenient, sensitive, and accurate measurement of activity. If 4MU-C3 is used it is important to ensure that the enzyme activity is truly proportional to the amount of chitotriosidase protein and the need to standardize the assay across laboratories is urgent and is underway through the IWGGD Biomarkers & Materials working group.

  • Recommendation #4: Both enzyme-linked immunosorbent assay (ELISA) and dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA) can be used for reliable estimation of PARC/CCL18 concentrations, while these cannot be reliably estimated using SELDI-TOF.

  • Recommendation #5: The most sensitive technique to assess GlcSph is LC–MS/MS. Reliable determination of absolute concentrations of GlcSph by mass spectrometry requires use of an appropriate internal standard. Its levels cannot be reliably estimated using SELDI-TOF.

  • Level of evidence: II (cohort studies /case series with consistent results/ research articles)

  • Grade: B (Recommendation)

Enzyme activity

The metabolic defect in Gaucher disease (GD) is an inherited deficiency of lysosomal membrane associated acid β-glucocerebrosidase (GCase) [83]. The basic function of GCase is degradation of the glycosphingolipid glucosylceramide (GlcCer), also known as glucocerebroside within acid pH to ceramide and glucose [84]. The gold standard for GD diagnosis is the demonstration of deficient GCase activity measured in peripheral blood leukocytes and/or cultured skin fibroblasts homogenates. Traditionally enzyme activity was measured by using the natural substrate glucocerebroside [85]. Nowadays, enzyme assay is carried out by the use of an artificial substrate 4-MU- β-D-glucoside. For this reason, and to avoid misinterpretation, enzyme activity assayed by the use of artificial substrate will be called BGLU.

In what samples BGLU activity can be measured?

The BGLU activity could be measured in different samples such as DBS, leukocytes, fibroblasts and in case of prenatal diagnosis in chorionic villi sampling (CVS) or cultured amniocytes [86]. BGLU could be measured in DBS samples as a first-line laboratory test. Pre-analytical requirements are critical for reliable BGLU results from DBS samples. DBS can be obtained by application of 50–75 μL drops of blood obtained by venipuncture into heparin tubes and spotted on the Whatman®903 or S&S903 filter paper. Another option is application of the same amount of blood after finger prick on filter paper collection device onto printed circles [87, 88]. DBS should be dried for 4 h at room temperature avoiding direct illumination, and then packed in a sealed plastic bag with desiccant, and stored at 4 °C until analysis [89]. Exposure of DBS to both heat and humidity can destroy enzyme functions rapidly. Moreover, an incomplete mixed blood before spotting can result in significant variation on enzyme activity [90].

The use of DBS as first line laboratory test offers many advantages over leukocytes or fibroblasts samples including easy collection methodologies, need of a small amount of blood, and simpler transportation as samples can be shipped via regular mail at room temperature. If the DBS sample is treated appropriately, the BGLU remain stable at least for 21 days [91,92,93]. DBS has limitations for measurement of BGLU activity. The volume of blood applied, hematocrit, recent blood transfusions and other preanalytical steps such as drying time, homogeneity and extraction of the analyte influences the quality of the DBS sample [94]. To ensure integrity of BGLU activity and to avoid false positive results, another lysosomal enzyme should be measured as a control enzyme with approximately same stability at room temperature. The value of the control (reference) sample should generally lie between the mean ± twostandard deviations [95].

Different studies have shown good sensitivity and specificity, above 95%. However, enzyme testing in DBS has a low positive predictive value (of < 45% on average) [96,97,98,99,100,101,102,103,104].

Patient leukocytes or cultured skin fibroblast homogenates are the gold standard for measurement of BGLU activity. Leukocytes as the BGLU source are obtained by separation from approximately 5–10 ml of blood, drawn from the patient in potassium EDTA or heparin tubes. Moreover, skin fibroblasts should be used when patients have received blood transfusions or when discordant results are obtained with white blood cells. The shipment of blood samples to the reference laboratory should be carried out at 4 °C [105]. The isolation of leukocytes from the whole blood should be completed within 24 h after blood collection using dextran sedimentation or the ammonium chloride lysis method [106,107,108,109]. The pellet of isolated leukocytes can be stored for at least 20 days at − 20 °C before enzyme activities are determined [108].

Homogenates prepared from cultured fibroblasts are labour intensive, since they require a skin biopsy (requiring no more than local anesthesia) transport in particular medium followed by transfer to medium for a primary cell culture of skin fibroblasts (avoiding the risk of contamination). The time taken for adequate fibroblast outgrowth to obtain a confluent cell monolayer varies but is generally about three weeks. Shipment of cultured fibroblasts should be at ambient room temperature, avoiding freezing, in a tube, dish or sealed flask (T25 or T75) containing culture media [110]. There are some potential interfering factors in the assays: excessive transport time, lack of viable cells, bacterial or mycoplasma contamination, exposure of the specimen to temperature extremes (freezing or > 30 °C).

The use of gold standard samples requires a homogenisation step with a metal tip sonicator, and total protein measurement of the homogenate [105, 111].

  • Recommendation #6: BGLU activity can be measured in dried blood spots (DBS) samples as a first-line test. However, GD diagnosis should never be rely solely on DBS enzyme activity measurement. Patient leukocytes or cultured skin fibroblast homogenates are the gold standard for measurement of BGLU activity and confirmation of GD diagnosis; skin fibroblasts, while more laborious and expensive to obtain, have the advantage that they can be cryopreserved in liquid nitrogen almost indefinitely and if adequately aliquoted, can be used repeatedly for study

  • Level of evidence: II, III and IV (Well-designed cohort, case–control study, case reports)

  • Grade: B (Recommendation)

How GCase activity can be measured?

BGLU activity can be measured using fluorometric methods, tandem mass spectrometry or by digital microfluidics platforms. Fluorometric methods are based on the artificial substrate 4-methylumbelliferyl-β-D-glucopyranoside (4-MUG). They are mostly performed in microtiter plates [112,113,114]. The sample is put into a reaction mixture of acidic pH, sodium deoxytaurocholate, and the fluorogenic substrate, 4-methylumbelliferyl β-D-glucopyranoside (4-MUG). Sodium deoxytaurocholate is added in order to inhibit the non-lysosomal isoenzyme BGLU activity [93, 115,116,117,118]. Fluorometric enzyme assays for BGLU onto digital microfluidic platforms have the potential for simple, rapid and high-throughput selective screening of BGLU activity [119,120,121,122]. Beside digital microfluidic fluorometry, there are other available compact digital microfluidic platforms (e.g. electro-wetting based digital microfluidics) [123].

Tandem mass spectrometry enzyme assays with (LC–MS/MS) or without (MS/MS) liquid chromatography are based on non-fluorometric synthetic substrates [124,125,126]. This approach may be particularly suitable for high-throughput analyses with a large number of individuals at-risk and/or for newborn screening for GD [103, 127, 128]. All three technologies (approaches) are suitable for selective screening BGLU activity [96].

  • Recommendation #7: BGLU activity could be measured using artificial substrate with fluorometric methods, tandem mass spectrometry or by digital microfluidics platform. The fluorometric method is accepted as the gold standard assay in leukocyte/fibroblast lysates. Tandem mass spectrometry or digital microfluidics platforms are generally used for DBS samples in screening studies.

  • Level of evidence: II, III and IV (Well-designed cohort, case–control study, case reports)

  • Grade: B (Recommendation)

What is the role of enzymatic activity in GD?

Enzyme determinations in DBS samples are useful screening tests in clinically suspected individuals. Samples with BGLU activity below cut-off values require confirmation by measuring BGLU activity in gold standard samples: homogenates of leukocytes or fibroblasts [92, 112]. Whenever subjects present suggestive GD symptoms they must be reassessed even in the presence of normal BGLU from DBS testing [104].

An enzyme activity result of less than 15% of normal activity in homogenates of leukocytes or fibroblasts is diagnostic of GD [129]. Residual enzyme activity does not correlate with disease severity. Enzyme testing is not suitable for identification of carriers of GD nor of saposin C deficiency [118, 130, 131]. Heterozygotes may have half-normal enzyme activity, but overlapping with activity levels of healthy controls, rendering enzymatic testing for carrier status unreliable [132,133,134].

  • Recommendation #8: The DBS samples are useful as a first line test in clinically suspected individuals. Samples with BGLU activity below cut-off values always require confirmation by measuring BGLU activity in gold standard samples: homogenates of leukocytes or fibroblasts. The demonstration of deficient (below 15% of mean normal activity) BGLU enzyme activity in leukocyte and/or skin fibroblast homogenates confirms GD diagnosis.

  • Residual enzyme activity does not correlate with disease severity and the test is not suitable for diagnosis of heterozygotes of GD nor of saposin C deficiency.

  • Level of evidence: II, III and IV (Well-designed cohort, case–control study, case reports)

  • Grade: B (Recommendation)

How to validate GCase assay in the lab?

To ensure the quality of BGLU testing performance, each laboratory should establish its own Quality Management (QM) system according to ISO15189 and participate in both internal and external quality assessments. The internal audit program monitors operations throughout the testing process and the quality system. For quality control purposes, it is necessary to include an appropriate blank and at least one affected control and one normal control sample for each run of enzyme assays. All assays should be performed in duplicate. The cut-off range, normal range, and disease range should be established by the laboratory based on its own analysis [135]. The inter-laboratory variance of numerical enzyme activity determinations could be large [136]. Reproducibility was demonstrated by intra- (n = 6) and inter-assay (n = 10) results using threshold of %CV < 15. Therefore, quality assurance and improvement in diagnostic proficiency have become essential in this area [137]. The enzyme assay is made in house by each laboratory based on the original published methods. It implies differences in units (pmol/h/disk, μmol/h/l, μmol/h/mg protein), disease cut-off; reference range, limit of detection (LOD) and limit of quantitation (LOQ). For this reason, laboratory reports from reference labs should include an interpretation of the result that reflects the conclusion of the result as normal or deficient, possible limitations of the test, and recommendations for additional testing if applicable.

The European Research Network for Evaluation and Improvement of Screening, Diagnosis, and Treatment of Inherited Disorders of Metabolism (ERNDIM) serves as an external proficiency testing program for clinical diagnostic laboratories, providing lyophilized fibroblasts for eight lysosomal storage diseases (LSD) enzymes [138]. For laboratories testing lysosomal enzymes on DBS, the Newborn Screening Quality Assurance Program (NSQAP) at Centers for Disease Control and Prevention (CDC) provides quality control (QC) materials, proficiency testing (PT) services, and technical support in collaboration with the Newborn Screening Translation Research Initiative (NSTRI) at CDC [139, 140].

  • Recommendation 9: Each laboratory should establish its own Quality Management (QM) system, if possible, according to ISO15189 international standards and participate in both internal and external quality assessments.

  • Level of evidence: V (Review, expert opinion)

  • Grade: D (Option)

Genetic testing

GD is caused by biallelic pathogenic variants in the gene encoding the acid β glucocerebrosidase protein, GBA1 (GRCh37/hg19 Chromosome 1: 155,204,239 to 155,214,653).

A highly homologous pseudogene, GBAP (96% identity), is located 16 kb downstream of the GBA1 gene [10]. The high degree of homology, which reaches 96% in exonic regions, and the proximity between GBA1 and GBAP favours the occurrence of recombination events resulting in complex gene-pseudogene rearrangements [141, 142].

The nascent GCase polypeptide is composed of 536 amino acids, including 39 that encode a signal sequence that is later cleaved after it directs the polypeptide to transit the endoplasmic reticulum. Historically, GBA1 variants were numbered from the first residue after the cleavage of the signal peptide as amino acid number one. This legacy nomenclature is still used (herein reported between brackets and without the prefix p.), although it does not comply with contemporary nomenclature standards of the Human Genome Variation Society (HGVS).

What is the role of genetic testing in the diagnosis of GD?

The identification of biallelic pathogenetic variants in the GBA1 gene confirms the diagnosis of GD.

Genetic testing is performed in subjects displaying absent or low residual BGLU activity in cells to support the diagnosis and provide appropriate genetic counseling to family members.

Genetic testing can be done as a primary test for GD diagnosis. However, since many GBA1 variants are private, the chances of finding variants of uncertain significance (VUS) are quite high [25, 143,144,145,146,147,148,149,150,151,152,153,154,155]. In this case, confirmation of diagnosis through the assessment of enzymatic activity in patient’s cells is mandatory.

Variants should be classified following the American College of Medical Genetics (ACMG) criteria and in the case of VUS, pathogenicity should be assessed by functional analysis.

In addition, molecular testing of known familial variants represents the most reliable method to identify GD carriers since enzymatic activity does not discriminate between carriers and normal subjects [132].

According to The Human Gene Mutation Database (HGMD-Professional 2021.1), 540 variants of the GBA1 gene have been reported to date, although not all of them are linked to GD. Indeed, 403 of them have been associated with GD.

Diverse variants have been reported: missense and nonsense variants, splice junction variants, deletions and insertions of one or more nucleotides and complex alleles (complex rearrangements) resulting from gene conversion or gene fusion with the downstream pseudogene GBAP. However, missense and nonsense variants, are the most frequently identified in GD patients worldwide [156].

The frequency distribution of GBA1 variants differs across ethnic groups. While 4 pathogenetic variants (N370S; L444P, c.84–85 insG; IVS + 1G > A) account for 90% of alleles within Ashkenazi Jews, they account only for about 50–75% of alleles in non-Jewish populations. In addition, about 10% of patients present large deletions/recombinant alleles [25, 143,144,145,146,147,148,149,150,151,152,153,154,155, 157,158,159,160,161,162,163,164].

  • Recommendation #10: Molecular analysis of the GBA1 gene should always be performed when biomarker results or phenotype are at odds with the enzymology and is highly recommended in subjects with BGLU activity below normal reference intervals in cells to further support/confirm the diagnosis of GD and provide genetic counseling. Testing of familial variants and genetic counseling should be made available to all at risk family members.

  • Recommendation #11: Genetic testing could be done as a primary test (before testing enzymatic activity). However, results should be interpreted with caution since GBA1 testing is challenging (see below), depending on the method used the detection of large deletions and/or recombinant alleles will not be possible and VUS are often identified. Therefore, confirmation of diagnosis through the assessment of enzymatic activity in patient’s cells is mandatory.

  • Recommendation #12: Genetic testing is the most reliable method to detect heterozygous carriers and it should be made available to family members at risk of being a carrier.

  • Recommendation #13: In all cases, molecular testing should be accompanied by a pre and post-test genetic counseling delivered by a counsellor experienced in GD to ensure informed choices.

  • Level of evidence: II and IV (retrospective cohort studies or case series with consistent results)

  • Grade: B (Recommendation)

How should molecular testing be performed?

Long template specific PCR amplification of the GBA1 gene (and not the pseudogene) followed by Sanger sequencing allows the identification of single base pair variants and most recombinant alleles leading to molecular diagnosis of GD about 95–98% of cases [25, 143,144,145,146,147,148,149,150,151,152,153,154,155]; however, this method fails to detect large deletions [144, 151, 165, 166].

GBA1 gene can also be analyzed using Next-Generation Sequencing (NGS) technologies, both as a single gene or as part of targeted gene panels, Whole exome sequencing (WES) or Whole genome sequencing (WGS). In all cases, the workflow should be optimized to avoid false positive or negative results due to misalignment of reads between the gene and the pseudogene.

Strategies to specifically analyze the GBA1 as a single gene using NGS technology have been developed [167,168,169]. Such NGS strategies allow the identification of single base pair variants and recombinant alleles (excluding the Recdelta55) with high specificity and sensitivity [167, 169]. Conversely, analysis of the GBA1 gene as part of gene panels using well designed NGS strategies that consider the presence of the pseudogene, allows only the identification of point mutations, while fail to identify both large deletions and recombinant alleles due misalignment of reads with the homologous pseudogene [170,171,172,173,174].

However, NGS data analysis is a field in continuous and rapid evolution and new solutions to improve sensitivity and specificity are expected to be available in the near future [175].

Indeed, the use of PacBio long-read Single Molecule Real-Time (SMRT) for GBA1 deep sequencing has recently been developed [176]. However, this technology is still not widely available in most genetic laboratories.

Multiplex ligation-probe amplification (MLPA) kits have been developed for the identification of recombinant/deleted GBA alleles. However, commercially available kits do not discriminate between L444P mutant and RecNci alleles and do not discriminate between recombination events and deletions [151, 174, 177].

  • Recommendation #14: Sequencing analysis of GBA1 exons and intron exon boundaries should be performed as the primary molecular test. It should be performed using specific long template amplification of the GBA gene (avoiding the amplification of the pseudogene) followed by Sanger sequencing or NGS specifically designed to avoid reads misalignments. This strategy allows detecting point mutations and most recombinant alleles but is not suitable to detect large deletions.

  • Recommendation #15: GBA1 could be included in gene panels analyzed by NGS. This technology allows the detection of point mutations, although false positive results have been reported. Therefore, point mutations detected by NGS methods should always be confirmed by Sanger sequencing. Standard workflows are not suitable for the detection of large deletions or recombinant alleles.

  • Recommendation #16: Segregation of alleles by identifying variants in parents, should be determined.

  • Recommendation #17: The presence of homozygous pathogenetic variants not confirmed in parents, as well as the absence of pathogenetic variants (in one or both allele) after sequencing should always be questioned and additional investigations should be performed. In particular, multiplex ligation-probe amplification (MLPA) and mRNA analysis should be done to identify possible undetected recombinant/deleted alleles or deep intronic pathogenetic variants, respectively.

  • Recommendation #18: Variants should be classified following the ACMG criteria and in case of identification of VUS, pathogenicity should be investiagted by functional analysis.

  • Level of evidence: II and IV (retrospective cohort studies or case series with consistent results)

  • Grade: B (Recommendation)

Conditions with a biochemical profile suggestive of GD and no pathogenetic variants in GBA1 gene

Although most cases of GD are due to mutations within the GBA1 gene, a small number of patients present mutations in the PSAP gene which encodes the GCase activator, saposin C (Sap C) [178,179,180,181,182,183].

Sap C is a member of a family of four small lysosomal glycoproteins (Saps A, B, C and D), all derived by proteolytic processing from a common precursor protein, prosaposin (PSAP), encoded by the PSAP gene (NM_001042465.3) located on chromosome 10 [184, 185].

Sap C promotes rearrangement of lipid organization in lysosomal membranes favoring substrate accessibility to GCase. Therefore, mutations in the Sap C domain of PSAP result in the inability of GCase to degrade GlcCer, with the consequent accumulation within the lysosomes, leading to a GD like phenotype. These patients display increased chitotriosidase activity and increased levels of GlcSph. However, the in vitro GCase activity in cells results reduced or even normal since Sap C is not needed for the hydrolysis of the artificial substrate used in the diagnostic test in vitro [179,180,181,182,183, 186].

  • Recommendation #19: In the absence of pathogenetic variants in the GBA1 gene in subjects with a clinical phenotype compatible with GD, increased chitotriosidase activity, increased levels of GlcSph and normal or low BGLU activity in cells, a Sap C deficiency should be suspected and the PSAP gene analyzed.

  • Level of evidence: V (case reports)

  • Grade: D (Option)

Use of DBS samples for diagnosis in external laboratories

The use of dried blood spots (DBS) in the diagnosis of lysosomal storage disorders has become increasingly popular mainly due to its convenience.

As stated above, BGLU activity and GlcSph can be measured in DBS. However, results have to be interpreted with caution since BGLU testing in DBS has a very poor positive predictive value (see enzyme activity section) and although recent monocentric studies have shown encouraging results in favor of using DBS to assess GlcSph, several points require clarification before this can be recommended. In particular, stability over time of the sample (to define storage and transport time recommendations) as well as correlation between standard and DBS assays and specificity (see biomarkers and enzyme activity sections) all need to be evaluated.

  • Recommendation # 20: DBS can be used for diagnosis of GD in patients without access to in house testing. In these cases, DBS can be sent to external laboratories with expertise in GD. Pre-analytical requirements are critical for reliable results. Both BGLU and/or GlcSph can be assessed as a first line test in this type of sample. However, interpretation of the results needs caution.

  • Therefore, diagnosis should never be relied on these tests only and they should be confirmed by demonstration of biallelic pathogenetic variants in the GBA1 gene (see genetic testing section).

  • In the absence of biallelic pathogenetic variants, the assessment of BGLU activity in cells is mandatory.

  • Level of evidence: II and IV (retrospective cohort studies or case series with consistent results)

  • Grade: B (Recommendation)

Final conclusions

These guidelines address the laboratory workup for the diagnosis of GD type 1 and are intended to facilitate accurate and timely diagnosis regardless of demography and access to health care. Based on the gathered evidences and the recommendations above, a diagnostic algorithm has been developed as shown in Fig. 1 (Algorithm 1).

Fig. 1
figure 1

Diagnostic algorithm 1. *The presence of the 24 bp deletion has to be excluded if this is the only biomarker assessed and it results normal. **Subjects presenting suggestive GD symptoms must be reassessed even in the presence of normal BGLU in DBS

Full size image

The group is aware that not all patients around the world have access to in-house testing and they are obliged to rely on external laboratories, sometimes commercial services, for diagnosis. In this case, dry blood spots can be used although results have to be interpreted with caution. An algorithm for diagnosis using DBS is shown in Fig. 2 (Algorithm 2):

Fig. 2
figure 2

Diagnostic algorithm 2. *Subjects presenting suggestive GD symptoms must be reassessed even in the presence of normal BGLU in DBS

Full size image

The interpretation of the test described in this workflow can be challenging and not always straight forward. Therefore, the group recommends that expert laboratories interpret the results in the context of the clinical description of the patient. Moreover, the group strongly recommends that the report includes a clear interpretation that reflects the conclusion of the result, possible limitations of the test, and recommendations for additional testing, where applicable.

Future challenges

  1. 1.

    A standardization of assays of various plasma biomarkers is recommended. A first step in this direction is undertaken by the IWGGD working group Biomarkers & Materials.

  2. 2.

    The use of DBS to assess biomarkers (e.g. GlcSph) should be confirmed by multiple centers with special attention to the influence of storage and shipment conditions.

  3. 3.

    The potential application of plasma biomarkers to monitor disease progression and efficacy of therapeutic intervention warrants further investigation, in consultation with other IWGGD working groups.

  4. 4.

    Collection of more information on plasma biomarkers in other conditions in which a (partial) deficiency of GCase activity occurs: Niemann Pick type C, Action Myoclonus Renal Failure Syndrome, Saposin C deficiency.

  5. 5.

    Identification of biomarkers able to predict the possible neurological involvement in newly identified patients.

  6. 6.

    Development of new methods for accurate and cost/effective analysis genetic testing of GBA1.

Availability of data and materials

Not applicable.

Abbreviations

GD:

Gaucher disease

GCase:

Acid β-glucosidase

GlcCer:

Glucosylceramide

GlcSph:

Glucosylsphingosine

IWGGD:

International Working Group of Gaucher Disease

ER:

Endoplasmic reticulum

LIMP-2:

Lysosomal integral membrane protein type 2

Sap C:

Saposin C

GDG:

Guideline development group

ACE:

Angiotensin-converting enzyme

TRAP:

Tartrate-resistant acid phosphatase

gpNMB:

Glycoprotein nonmetastatic melanoma protein B

DBS:

Dried Blood Spot

COPD:

Chronic obstructive pulmonary disease

PARC:

Pulmonary and activation-regulated chemokine

4MU-C3:

4- Methylumbelliferyl-β-D-N,N′,N″-triacetyl-chitotrioside

4MU-dC2:

4-Methylumbelliferyl-deoxychitobiose

ELISA:

Enzyme-linked immunosorbent assay

DELFIA:

Dissociation-enhanced lanthanide fluoroimmunoassay

CVS:

Chorionic villi sampling

4-MUG:

4-Methylumbelliferyl β-D-glucopyranoside

BGLU:

GCase enzymatic activity assayed with the artificial substrate in vitro

LOD:

Limit of detection

LOQ:

Limit of quantitation

ERNDIM:

European Research Network for Evaluation and Improvement of Screening, Diagnosis, and Treatment of Inherited Disorders of Metabolism

LSD:

Lysosomal storage diseases

NSQAP:

Newborn Screening Quality Assurance Program

CDC:

Centers for Disease Control and Prevention

QC:

Quality control

PT:

Proficiency testing

NSTRI:

Newborn Screening Translation Research Initiative

QM:

Quality Management

VUS:

Variants of uncertain significance

ACMG:

American College of Medical Genetics

HGMD:

Human Gene Mutation Database

WES:

Whole exome sequencing

WGS:

Whole Genome Sequencing

NGS:

Next-Generation Sequencing

SMRT:

Single Molecule Real-Time

MLPA:

Multiplex ligation-probe amplification

References

  1. Beutler E, Grabowski GA. Gaucher disease. In: Scriver CR, Beaudet AL, Sly WS, Valle D, editors. The metabolic and molecular bases of inherited disease. New York: McGraw-Hill; 2001. p. 3635–68.

    Google Scholar 

  2. Knudson AG. Inborn errors of sphingolipid metabolism. Am J Clin Nutr. 1961;9:55–62.

    Article  CAS  Google Scholar 

  3. Khan A, Stimpson P, Karmolinski A, Patel N. Middle-ear involvement in type I Gaucher’s disease—a unique case. J Laryngol Otol. 2013;127:1226–9.

    Article  CAS  Google Scholar 

  4. Potnis KC, Flueckinger LB, DeArmey SM, Alcalay RN, Cooney JW, Kishnani PS. Corticobasal syndrome in a man with Gaucher disease type 1: Expansion of the understanding of the neurological spectrum. Mol Genet Metab Rep. 2018;17:69–72.

    Article  Google Scholar 

  5. D’Amore S, Page K, Donald A, Taiyari K, Tom B, Deegan P, et al. In-depth phenotyping for clinical stratification of Gaucher disease. Orphanet J Rare Dis. 2021;16:431.

    Article  Google Scholar 

  6. Biegstraaten M, Schaik IN, Aerts JMFG, Hollak CEM. “Non-neuronopathic” Gaucher disease reconsidered. Prevalence of neurological manifestations in a Dutch cohort of type I Gaucher disease patients and a systematic review of the literature. J Inherit Metab Dis. 2008;31:337–49.

    Article  CAS  Google Scholar 

  7. Chérin P, Rose C, de Roux-Serratrice C, Tardy D, Dobbelaere D, Grosbois B, et al. The neurological manifestations of Gaucher disease type 1: the French Observatoire on Gaucher disease (FROG). J Inherit Metab Dis. 2010;33:331–8.

    Article  Google Scholar 

  8. Roshan Lal T, Sidransky E. The spectrum of neurological manifestations associated with Gaucher disease. Diseases. 2017;5:10.

    Article  Google Scholar 

  9. Daykin EC, Ryan E, Sidransky E. Diagnosing neuronopathic Gaucher disease: New considerations and challenges in assigning Gaucher phenotypes. Mol Genet Metab. 2021;132:49–58.

    Article  CAS  Google Scholar 

  10. Horowitz M, Wilder S, Horowitz Z, Reiner O, Gelbart T, Beutler E. The human glucocerebrosidase gene and pseudogene: Structure and evolution. Genomics. 1989;4:87–96.

    Article  CAS  Google Scholar 

  11. Ericksonss AH, Ginnsl EI, Barrangerl JA. Biosynthesis of the Lysosomal Enzyme Glucocerebrosidase. J Biol Chem. 1985;260:14319–24.

    Article  Google Scholar 

  12. Reczek D, Schwake M, Schröder J, Hughes H, Blanz J, Jin X, et al. LIMP-2 is a receptor for lysosomal mannose-6-phosphate-independent targeting of beta-glucocerebrosidase. Cell. 2007;131:770–83.

    Article  CAS  Google Scholar 

  13. Fukuda M. Lysosomal membrane glycoproteins. Structure, biosynthesis, and intracellular trafficking. J Biol Chem. 1991;266:21327–30.

    Article  CAS  Google Scholar 

  14. Fujita H, Takata Y, Kono A, Tanaka Y, Takahashi T, Himeno M, et al. Isolation and sequencing of a cDNA clone encoding the 85 kDa human lysosomal sialoglycoprotein (hLGP85) in human metastatic pancreas islet tumor cells. Biochem Biophys Res Commun. 1992;184:604–11.

    Article  CAS  Google Scholar 

  15. Vaccaro AM, Tatti M, Ciaffoni F, Salvioli R, Barca A, Scerch C. Effect of saposins A and C on the enzymatic hydrolysis of liposomal glucosylceramide. J Biol Chem. 1997;272:16862–7.

    Article  CAS  Google Scholar 

  16. Salvioli R, Tatti M, Ciaffoni F, Vaccaro AM. Further studies on the reconstitution of glucosylceramidase activity by Sap C and anionic phospholipids. FEBS Lett. 2000;472:17–21.

    Article  CAS  Google Scholar 

  17. Abdul-Hammed M, Breiden B, Schwarzmann G, Sandhoff K. Lipids regulate the hydrolysis of membrane bound glucosylceramide by lysosomal β-glucocerebrosidase. J Lipid Res. 2017;58:563–77.

    Article  CAS  Google Scholar 

  18. Atrian S, López-Viñas E, Gómez-Puertas P, Chabás A, Vilageliu L, Grinberg D. An evolutionary and structure-based docking model for glucocerebrosidase-saposin C and glucocerebrosidase-substrate interactions - relevance for Gaucher disease. Proteins. 2008;70:882–91.

    Article  CAS  Google Scholar 

  19. Burns PB, Rohrich RJ, Chung KC. The Levels of Evidence and their role in Evidence-Based Medicine. Plast Reconstr Surg. 2011;128:305.

    Article  CAS  Google Scholar 

  20. Hollak CEM, van Weely S, van Oers MHJ, Aerts JMFG. Marked elevation of plasma chitotriosidase activity. A novel hallmark of Gaucher disease. J Clin Invest. 1994;93:1288–92.

    Article  CAS  Google Scholar 

  21. van Dussen L, Hendriks EJ, Groener JEM, Boot RG, Hollak CEM, Aerts JMFG. Value of plasma chitotriosidase to assess non-neuronopathic Gaucher disease severity and progression in the era of enzyme replacement therapy. J Inherit Metab Dis. 2014;37:991–1001.

    Article  Google Scholar 

  22. Stirnemann J, Vigan M, Hamroun D, Heraoui D, Rossi-Semerano L, Berger MG, et al. The French Gaucher’s disease registry: Clinical characteristics, complications and treatment of 562 patients. Orphanet J Rare Dis. 2012;7:77.

    Article  Google Scholar 

  23. Deegan PB, Moran MT, McFarlane I, Schofield JP, Boot RG, Aerts JMFG, et al. Clinical evaluation of chemokine and enzymatic biomarkers of Gaucher disease. Blood Cells Mol Dis. 2005;35:259–67.

    Article  CAS  Google Scholar 

  24. Raskovalova T, Deegan PB, Mistry PK, Pavlova E, Yang R, Zimran A, et al. Accuracy of chitotriosidase activity and CCL18 concentration in assessing type I Gaucher disease severity. A systematic review with meta-analysis of individual participant data. Haematologica. 2020;105:437–45.

    Article  Google Scholar 

  25. Dimitriou E, Moraitou M, Cozar M, Serra-Vinardell J, Vilageliu L, Grinberg D, et al. Gaucher disease: Biochemical and molecular findings in 141 patients diagnosed in Greece. Mol Genet Metab Rep. 2020;24:100614.

    Article  CAS  Google Scholar 

  26. Boot RG, Renkema GH, Verhock M, Strijland A, Bliek J, de Meulemeester TMAMO, et al. The human chitotriosidase gene—Nature of inherited enzyme deficiency. J Biol Chem. 1998;273:25680–5.

  27. Grace ME, Balwani M, Nazarenko I, Prakash-Cheng A, Desnick RJ. Type 1 Gaucher disease: Null and hypornorphic novel chitotriosidase mutations - Implications for diagnosis and therapeutic monitoring. Hum Mutat. 2007;28:866–73.

    Article  CAS  Google Scholar 

  28. Arndt S, Hobbs A, Sinclaire I, Lane AB. Chitotriosidase deficiency: A mutation update in an African population. JIMD Rep. 2013;10:11–6.

    Article  Google Scholar 

  29. Lee P, Waalen J, Crain K, Smargon A, Beutler E. Human chitotriosidase polymorphisms G354R and A442V associated with reduced enzyme activity. Blood Cells Mol Dis. 2007;39:353–60.

    Article  CAS  Google Scholar 

  30. Mavrikiou G, Petrou P, Georgiou T, Drousiotou A. Chitotriosidase deficiency in the Cypriot population: identification of a novel deletion in the CHIT1 gene. Clin Biochem. 2016;49:885–9.

    Article  CAS  Google Scholar 

  31. Csongrádi A, Altorjay IT, Fülöp G, Enyedi A, Enyedi EE, Hajnal P, et al. Chitotriosidase gene polymorphisms and mutations limit the determination of chitotriosidase expression in sarcoidosis. Clin Chim Acta. 2021;513:50–6.

    Article  Google Scholar 

  32. Guo Y, He W, Boer AM, Wevers RA, de Bruijn AM, Groener JEM, et al. Elevated plasma chitotriosidase activity in various lysosomal storage disorders. J Inherit Metab Dis. 1995;18:717–22.

    Article  CAS  Google Scholar 

  33. Michelakakis H, Dimitriou E, Labadaridis I. The expanding spectrum of disorders with elevated plasma chitotriosidase activity: an update. J Inherit Metab Dis. 2004;27:705–6.

    Article  CAS  Google Scholar 

  34. Brinkman J, Wijburg FA, Hollak CE, Groener JE, Verhoek M, Scheij S, et al. Plasma chitotriosidase and CCL18: early biochemical surrogate markers in type B Niemann-Pick disease. J Inherit Metab Dis. 2005;28:13–20.

    Article  CAS  Google Scholar 

  35. Vedder AC, Cox-Brinkman J, Hollak CEM, Linthorst GE, Groener JEM, Helmond MTJ, et al. Plasma chitotriosidase in male Fabry patients: a marker for monitoring lipid-laden macrophages and their correction by enzyme replacement therapy. Mol Genet Metab. 2006;89:239–44.

    Article  CAS  Google Scholar 

  36. Boot RG, Hollak CEM, Verhoek M, Alberts C, Jonkers RE, Aerts JM. Plasma chitotriosidase and CCL18 as surrogate markers for granulomatous macrophages in sarcoidosis. Clin Chim Acta. 2010;411:31–6.

    Article  CAS  Google Scholar 

  37. Iyer A, van Eijk M, Silva E, Hatta M, Faber W, Aerts JMFG, et al. Increased chitotriosidase activity in serum of leprosy patients: Association with bacillary leprosy. Clin Immunol. 2009;131:501–9.

    Article  CAS  Google Scholar 

  38. Boven LA, van Meurs M, van Zwam M, Wierenga-Wolf A, Hintzen RQ, Boot RG, et al. Myelin-laden macrophages are anti-inflammatory, consistent with foam cells in multiple sclerosis. Brain. 2006;129:517–26.

    Article  Google Scholar 

  39. Boot RG, van Achterberg TAE, van Aken BE, Renkema GH, Jacobs MJHM, Aerts JMFG, et al. Strong induction of members of the chitinase family of proteins in atherosclerosis: Chitotriosidase and human cartilage gp-39 expressed in lesion macrophages. Arterioscler Thromb Vasc Biol. 1999;19:687–94.

    Article  CAS  Google Scholar 

  40. Labadaridis J, Dimitriou E, Costalos C, Aerts J, van Weely S, Donker- Koopman WE, et al. Serial chitotriosidase activity estimations in neonatal systemic candidiasis. Acta Paediatr. 1998;87:605.

    Article  CAS  Google Scholar 

  41. vom Dahl S, Harzer K, Rolfs A, Albrecht B, Niederau C, Vogt C, et al. Hepatosplenomegalic lipidosis: What unless Gaucher? Adult cholesteryl ester storage disease (CESD) with anemia, mesenteric lipodystrophy, increased plasma chitotriosidase activity and a homozygous lysosomal acid lipase -1 exon 8 splice junction mutation. J Hepatol. 1999;31:741–6.

    Article  Google Scholar 

  42. Moran MT, Schofield JP, Hayman AR, Shi G-P, Young E, Cox TM. Pathologic gene expression in Gaucher disease: up-regulation of cysteine proteinases including osteoclastic cathepsin K. Blood. 2000;96:1969–78.

    Article  CAS  Google Scholar 

  43. Boot RG, Verhoek M, de Fost M, Hollak CEM, Maas M, Bleijlevens B, et al. Marked elevation of the chemokine CCL18/PARC in Gaucher disease: A novel surrogate marker for assessing therapeutic intervention. Blood. 2004;103:33–9.

    Article  CAS  Google Scholar 

  44. Reape TJ, Rayner K, Manning CD, Gee AN, Barnette MS, Burnand KG, et al. Expression and cellular localization of the CC chemokines PARC and ELC in human atherosclerotic plaques. Am J Pathol. 1999;154:365–74.

    Article  CAS  Google Scholar 

  45. Struyf S, Schutyser E, Gouwy M, Gijsbers K, Proost P, Benoit Y, et al. PARC/CCL18 Is a plasma CC chemokine with increased levels in childhood acute lymphoblastic leukemia. Am J Pathol. 2003;163:2065–75.

    Article  CAS  Google Scholar 

  46. Dimitriou E, Verhoek M, Altun S, Karabatsos F, Moraitou M, Youssef J, et al. Elevated plasma chemokine CCL18/PARC in β-thalassemia. Blood Cells Mol Dis. 2005;35:328–31.

    Article  CAS  Google Scholar 

  47. Dekker N, van Dussen L, Hollak CEM, Overkleeft H, Scheij S, Ghauharali K, et al. Elevated plasma glucosylsphingosine in Gaucher disease: relation to phenotype, storage cell markers, and therapeutic response. Blood. 2011;118:e118–27.

    Article  Google Scholar 

  48. Beasley J, McCaw P, Zhang H, Young SP, Stiles AR. Combined analysis of plasma or serum glucosylsphingosine and globotriaosylsphingosine by UPLC-MS/MS. Clin Chim Acta. 2020;511:132–7.

    Article  CAS  Google Scholar 

  49. Keatinge M, Bui H, Menke A, Chen YC, Sokol AM, Bai Q, et al. Glucocerebrosidase 1 deficient Danio rerio mirror key pathological aspects of human Gaucher disease and provide evidence of early microglial activation preceding alpha-synuclein-independent neuronal cell death. Hum Mol Genet. 2015;24:6640–52.

    Article  CAS  Google Scholar 

  50. Lelieveld LT, Mirzaian M, Kuo CL, Artola M, Ferraz MJ, Peter REA, et al. Role of β-glucosidase 2 in aberrant glycosphingolipid metabolism: Model of glucocerebrosidase deficiency in zebrafish. J Lipid Res. 2019;60:1851–67.

    Article  CAS  Google Scholar 

  51. Dahl M, Smith EMK, Warsi S, Rothe M, Ferraz MJ, Aerts JMFG, et al. Correction of pathology in mice displaying Gaucher disease type 1 by a clinically-applicable lentiviral vector. Mol Ther Methods Clin Dev. 2021;20:312–23.

    Article  CAS  Google Scholar 

  52. Murugesan V, Chuang WL, Liu J, Lischuk A, Kacena K, Lin H, et al. Glucosylsphingosine is a key biomarker of Gaucher disease. Am J Hematol. 2016;91:1082–9.

    Article  CAS  Google Scholar 

  53. Rolfs A, Giese AK, Grittner U, Mascher D, Elstein D, Zimran A, et al. Glucosylsphingosine is a highly sensitive and specific biomarker for primary diagnostic and follow-up monitoring in gaucher disease in a non-jewish, caucasian cohort of gaucher disease patients. PLoS ONE. 2013;8:e79732.

    Article  Google Scholar 

  54. Revel-Vilk S, Fuller M, Zimran A. Value of glucosylsphingosine (Lyso-Gb1) as a biomarker in gaucher disease: a systematic literature review. Int J Mol Sci. 2020;21:7159.

    Article  CAS  Google Scholar 

  55. van Eijk M, Ferra MJ, Boot RG, Aerts JMFG. Lyso-glycosphingolipids: Presence and consequences. Essays Biochem. 2020;64:565–78.

    Article  Google Scholar 

  56. Gaspar P, Kallemeijn WW, Strijland A, Scheij S, van Eijk M, Aten J, et al. Action myoclonus-renal failure syndrome: diagnostic applications of activity-based probes and lipid analysis. J Lipid Res. 2014;55:138–45.

    Article  CAS  Google Scholar 

  57. Motta M, Tatti M, Furlan F, Celato A, di Fruscio G, Polo G, et al. Clinical, biochemical and molecular characterization of prosaposin deficiency. Clin Genet. 2016;90:220–9.

    Article  CAS  Google Scholar 

  58. Ferraz MJ, Marques ARA, Gaspar P, Mirzaian M, van Roomen C, Ottenhoff R, et al. Lyso-glycosphingolipid abnormalities in different murine models of lysosomal storage disorders. Mol Genet Metab. 2016;117:186–93.

    Article  CAS  Google Scholar 

  59. Lieberman J, Beutler E. Elevation of serum angiotensin-converting enzyme in Gaucher’s disease. N Engl J Med. 1976;294:1442–4.

    Article  CAS  Google Scholar 

  60. Silverstein E, Pertschuk LP, Friedland J. Immunofluorescent detection of angiotensin-converting enzyme (ACE) in Gaucher cells. Am J Med. 1980;69:408–10.

    Article  CAS  Google Scholar 

  61. Cabrera-Salazar MA, O’Rourke E, Henderson N, Wessel H, Barranger JA. Correlation of surrogate markers of Gaucher disease. Implications for long-term follow up of enzyme replacement therapy. Clin Chim Acta. 2004;344:101–7.

    Article  CAS  Google Scholar 

  62. Aerts JMF, Hollak CEM. Plasma and metabolic abnormalities in Gaucher’s disease. Baillieres Clin Haematol. 1997;10:691–709.

    Article  CAS  Google Scholar 

  63. Šumarac Z, Suvajdžì N, Ignjatoviz̀ S, Majkí-Singh N, Janí D, Petakov M, et al. Biomarkers in Serbian patients with Gaucher disease. Clin Biochem. 2011;44:950–4.

  64. Danilov SM, Tikhomirova VE, Metzger R, Naperova IA, Bukina TM, Goker-Alpan O, et al. ACE phenotyping in Gaucher disease. Mol Genet Metab. 2018;123:501–10.

    Article  CAS  Google Scholar 

  65. van der Lienden MJC, Gaspar P, Boot R, Aerts JMFG, van Eijk M. Glycoprotein non-metastatic protein B: an emerging biomarker for lysosomal dysfunction in macrophages. Int J Mol Sci. 2018;20:66.

    Article  Google Scholar 

  66. Beneteau-Burnat B, Baudin B. Angiotensin-converting enzyme: Clinical applications and laboratory investigations on serum and other biological fluids. Crit Rev Clin Lab Sci. 1991;28:337–56.

    Article  CAS  Google Scholar 

  67. Danilov SM, Jain MS, Petukhov PA, Goldman C, DiSanto-Rose M, Vancavage R, et al. Novel ACE mutations mimicking sarcoidosis by increasing blood ACE levels. Transl Res. 2021;230:5–20.

    Article  CAS  Google Scholar 

  68. Nesterovitch AB, Hogarth KD, Adarichev VA, Vinokour EI, Schwartz DE, Solway J, et al. Angiotensin I-converting enzyme mutation (Trp1197Stop) causes a dramatic increase in blood ACE. PLoS ONE. 2009;4.

  69. Struthers AD, MacFadyen R, Fraser C, Robson J, Morton JJ, Junot C, et al. Nonadherence with angiotensin-converting enzyme inhibitor therapy: a comparison of different ways of measuring it in patients with chronic heart failure. J Am Coll Cardiol. 1999;34:2072–7.

    Article  CAS  Google Scholar 

  70. Tuchman LR, Suna H, Carr JJ. Elevation of serum acid phosphatase in Gaucher’s disease. J Mt Sinai Hosp N Y. 1956;23:227–9.

    CAS  Google Scholar 

  71. Lam WKW, Ted D, Li C-Y, Yam LT. Biochemical properties of tartrate-resistant acid phosphatasein serum of adultsand children. Clin Chem. 1978;24:1105.

    Article  CAS  Google Scholar 

  72. Kramer G, Wegdam W, Donker-Koopman W, Ottenhoff R, Gaspar P, Verhoek M, et al. Elevation of glycoprotein nonmetastatic melanoma protein B in type 1 Gaucher disease patients and mouse models. FEBS Open Bio. 2016;6:902–13.

    Article  CAS  Google Scholar 

  73. Murugesan V, Liu J, Yang R, Lin H, Lischuk A, Pastores G, et al. Validating glycoprotein non-metastatic melanoma B (gpNMB, osteoactivin), a new biomarker of Gaucher disease. Blood Cells Mol Dis. 2018;68:47–53.

    Article  CAS  Google Scholar 

  74. Saville JT, McDermott BK, Chin SJ, Fletcher JM, Fuller M. Expanding the clinical utility of glucosylsphingosine for Gaucher disease. J Inherit Metab Dis. 2020;43:558–63.

    Article  CAS  Google Scholar 

  75. Pacheco N, Uribe A. Enzymatic analysis of biomarkers for the monitoring of Gaucher patients in Colombia. Gene. 2013;521:129–35.

    Article  CAS  Google Scholar 

  76. Rodrigues MDB, de Oliveira AC, Müller KB, Martins AM, D’Almeida V. Chitotriosidase determination in plasma and in dried blood spots: a comparison using two different substrates in a microplate assay. Clin Chim Acta. 2009;406:86–8.

    Article  CAS  Google Scholar 

  77. Chaves RG, Coelho JC, Michelin-Tirelli K, Michelin-Tirelli K, Freitas Maurıcio T, de Freitas Maia Chaves E, et al. Successful screening for Gaucher disease in a high-prevalence population in tabuleirodo norte (Northeastern Brazil): A cross-sectional study. J Inherit Metab Dis. 2011;1:73–8.

  78. Aguilera B, Ghauharali-van der Vlugt K, Helmond MTJ, Out JMM, Donker-Koopman WE, Groener JEM, et al. Transglycosidase Activity of Chitotriosidase: improved enzymatic assay for the human macrophage chitinase. J Biol Chem. 2003;278:40911–6.

  79. Schoonhoven A, Rudensky B, Elstein D, Zimran A, Hollak CEM, Groener JE, et al. Monitoring of Gaucher patients with a novel chitotriosidase assay. Clin Chim Acta. 2007;381:136–9.

    Article  CAS  Google Scholar 

  80. van Breemen MJ, Bleijlevens B, de Koster CG, Aerts JMFG. Limitations in quantitation of the biomarker CCL18 in Gaucher disease blood samples by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry. Biochim Biophys Acta Proteins Proteom. 2006;1764:1626–32.

    Article  Google Scholar 

  81. Polo G, Burlina AP, Ranieri E, Colucci F, Rubert L, Pascarella A, et al. Plasma and dried blood spot lysosphingolipids for the diagnosis of different sphingolipidoses: a comparative study. Clin Chem Lab Med. 2019;57:1863–74.

    Article  CAS  Google Scholar 

  82. Dinur T, Bauer P, Beetz C, Kramp G, Cozma C, Iurașcu M-I, et al. Gaucher disease diagnosis using Lyso-Gb1 on dry blood spot samples: time to change the paradigm? Int J Mol Sci. 2022;23:1627.

    Article  CAS  Google Scholar 

  83. Brady RO, Kanfer J, Shapiro D. The metabolism of glucocerebrosides. I purification and properties of a glucocerebroside-cleaving enzyme from spleen tissue. J Biol Chem. 1965;240:39–43.

  84. Beutler E. Gaucher disease: new molecular approaches to diagnosis and treatment. Science. 1992;256:794–9.

    Article  CAS  Google Scholar 

  85. Patrick AD. A deficiency of Glucocerebrosidase in Gaucher’s disease. Biochem J. 1965;97:17C-24C.

    Article  CAS  Google Scholar 

  86. Bodamer OA, Hung C. Laboratory and genetic evaluation of Gaucher disease. Wien Med Wochenschr. 2010;160:600–4.

    Article  Google Scholar 

  87. Hannon HW. Blood collection on filter paper for Newborn screening programs; approved standard. 6th ed. Wayne: Clinical and Laboratory Standards Institute; 2013.

    Google Scholar 

  88. Gasparotto N, Tomanin R, Frigo AC, Niizawa G, Pasquini E, Blanco M, et al. Rapid diagnostic testing procedures for lysosomal storage disorders: alpha-glucosidase and beta-galactosidase assays on dried blood spots. Clin Chim Acta. 2009;402:38–41.

    Article  CAS  Google Scholar 

  89. Adam BW, Hall EM, Sternberg M, Lim TH, Flores SR, O’Brien S, et al. The stability of markers in dried-blood spots for recommended newborn screening disorders in the United States. Clin Biochem. 2011;44:1445–50.

    Article  CAS  Google Scholar 

  90. Elbin CS, Olivova P, Marashio CA, Cooper SK, Cullen E, Keutzer JM, et al. The effect of preparation, storage and shipping of dried blood spots on the activity of five lysosomal enzymes. Clin Chim Acta. 2011;412:1207–12.

    Article  CAS  Google Scholar 

  91. Reuser AJ, Verheijen FW, Bali D, van Diggelen OP, Germain DP, Hwu WL, et al. The use of dried blood spot samples in the diagnosis of lysosomal storage disorders—current status and perspectives. Mol Genet Metab. 2011;104:144–8.

    Article  CAS  Google Scholar 

  92. Ceci R, de Francesco PN, Mucci JM, Cancelarich LN, Fossati CA, Rozenfeld PA. Reliability of enzyme assays in dried blood spots for diagnosis of 4 lysosomal storage disorders. Adv Biol Chem. 2011;01:58–64.

    Article  CAS  Google Scholar 

  93. Chamoles NA, Blanco M, Gaggioli D, Casentini C. Gaucher and Niemann-Pick diseases—enzymatic diagnosis in dried blood spots on filter paper: retrospective diagnoses in newborn-screening cards. Clin Chim Acta. 2002;317:191–7.

    Article  CAS  Google Scholar 

  94. Moat SJ, George RS, Carling RS. Use of dried blood spot specimens to monitor patients with inherited metabolic disorders. Int J Neonatal Screen. 2020;26(6):26.

    Article  Google Scholar 

  95. Lukacs Z, Keil A, Peters V, Kohlschütter A, Hoffmann GF, Cantz M, et al. Towards quality assurance in the determination of lysosomal enzymes: a two-centre study. J Inherit Metab Dis. 2003;26:571–81.

    Article  CAS  Google Scholar 

  96. Sanders KA, Gavrilov DK, Oglesbee D, Raymond KM, Tortorelli S, Hopwood JJ, et al. A comparative effectiveness study of newborn screening methods for four lysosomal storage disorders. Int J Neonatal Screen. 2020;6:44.

    Article  Google Scholar 

  97. Bender F, Burin MG, Tirelli KM, Medeiros F, de Bitencourt FH, Civallero G, et al. Newborn screening for lysosomal disorders in Brazil: a pilot study using customized fluorimetric assays. Genet Mol Biol. 2020;43:e20180334.

    Article  CAS  Google Scholar 

  98. Huang Y, Jia X, Tang C, Liu S, Sheng H, Zhao X, et al. High risk screening for Gaucher disease in patients with splenomegaly and/or thrombocytopenia in China: 55 cases identified. Clin Chim Acta. 2020;506:22–7.

    Article  CAS  Google Scholar 

  99. Burlina AB, Polo G, Rubert L, Gueraldi D, Cazzorla C, Duro G, et al. Implementation of second-tier tests in newborn screening for lysosomal disorders in North Eastern Italy. Int J Neonatal Screen. 2019;5:24.

    Article  Google Scholar 

  100. Wasserstein MP, Caggana M, Bailey SM, Desnick RJ, Edelmann L, Estrella L, et al. The New York pilot newborn screening program for lysosomal storage diseases: report of the First 65,000 Infants. Genet Med. 2019;21:631–40.

    Article  CAS  Google Scholar 

  101. Kang L, Zhan X, Gu X, Zhang H. Successful newborn screening for Gaucher disease using fluorometric assay in China. J Hum Genet. 2017;62:763–8.

    Article  CAS  Google Scholar 

  102. Tortorelli S, Turgeon CT, Gavrilov DK, Oglesbee D, Raymond KM, Rinaldo P, et al. Simultaneous testing for 6 lysosomal storage disorders and x-adrenoleukodystrophy in dried blood spots by tandem mass spectrometry. Clin Chem. 2016;62:1248–54.

    Article  CAS  Google Scholar 

  103. Elliott S, Buroker N, Cournoyer JJ, Potier AM, Trometer JD, Elbin C, et al. Pilot study of newborn screening for six lysosomal storage diseases using Tandem Mass Spectrometry. Mol Genet Metab. 2016;118:304–9.

    Article  CAS  Google Scholar 

  104. Stroppiano M, Calevo MG, Corsolini F, Cassanello M, Cassinerio E, Lanza F, et al. Validity of β-d-glucosidase activity measured in dried blood samples for detection of potential Gaucher disease patients. Clin Biochem. 2014;47:1293–6.

    Article  CAS  Google Scholar 

  105. Kolodny EH, Mumford RA. Human leukocyte acid hydrolases: Characterization of eleven lysosomal enzymes and study of reaction conditions for their automated analysis. Clin Chim Acta. 1976;70:247–57.

    Article  CAS  Google Scholar 

  106. Roos D, Loos JA. Changes in the carbohydrate metabolism of mitogenicellay stimulated human peripheral lymphocytes I. Stimulation by phytohaemagglutinin. Biochim Biophys Acta Gen Subj. 1970;222:565–82.

    Article  CAS  Google Scholar 

  107. Peters SP, Lee RE, Glew RH. A microassay for Gaucher’s disease. Clin Chim Acta. 1975;60:391–6.

    Article  CAS  Google Scholar 

  108. Karatas M, Dogan S, Spahiu E, Ašić A, Bešić L, Turan Y. Enzyme kinetics and inhibition parameters of human leukocyte glucosylceramidase. Heliyon. 2020;6.

  109. Skoog WA, Beck WS. Studies on the fibrinogen, dextran and phytohemagglutinin methods of isolating leukocytes. Blood. 1956;11:436–54.

    Article  CAS  Google Scholar 

  110. Coelho JC, Giugliani R. Fibroblasts of skin fragments as a tool for the investigation of genetic diseases: technical recommendations. Genet Mol Biol. 2000;23:269–71.

    Article  Google Scholar 

  111. Bradford M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–54.

    Article  CAS  Google Scholar 

  112. Wenger DA, Clark C, Sattler M, Wharton C. Synthetic substrate beta-glucosidase activity in leukocytes: a reproducible method for the identification of patients and carriers of Gaucher’s disease. Clin Genet. 1978;13:145–53.

    Article  CAS  Google Scholar 

  113. Olivova P, Cullen E, Titlow M, Kallwass H, Barranger J, Zhang K, et al. An improved high-throughput dried blood spot screening method for Gaucher disease. Clin Chim Acta. 2008;398:163–4.

    Article  CAS  Google Scholar 

  114. Chiao YB, Glew RH, Driven W, Lee RE. Comparison of various β-glucosidase assays used to diagnose Gaucher’s disease. Clin Chim Acta. 1980;105:41–50.

    Article  CAS  Google Scholar 

  115. Daniels LB, Glew RH, Diven WF, Lee RE, Radin NS. An improved fluorometric leukocyte β-glucosidase assay for Gaucher’s disease. Clin Chim Acta. 1981;115:369–75.

    Article  CAS  Google Scholar 

  116. Shapira E, Blitzer MG, Africk DK, Miller JB. Biochemical genetics: a laboratory manual. 1st ed. Oxford: Oxford University Press; 1989.

    Google Scholar 

  117. Magalhães J, SáMiranda MC, Pinto R, Lemos M, Poenaru L. Sodium taurocholate effect on β-glucosidase activity: a new approach for identification of Gaucher disease using the synthetic substrate and leucocytes. Clin Chim Acta. 1984;141:111–8.

    Article  Google Scholar 

  118. Michelin K, Wajner A, Goulart LDS, Fachel ÂA, Pereira MLS, de Mello AS, et al. Biochemical study on β-glucosidase in individuals with Gaucher’s disease and normal subjects. Clin Chim Acta. 2004;343:145–53.

    Article  CAS  Google Scholar 

  119. Sista RS, Wang T, Wu N, Graham C, Eckhardt A, Bali D, et al. Rapid assays for Gaucher and Hurler diseases in dried blood spots using digital microfluidics. Mol Genet Metab. 2013;109:218–20.

    Article  CAS  Google Scholar 

  120. Sista RS, Wang T, Wu N, Graham C, Eckhardt A, Winger T, et al. Multiplex newborn screening for Pompe, Fabry, Hunter, Gaucher, and Hurler diseases using a digital microfluidic platform. Clin Chim Acta. 2013;424:12–8.

    Article  CAS  Google Scholar 

  121. Camargo Neto E, Schulte J, Pereira J, Bravo H, Sampaio-Filho C, Giugliani R. Neonatal screening for four lysosomal storage diseases with a digital microfluidics platform: Initial results in Brazil. Genet Mol Biol. 2018;41:414–6.

    Article  Google Scholar 

  122. Hopkins P v., Campbell C, Klug T, Rogers S, Raburn-Miller J, Kiesling J. Lysosomal storage disorder screening implementation: findings from the first six months of full population pilot testing in Missouri. J Pediatr 2015;166:172–7.

  123. Millington D, Norton S, Singh R, Sista R, Srinivasan V, Pamula V. Digital microfluidics comes of age: high-throughput screening to bedside diagnostic testing for genetic disorders in newborns. Expert Rev Mol Diagn. 2018;18:701–12.

    Article  CAS  Google Scholar 

  124. Li Y, Scott CR, Chamoles NA, Ghavami A, Pinto BM, Turecek F, et al. Direct multiplex assay of lysosomal enzymes in dried blood spots for newborn screening. Clin Chem. 2004;50:1785–96.

    Article  CAS  Google Scholar 

  125. Zhang XK, Elbin CS, Chuang WL, Cooper SK, Marashio CA, Beauregard C, et al. Multiplex enzyme assay screening of dried blood spots for lysosomal storage disorders by using tandem mass spectrometry. Clin Chem. 2008;54:1725–8.

    Article  CAS  Google Scholar 

  126. Gelb MH, Turecek F, Scott CR, Chamoles NA. Direct multiplex assay of enzymes in dried blood spots by tandem mass spectrometry for the newborn screening of lysosomal storage disorders. J Inherit Metab Dis. 2006;29:397–404.

    Article  CAS  Google Scholar 

  127. Wolf P, Alcalay RN, Liong C, Cullen E, Pauciulo MW, Nichols WC, et al. Tandem mass spectrometry assay of β-glucocerebrosidase activity in dried blood spots eliminates false positives detected in fluorescence assay. Mol Genet Metab. 2018;123:135–9.

    Article  CAS  Google Scholar 

  128. Orsini JJ, Martin MM, Showers AL, Bodamer OA, Zhang XK, Gelb MH, et al. Lysosomal storage disorder 4+1 multiplex assay for newborn screening using tandem mass spectrometry: application to a small-scale population study for five lysosomal storage disorders. Clin Chim Acta. 2012;413:1270–3.

    Article  CAS  Google Scholar 

  129. Stirnemann J, Belmatoug N, Camou F, Serratrice C, Froissart R, Caillaud C, et al. A review of Gaucher disease pathophysiology, clinical presentation and treatments. Int J Mol Sci. 2017;18:441.

    Article  Google Scholar 

  130. Baris HN, Cohen IJ, Mistry PK. Gaucher disease: The metabolic defect, pathophysiology, phenotypes and natural history. Pediatr Endocrinol Rev. 2014;12:72–81.

    Google Scholar 

  131. Fateen E, Abdallah ZY. Twenty- five years of biochemical diagnosis of Gaucher disease: the Egyptian experience. Heliyon. 2019;5:e02574.

    Article  Google Scholar 

  132. Butcher BA, Gopalan V, Lee RE, Richards TC, Waggoner AS, Glew RH. Use of 4-heptylumbelliferyl-β-d-glucoside to identify Gaucher’s disease heterozygotes. Clin Chim Acta. 1989;184:235–42.

    Article  CAS  Google Scholar 

  133. Mistry P, Germain DP. Phenotype variations in Gaucher disease. Rev Med Int. 2006;27(Suppl 1):S3-10.

    Article  Google Scholar 

  134. Essabar L, Meskini T, Lamalmi N, Ettair S, Erreimi N, Mouane N. Gaucher’s disease: report of 11 cases with review of literature. Pan Afr Med J. 2015;20.

  135. Blau N, Duran M, Gibson KM. Laboratory guide to the methods in biochemical genetics. 1st ed. Heidelberg: Springer; 2008.

    Book  Google Scholar 

  136. Martins AM, Valadares ER, Porta G, Coelho J, Filho JS, Dudeque Pianovski MA, et al. Recommendations on Diagnosis, Treatment, and Monitoring for Gaucher Disease. J Pediatr. 2009;155.

  137. Verma J, Thomas DC, Sharma S, Jhingan G, Singh A, Hsiao KJ, et al. Inherited metabolic disorders: quality management for laboratory diagnosis. Clin Chim Acta. 2015;447:1–7.

    Article  CAS  Google Scholar 

  138. Erndim. http://cms.erndimqa.nl/Home/Lysosomal-Enzymes.aspx. Accessed 18 January 2022.

  139. Yu C, Sun Q, Zhou H. Enzymatic screening and diagnosis of lysosomal storage diseases. N Am J Med Sci (Boston). 2013;06:186.

    Article  Google Scholar 

  140. Fowler B, Burlina A, Kozich V, Vianey-Saban C. Quality of analytical performance in inherited metabolic disorders: the role of ERNDIM. J Inherit Metab Dis. 2008;31:680–9.

    Article  CAS  Google Scholar 

  141. Hong CM, Ohashi T, Yu XJ, Weiler S, Barranger JA. Sequence of two alleles responsible for Gaucher disease. DNA Cell Biol. 1990;9:233–41.

    Article  CAS  Google Scholar 

  142. Latham TE, Theophilus BDM, Grabowski GA, Smith FI. Heterogeneity of mutations in the acid β-glucosidase gene of Gaucher disease patients. DNA Cell Biol. 1991;10:15–21.

    Article  CAS  Google Scholar 

  143. Filocamo M, Mazzotti R, Stroppiano M, Seri M, Giona F, Parenti G, et al. Analysis of the glucocerebrosidase gene and mutation profile in 144 Italian gaucher patients. Hum Mutat. 2002;20:234–5.

    Article  Google Scholar 

  144. Koprivica V, Stone DL, Park JK, Callahan M, Frisch A, Cohen IJ, et al. Analysis and classification of 304 mutant alleles in patients with type 1 anti type 3 gaucher disease. Am J Hum Genet. 2000;66:1777–86.

    Article  CAS  Google Scholar 

  145. Alfonso P, Cenarro A, Pérez-Calvo JI, Giralt M, Giraldo P, Pocoví M. Mutation prevalence among 51 unrelated Spanish patients with Gaucher disease: identification of 11 novel mutations. Blood Cells Mol Dis. 2001;27:882–91.

    Article  CAS  Google Scholar 

  146. Miocić S, Filocamo M, Dominissini S, Montalvo ALE, Vlahovicek K, Deganuto M, et al. Identification and functional characterization of five novel mutant alleles in 58 Italian patients with Gaucher disease type 1. Hum Mutat. 2005;25:100.

    Article  Google Scholar 

  147. Erdos M, Hodanova K, Taskó S, Palicz A, Stolnaja L, Dvorakova L, et al. Genetic and clinical features of patients with Gaucher disease in Hungary. Blood Cells Mol Dis. 2007;39:119–23.

    Article  CAS  Google Scholar 

  148. Emre S, Gürakan F, Yüce A, Rolf A, Scott R, Özen H. Molecular analysis of Turkish Gaucher disease patients: identification of novel mutations in glucocerebrosidase (GBA) gene. Eur J Med Genet. 2008;51:315–21.

    Article  Google Scholar 

  149. Mattošová S, Chandoga J, Hlavatá A, Šaligová J, Maceková D. Spectrum of GBA mutations in patients with gaucher disease from Slovakia: identification of five novel mutations. Isr Med Assoc J. 2015;17:166–70.

    Google Scholar 

  150. Jeong SY, Park SJ, Kim HJ. Clinical and genetic characteristics of Korean patients with Gaucher disease. Blood Cells Mol Dis. 2011;46:11–4.

    Article  CAS  Google Scholar 

  151. Ortiz-Cabrera N v., Gallego-Merlo J, Vélez-Monsalve C, de Nicolas R, Mas SF, Ayuso C, et al. Nine-year experience in Gaucher disease diagnosis at the Spanish reference center Fundación Jiménez Díaz. Mol Genet Metab Rep. 2016;9:79–85.

  152. Feng Y, Huang Y, Tang C, Hu H, Zhao X, Sheng H, et al. Clinical and molecular characteristics of patients with Gaucher disease in Southern China. Blood Cells Mol Dis. 2018;68:30–4.

    Article  CAS  Google Scholar 

  153. Sheth J, Bhavsar R, Mistri M, Pancholi D, Bavdekar A, Dalal A, et al. Gaucher disease: Single gene molecular characterization of one-hundred Indian patients reveals novel variants and the most prevalent mutation. BMC Med Genet. 2019;20.

  154. Lepe-Balsalobre E, Santotoribio JD, Nuñez-Vazquez R, García-Morillo S, Jiménez-Arriscado P, Hernández-Arévalo P, et al. Genotype/phenotype relationship in Gaucher disease patients. Novel mutation in glucocerebrosidase gene. Clin Chem Lab Med. 2020;58:2017–24.

  155. Duran R, McNeill A, Mehta A, Hughes D, Cox T, Deegan P, et al. Novel pathogenic mutations in the glucocerebrosidase locus. Mol Genet Metab. 2012;106:495–7.

    Article  CAS  Google Scholar 

  156. Hruska KS, LaMarca ME, Scott CR, Sidransky E. Gaucher disease: Mutation and polymorphism spectrum in the glucocerebrosidase gene (GBA). Hum Mutat. 2008;29:567–83.

    Article  CAS  Google Scholar 

  157. Giraldo P, Pocoví M, Pérez-Calvo J, Rubio-Félix D, Giralt M. Report of the Spanish Gaucher’s disease registry: clinical and genetic characteristics. Haematologica. 2000;85:792–9.

    CAS  Google Scholar 

  158. Cherif W, ben Turkia H, ben Rhouma F, Riahi I, Chemli J, Kefi R, et al. Gaucher disease in Tunisia: High frequency of the most common mutations. Blood Cells Mol Dis. 2009;43:161–2.

  159. Bronstein S, Karpati M, Peleg L. An update of Gaucher mutations distribution in the Ashkenazi Jewish population: prevalence and country of origin of the Mutation R496H. Isr Med Assoc J. 2014;16:683–5.

    Google Scholar 

  160. Orenstein M, Barbouth D, Bodamer OA, Weinreb NJ. Patients with type 1 Gaucher disease in South Florida, USA: Demographics, genotypes, disease severity and treatment outcomes. Orphanet J Rare Dis. 2014;9.

  161. Grabowski GA, Zimran A, Ida H. Gaucher disease types 1 and 3: Phenotypic characterization of large populations from the ICGG Gaucher Registry. Am J Hematol. 2015;90(Suppl 1):S12–8.

    Article  Google Scholar 

  162. Horowitz M, Tzuri G, Eyal N, Berebi A, Kolodny EH, Brady RO, et al. Prevalence of nine mutations among Jewish and non-Jewish Gaucher disease patients. American J Hum Genet. 1993;53:930.

    Google Scholar 

  163. Beutler E, Gelbart T. Gaucher disease mutations in non-Jewish patients. Br J Haematol. 1993;85:401–5.

    Article  CAS  Google Scholar 

  164. Hatton CE, Cooper A, Whitehouse C, Wraith JE. Mutation analysis in 46 British and Irish patients with Gaucher’s disease. Arch Dis Child. 1997;77:17–22.

    Article  CAS  Google Scholar 

  165. Beutler E, Gelbart T. Erroneous assignment of Gaucher disease genotype as a consequence of a complete gene deletion. Hum Mutat. 1994;4:212–6.

    Article  CAS  Google Scholar 

  166. Cozar M, Bembi B, Dominissini S, Zampieri S, Vilageliu L, Grinberg D, et al. Molecular characterization of a new deletion of the GBA1 gene due to an inter Alu recombination event. Mol Genet Metab. 2011;102:226–8.

    Article  CAS  Google Scholar 

  167. Zampieri S, Cattarossi S, Bembi B, Dardis A. GBA analysis in next-generation era: pitfalls, challenges, and possible solutions. J Mol Diagn. 2017;19:733–41.

    Article  CAS  Google Scholar 

  168. den Heijer JM, Cullen VC, Quadri M, Schmitz A, Hilt DC, Lansbury P, et al. A large-scale full GBA1 gene screening in Parkinson’s disease in the Netherlands. Mov Disord. 2020;35:1667–74.

    Article  Google Scholar 

  169. den Heijer JM, Schmitz A, Lansbury P, Cullen VC, Hilt DC, Bonifati V, et al. False negatives in GBA1 sequencing due to polymerase dependent allelic imbalance. Sci Rep. 2021;11.

  170. Málaga DR, Brusius-Facchin AC, Siebert M, Pasqualim G, Pereira MLS, de Souza CFM, et al. Sensitivity, advantages, limitations, and clinical utility of targeted next-generation sequencing panels for the diagnosis of selected lysosomal storage disorders. Genet Mol Biol. 2019;42:197–206.

    Article  Google Scholar 

  171. Lee CY, Yen HY, Zhong AW, Gao H. Resolving misalignment interference for NGS-based clinical diagnostics. Hum Genet. 2021;140:477–92.

    Article  Google Scholar 

  172. Muñoz G, García-Seisdedos D, Ciubotariu C, Piris-Villaespesa M, Gandía M, Martín-Moro F, et al. Early detection of lysosomal diseases by screening of cases of idiopathic splenomegaly and/or thrombocytopenia with a next-generation sequencing gene panel. JIMD Rep. 2020;51:53–61.

    Article  Google Scholar 

  173. Zanetti A, D’Avanzo F, Bertoldi L, Zampieri G, Feltrin E, de Pascale F, et al. Setup and validation of a targeted next-generation sequencing approach for the diagnosis of lysosomal storage disorders. J Mol Diagn. 2020;22:488–502.

    Article  CAS  Google Scholar 

  174. Zampieri S, Cattarossi S, Pavan E, Barbato A, Fiumara A, Peruzzo P, et al. Accurate molecular diagnosis of Gaucher disease using clinical exome sequencing as a first-tier test. Int J Mol Sci. 2021;22:5538.

    Article  CAS  Google Scholar 

  175. Spataro N, Roca-Umbert A, Cervera-Carles L, Vallès M, Anglada R, Pagonabarraga J, et al. Detection of genomic rearrangements from targeted resequencing data in Parkinson’s disease patients. Mov Disord. 2017;32:165–9.

    Article  CAS  Google Scholar 

  176. Drelichman G, Fernández Escobar N, Soberon B, Basack N, Frabasil J, Schenone A, et al. Long-read single molecule real-time (SMRT) sequencing of GBA1 locus in Gaucher disease national cohort from Argentina reveals high frequency of complex allele underlying severe skeletal phenotypes: Collaborative study from the Argentine Group for Diagnosis and Treatment of Gaucher Disease. Mol Genet Metab Rep. 2021;29.

  177. Amico G, Grossi S, Vijzelaar R, Lanza F, Mazzotti R, Corsolini F, et al. MLPA-based approach for initial and simultaneous detection of GBA deletions and recombinant alleles in patients affected by Gaucher Disease. Mol Genet Metab. 2016;119:329–37.

    Article  CAS  Google Scholar 

  178. Schnabel D, Schröder M, Sandhoff K. Mutation in the sphingolipid activator protein 2 in a patient with a variant of Gaucher disease. FEBS Lett. 1991;284:57–9.

    Article  CAS  Google Scholar 

  179. Rafi MA, de Gala G, Zhang X ling, Wenger DA. Mutational analysis in a patient with a variant form of Gaucher disease caused by SAP-2 deficiency. Somat Cell Mol Genet. 1993;19:1–7.

  180. Christomanou H, Chabás A, Pámpols T, Guardiola A. Activator protein deficient Gaucher’s disease. A second patient with the newly identified lipid storage disorder. Klin Wochenschr. 1989;67:999–1003.

  181. Vaccaro AM, Motta M, Tatti M, Scarpa S, Masuelli L, Bhat M, et al. Saposin C mutations in Gaucher disease patients resulting in lysosomal lipid accumulation, saposin C deficiency, but normal prosaposin processing and sorting. Hum Mol Genet. 2010;19:2987–97.

    Article  CAS  Google Scholar 

  182. Kang L, Zhan X, Ye J, Han L, Qiu W, Gu X, et al. A rare form of Gaucher disease resulting from saposin C deficiency. Blood Cells Mol Dis. 2018;68:60–5.

    Article  CAS  Google Scholar 

  183. Tylki-Szymańska A, Czartoryska B, Vanier MT, Poorthuis BJMH, Groener JAE, Ługowska A, et al. Non-neuronopathic Gaucher disease due to saposin C deficiency. Clin Genet. 2007;72:538–42.

    Article  Google Scholar 

  184. Hiraiwa M, Martin BM, Kishimoto Y, Conner GE, Tsuji S, O’Brien JS. Lysosomal proteolysis of prosaposin, the precursor of saposins (sphingolipid activator proteins): its mechanism and inhibition by ganglioside. Arch Biochem Biophys. 1997;341:17–24.

    Article  CAS  Google Scholar 

  185. Vielhaber G, Hurwitz R, Sandhoff K. Biosynthesis, processing, and targeting of sphingolipid activator protein (SAP) precursor in cultured human fibroblasts. Mannose 6-phosphate receptor-independent endocytosis of SAP precursor. J Biol Chem. 1996;271:32438–46.

    Article  CAS  Google Scholar 

  186. Motta M, Camerini S, Tatti M, Casella M, Torreri P, Crescenzi M, et al. Gaucher disease due to saposin C deficiency is an inherited lysosomal disease caused by rapidly degraded mutant proteins. Hum Mol Genet. 2014;23:5814–26.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors would like to thank all members of the IWGGD for their constructive discussion and suggestions.

Funding

Not applicable.

Author information

Authors and Affiliations

  1. Regional Coordinator Centre for Rare Disease, University Hospital of Udine, P.Le Santa Maria Della Misericordia 15, 33100, Udine, Italy

    A. Dardis & E. Pavan

  2. Department of Enzymology and Cellular Function, Institute of Child Health, Athens, Greece

    H. Michelakakis

  3. Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Instituto de Estudios Inmunológicos Y Fisiopatológicos (IIFP), UNLP, CONICET, Asociado CIC PBA, La Plata, Argentina

    P. Rozenfeld

  4. Department for Laboratory Diagnostics, University Hospital Centre Zagreb and School of Medicine, Zagreb, Croatia

    K. Fumic

  5. Department of Medical Biology and Genetics, Faculty of Medicine, J.J. Strossmayer University, Osijek, Croatia

    J. Wagner

  6. International Gaucher Alliance, Dursley, UK

    J. Wagner

  7. Genetics and Molecular Pathology, SA Pathology at Women’s and Children’s Hospital and Adelaide Medical School, University of Adelaide, Adelaide, SA, 5005, Australia

    M. Fuller

  8. Gaucher Unit, Shaare Zedek Medical Center, Jerusalem, Israel

    S. Revel-Vilk

  9. Faculty of Medicine, Hebrew University, Jerusalem, Israel

    S. Revel-Vilk

  10. Lysosomal Storage Disorders Unit, Royal Free London NHS Foundation Trust and University College London, London, UK

    D. Hughes

  11. Department of Medicine, University of Cambridge, Cambridge, UK

    T. Cox

  12. Department of Medical Biochemistry, Leiden Institute of Chemistry, Leiden, The Netherlands

    J. Aerts

Authors
  1. A. Dardis
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. H. Michelakakis
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. P. Rozenfeld
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. K. Fumic
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. J. Wagner
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. E. Pavan
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. M. Fuller
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. S. Revel-Vilk
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. D. Hughes
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. T. Cox
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. J. Aerts
    View author publications

    You can also search for this author in PubMed Google Scholar

Consortia

the International Working Group of Gaucher Disease (IWGGD)

Contributions

AD, HM, PR, KF, JW, JA, EP: contributed to the guidelines development by planning and drafting the manuscript; SR-V, MF, DH, TC: critically revised the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to A. Dardis.

Ethics declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

AD received consulting fees, speaker honoraria and research grants from Amicus, Takeda and Sanofi. PR have received consulting and research grants from Takeda, Biomarin and Amicus. EP received a doctoral fellowship from Sanofi. MF received research and travel funds from Abeona Therapeutics, Paradigm Biopharmaceuticals, Sanofi, Takeda and Taysha Gene Therapies. SR-V received research support/honoraria from Sanofi, Takeda and Pifzer. DH performed consultancy work and speaking engagements through UCL consultants for Takeda, Sanofi and Freeline. TC advises AvroBio, Sanofi, Takeda and receives honoraria for ABPI approved lectures. University of Cambridge has received research funds from Takeda and Sanofi to support investigator-led studies in Gaucher disease. Cambridge University hospitals receives fees from Sanofi for the conduct of a clinical trial in Gaucher disease.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dardis, A., Michelakakis, H., Rozenfeld, P. et al. Patient centered guidelines for the laboratory diagnosis of Gaucher disease type 1. Orphanet J Rare Dis 17, 442 (2022). https://doi.org/10.1186/s13023-022-02573-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1186/s13023-022-02573-6

Keywords

Orphanet Journal of Rare Diseases

ISSN: 1750-1172