Printer Friendly

Detection of mucopolysaccharidosis type II by measurement of iduronate-2-sulfatase in dried blood spots and plasma samples.

Mucopolysaccharidosis II (MPS II, (3) Hunter syndrome) is an X-linked, recessively inherited, lysosomal storage disorder (LSD) (1). MPS II is caused by accumulation of the glycosaminoglycans heparan sulfate and dermatan sulfate, which occurs as a result of a deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS; EC 3.1.6.13). Clinically, MPS II patients can present with variable symptoms, with disease categories ranging from severe to attenuated forms (1). Patients with a severe phenotype have pronounced neurologic impairment and often die in early childhood, whereas individuals with an attenuated phenotype may have little or no neurologic impairment and can survive into adulthood. Other symptoms include skeletal deformities, organomegaly, short stature, stiff joints, and coarse facial features (1). At least 309 IDS gene mutations have been cataloged in the Human Gene Mutation Database (http://www.hgmd.org) (2), and genetic heterogeneity is presumed to underlie most variations in clinical phenotype. However, genotype /phenotype studies have not provided the correlation's required to accurately predict disease severity and progression in all MPS II patients (3-5).

Bone marrow transplantation is a treatment option for some MPS II patients, but it is not a panacea, as relatively few patients at the severe end of the clinical spectrum have positive outcomes (6). Enzyme replacement therapy has been successfully used in humans to treat several other LSDs, including Gaucher disease (7), Fabry disease (8), MPS I (9), and MPS VI (10). The correction of glycosaminoglycan storage by enzyme replacement therapy in a mouse model of MPS II (11) was followed by human clinical trials, which are ongoing. Because these treatments will probably be most successful if given early, before the onset of irreversible pathology, an effective method for the identification of MPS II patients is needed. Protein markers have already been effective for the detection of individuals with other LSDs (12-15). Assays of IDS activity (16-18) and a direct assay for dried blood spots (19) have been described, but a sensitive assay to quantify IDS protein from a 3-mm dried blood spot has not been reported. Because IDS does not efficiently hydrolyze the fluorescent substrate 4-methylumbelliferyl sulfate (4MU sulfate) under uncaptured assay conditions, existing assays rely on specialized or radiolabeled substrates (17), complex substrate /product detection protocols (19), or additional enzyme-linked systems (16,18). An assay using the readily available 4MU sulfate would facilitate the early identification of MPS II patients. We describe an immunoassay for IDS protein and a solid-phase fluorometric assay that uses a polyclonal antibody to capture IDS and 4MU sulfate to detect enzyme activity, and we report on the diagnostic accuracy of these assays.

Materials and Methods

Recombinant human IDS was isolated from Chinese hamster ovary expression cells and purified by DEAE ion-exchange and phenyl-Sepharose chromatography to yield IDS, with characteristics similar to those described previously (20). Purified recombinant IDS was kept at 4[degrees]C before use. A sheep anti-IDS polyclonal antibody was produced against this IDS and purified from serum by protein G chromatography and affinity chromatography with recombinant IDS protein, as described previously (5). The affinity-purified antibody was adsorbed with sheep IgG coupled to a 1-mL Hitrap N-hydroxysuccinimide-activated high-performance affinity column (Amersham Pharmacia Biotech; 2 mg of sheep IgG on a 1-mL column), prepared according to the manufacturer's instructions. Purified antibody was stored frozen at -20[degrees]C before use. The sheep polyclonal was labeled with europium as described previously (21) and stored at 4[degrees]C. Recombinant human N-acetylgalactosamine-4-sulfatase was expressed in Chinese hamster ovary cells (22), purified by monoclonal antibody affinity chromatography (13), and stored at 4[degrees]C before use. Unless stated otherwise, all other reagents were of analytical grade and were obtained from Sigma Chemical Company.

PATIENT SAMPLES

The Human Research Ethics Committee of the Children, Youth and Women's Health Service, Adelaide, Australia, approved the use of dried-blood-spot and plasma samples in this study. Retrospective dried-blood-spot (3-mm diameter) and plasma samples from unaffected consecutive control individuals (age range, 0.5-50.5 years) and from MPS II-affected individuals (age range, 0.6-9.7 years), in whom MPS II was diagnosed enzymatically with a radiolabeled oligosaccharide substrate (23), were from specimens submitted to the National Referral Laboratory for the Diagnosis of Lysosomal, Peroxisomal and Related Genetic Disorders (Department of Genetic Medicine, Children, Youth and Women's Health Service, Adelaide). Plasma samples were stored at -20[degrees]C before use. Blood samples were also collected, with informed consent, from unaffected human controls (age range, 22.8-55.7 years) within the Department of Genetic Medicine. These latter samples were collected to evaluate several potential diagnostic assays for LSDs. Blood samples were collected into either EDTA or heparin. Dried-blood-spot samples were stored in sealed plastic bags inside plastic containers with desiccant at -20[degrees]C up to 4 years before use. Clinical follow-up data were not available for the MPS II or control individuals, and the disease diagnoses of sample donors were known to the persons performing IDS protein and enzyme activity assays.

MEASUREMENT OF IDS PROTEIN BY IMMUNOASSAY

We measured IDS protein in dried blood spots and plasma samples with a 2-step time-delayed, dissociationenhanced, lanthanide fluorescence immunoassay (DELFIA[R]). Microtiter plates (Immulon 4; DYNEX Technologies, Inc.) were coated with 100 [micro]L of sheep anti-IDS polyclonal antibody at a concentration of 5 mg/L in 0.1 mol/L NaHC[O.sub.3] (pH 8.3) and incubated, without shaking, overnight at 4[degrees]C. Plates were then washed 6 times (0.8 mL each time) with wash buffer [0.02 mol/L Tris-HCl, 0.25 mol/L NaCl, 0.05 mL/L Tween 20 (BDH), and 0.02 g/L Thiomerosal (pH 7.8)] in a DELFIA plate washer (1296-026; Perkin-Elmer Life and Analytical Sciences). Dried blood spots (duplicate samples) were placed in the coated microtiter wells with 100 [micro]L of assay buffer [0.05 mol/L Tris-HCl, 0.15 mol/L NaCl, 20 [micro]mol/L diethylenetriamine pentaacetic acid, 0.1 mL/L Tween 40, 5 g/L bovine serum albumin (BSA), 0.5 g/L bovine [gamma]-globulin, and 0.5 g/L sodium azide (pH 7.8)]. Plasma samples (8 [micro]L/well, assayed in duplicate) were diluted with assay buffer and added in a final volume of 100 [micro]L/well. Liquid calibrators of purified IDS were included on every assay plate. The plates were covered and incubated at room temperature for 1 h with shaking (amplitude 5; Milenia Micromix 4 plate shaker; Model 602002; DPC), then placed at 4[degrees]C overnight without shaking, before a 1-h incubation with shaking at room temperature. The blood spots were removed by aspiration, and the plates were washed (6 times) with wash buffer. Europium-labeled anti-IDS polyclonal antibody was diluted in assay buffer (100 [micro]L; final concentration, 200 [micro]g/L) and added to each well. The plates were incubated overnight as described above. After washing (6 times) with wash buffer, DELFIA enhancement solution (200 [micro]L/well; Wallac) was added to the plates and incubated at room temperature for 10 min with shaking. Fluorescence was measured on a DELFIA 1234 research fluorometer (Wallac). The IDS concentrations in the blood spots and plasma were calculated with spline fit curves generated by Multicalc Data Analysis software (Ver. 2.4; Wallac).

MEASUREMENT OF IDS ACTIVITY BY FLUOROGENIC IMMUNE-CAPTURE ASSAY

Microtiter plates were coated with affinity-purified sheep anti-IDS polyclonal antibody as described above, then washed (6 times) with 0.02 mol/L Tris-HCl (pH 7.0) containing 0.25 mol/L NaCl. Blood spots, plasma samples, and recombinant IDS calibrators were assayed in duplicate and diluted in 100 [micro]L of phosphate-buffered saline (10 mmol/L [Na.sub.2]HP[O.sub.4]/NaCH, 138 mmol/L NaCl, 2.7 mmol/L KCl) containing 0.5 mL/L Tween 20, 5 g/L BSA, and 0.5 g/L [gamma]-globulin. To demonstrate linearity, a 9-point calibration curve of recombinant human IDS (0-2000 pg/well) was run on every plate. Plates were covered and incubated at room temperature for 1 h with shaking, stored overnight at 4[degrees]C without shaking, and then incubated again at room temperature for 1 h with shaking. Blood spots were removed by aspiration, and the plates were washed (twice) with 0.02 mol/L Tris-HCl containing 0.25 mol/L NaCl (pH 7.0). Enzyme activity of the immune-captured IDS was determined by the addition of 5 mmol/L 4NN sulfate (stored at -20[degrees]C) in 0.1 mol/L sodium acetate buffer (pH 5.6) containing 10 g/L BSA to each well. Plates were sealed and shaken for 1 min and then incubated without shaking for 24 h at 37[degrees]C. Reactions were stopped by the addition of 100 [micro]L of 0.2 mol/L glycine, 0.125 mol/L [Na.sub.2]C[O.sub.3], and 0.16 mol/L NaOH (pH 10.7). Plates were shaken for an additional 10 min, and fluorescence was read on a Wallac Victor (2) 1420 Multilabel Counter (Perkin-Elmer Life and Analytical Sciences). With each analysis, a point calibration of 4-methylumbelliferone (710 pmol) was determined in duplicate. The enzyme activities of the samples were calculated by reference to the point calibration and corrected for incubation time (24 h) and sample volume (blood spots, 3 [micro]L; plasma samples, 8 [micro]L).

STATISTICAL ANALYSIS

Inter- and intraassay CVs were calculated for results from plasma and blood spots obtained from 2 different controls. Data were analyzed with SPSS (Ver. 11.0 for Windows) statistical software (SPSS Inc.).

Results

DEVELOPMENT OF IDS ASSAYS

The IDS protein and activity calibration curves were linear over the range 0-2000 pg/well (Fig. 1). We calculated the CV for the IDS protein assay by use of 10 replicates of each IDS calibrator (7.8 to 2000 pg/well), with 2-fold dilutions, and used a precision profile of CV against IDS concentration to determine the working range of the IDS protein assay (defined as protein concentrations with CVs <10%), which was 31.3 to 2000 pg/well. Samples with lower concentrations were deemed to have no detectable protein and assigned a zero concentration of IDS protein. The detection limit for IDS activity was defined by the concentration of activity required to give a signal 2 SD above the mean of the assay blank (n = 12). Any sample with activity below this value was deemed to have no detectable activity, and a zero degree of IDS activity was reported.

[FIGURE 1 OMITTED]

Intraassay CVs for the IDS protein assay, calculated from results for 2 different plasma samples with 20 replicates for each, were <6%. Intraassay CVs for the IDS protein assay, from results for 2 different blood-spot samples, were 9.6% and 9.2% (n = 20). Intraassay CVs for IDS activity, from 1 blood spot and 1 plasma sample, were 11% and 7.5%, respectively (n = 10). Interassay CVs (n = 6) for the IDS protein were 13% and 10% for 2 plasma samples and 18% and 16% for 2 blood-spot samples. Interassay CVs (n = 6) for IDS activity were 9.6% for a plasma sample and 8.6% for a blood-spot sample.

We determined the stability of IDS in plasma (n = 61) and blood spots (n = 64) stored at -20[degrees]C by assaying control samples stored for up to 4 years. No statistically significant correlations were seen between sample age and either IDS protein concentration or activity (data not shown). Only samples stored for <4 years were used in subsequent analyses for both the IDS protein concentration and activity assays. There was a significant correlation between IDS protein concentration and enzyme activity (Fig. 2) for both plasma [n = 38; Spearman correlation coefficient ([r.sub.s]) = 0.732; P <0.01, two-tailed] and blood-spot samples (n = 51; [r.sub.s] = 0.751; P <0.01, two-tailed).

The IDS enzyme did not hydrolyze the 4MU sulfate in an uncaptured activity assay (i.e., enzyme not captured on an antibody-coated microtiter well; data not shown). Similarly, the addition of various concentrations (1, 5, and 10 mg/L) of anti-IDS sheep polyclonal antibody to IDS protein in solution did not produce activity against 4MU sulfate (data not shown). There was negligible cross-reactivity with N-acetylgalactosamine-4-sulfatase in the immune-capture activity assay. N-Acetylgalactosamine-4-sulfatase (2000 pg/well) activity against the 4MU sulfate was only 3% that of IDS added at the same concentration (data not shown).

QUANTIFICATION OF IDS PROTEIN AND ACTIVITY IN CONTROL AND MPS II PLASMAS AND BLOOD SPOTS

We investigated the relationship between the age of the control individuals and the concentration of IDS protein or enzyme activity. IDS protein concentrations decreased significantly with age for both plasma (n = 38; [r.sub.s] = -0.578; P <0.01, two-tailed) and blood spots (n = 51; [r.sub.s] = -0.325; P <0.05, two-tailed). We also found a significant decrease with increasing age for IDS activity in both plasma samples (n = 59; [r.sub.s] = -0.453; P <0.01, two-tailed) and blood spots (n = 62; [r.sub.s] = -0.570; P <0.01, two-tailed), but the lower limits did not change for either activity or protein concentrations over the range 0.5-55.7 years of age (data not shown).

In plasma, MPS II patient samples were distinguishable from control samples for both IDS protein concentration (Fig. 3A) and enzyme activity (Fig. 3B). One of the controls had an extremely high plasma protein concentration and activity (349.4 [micro]g/L and 8.1 [micro]mol * [h.sup.-1] * [L.sup.-1], respectively); another control had low plasma IDS activity (0.168 [micro]mol * [h.sup.-1] * [L.sup.-1]), but this value was still 2-fold higher than that of the MPS II patient with the highest residual enzyme activity.

[FIGURE 2 OMITTED]

The control with high plasma IDS protein concentration and enzyme activity also had high values in blood spots (Fig. 4); however, no clinical data were available for this individual. In blood spots, 11 of 12 MPS II patients had no detectable protein (Fig. 4A). One MPS II patient blood spot did have an IDS protein concentration of 24.8 [micro]g/L, a value just below the lowest control value (25.8 [micro]g/L). No genotype or clinical data were available for the patient who provided this sample, which along with 11 other MPS II patient samples had negligible IDS activity in blood spots and was clearly distinguishable from control samples (Fig. 4B).

Discussion

Enzyme replacement therapy for the treatment of MPS II patients is likely to be available in the near future. Early diagnosis and treatment of asymptomatic individuals will be imperative to maximize the efficacy of this therapy. Current diagnostic techniques for MPS II involve the screening of urine to detect patterns of glycosaminoglycans and/or oligosaccharides, followed by a specific enzyme assay in either leukocyte or cultured skin fibroblast extracts (24). These methods are not amenable to high-throughput, large-scale screening programs. Thus, changing diagnostic requirements for this disease have prompted the development of the new assays reported here.

[FIGURE 3 OMITTED]

The method for quantification of IDS protein reported here was more sensitive than previously reported assays, with a working range of 31 to 2000 pg/well compared with a lower limit of detection of 1000 pg/well reported by Parkinson et al. (5) for nondenatured IDS and 5000 pg/well reported by Villani et al. (25). The current assay has the sensitivity to quantify IDS protein from a single 3-mm dried blood spot. Furthermore, only 8 [micro]L of plasma was required in the current assays compared with 50 [micro]L used previously (5). The large increases in assay sensitivity can be attributed to the use of the DELFIA detection technology instead of the less sensitive horseradish peroxidase detection methods (25) and to the use of a polyclonal antibody for both capture and detection steps.

A dried-blood-spot assay for IDS activity has been described previously (19), but this assay required the use of lead acetate to inhibit other lysosomal enzymes, a specific radiolabeled substrate, and an ion-exchange column step, making it impractical for large-scale screening applications. The novel fluorogenic immune-capture activity assay described here uses a commercially available 4MU sulfate substrate that could be measured directly in the reaction well, providing a simplified procedure amenable to high-throughput screening applications.

[FIGURE 4 OMITTED]

The immune assays for IDS protein and activity described here enabled the differentiation of all MPS II individuals from controls by use of either plasma or dried-blood-spot samples. No IDS protein was detected in any of the 9 MPS II plasma samples or in 11 of 12 MPS II blood-spot samples. One MPS II blood spot had detectable IDS protein, with a concentration just below the control range, although this sample had no IDS activity. This finding was consistent with previous reports of mutations such as K347T, 473delTCC, and N265I, which were shown to produce inactive IDS protein (26).

Of the sulfatases involved in LSDs, only arylsulfatase A (galactose-3-sulfatase) and arylsulfatase B (N-acetyl-galactosamine-4-sulfatase) efficiently hydrolyze 4MU sulfate. IDS, sulfamidase, N-acetylglucosamine-6-sulfatase, and galactose-6-sulfatase show little activity toward 4MU sulfate (27, 28). It appears that the capture of IDS by the antibody coated to the solid phase is critical for this enzyme to react with the 4MU sulfate substrate, because no reactivity was detected in the absence of capture antibody or when the antibody and enzyme were together in solution. Surface adsorption has been known to alter protein characteristics (29), and presumably, interaction between the protein and the adsorbed capture antibody led to an adsorption-induced conformation change, enhancing enzyme activity. Of note, a similar system for immune capture did not show increased activity with sulfamidase and 4MU sulfate (our unpublished observations).

The methods for IDS protein and activity measurements described here are simple to perform, have potential as diagnostic assays, and are adaptable to large-scale, high-throughput protocols. We estimate that the cost for production and purification of 2.5 g of sheep anti-IDS polyclonal antibody, enough to cover 5 million newborns (annual US birth rate), to be approximately AUS $0.01 per sample. This cost includes chromatography columns for purification (approximately AUS $4000.00),100 mg of IDS protein (approximately AUS $20 000.00), and labor (approximately AUS $26 000.00), to give a final cost of AUS $50 000.00. The preferred use of either enzyme activity or protein concentration determinations will depend to a large degree on the application. For a diagnostic assay, the use of enzyme activity measurements on dried blood spots or plasma would enable identification of all patients, including those with significant concentrations of mutant protein. However, newborn screening for a single rare disorder such as MPS II, with a reported incidence of 1 in 162 000 (30), would not be economically viable. A screening program for LSDs will require screening for multiple LSDs in a single-assay format. Although there is currently no technology available to meld multiple fluorogenic enzyme assays together, multiplex technology is available for protein quantification (31). As such, this method provides a viable option for the development of a newborn-screening program for multiple LSDs. A potential drawback of this approach is that immune quantification of IDS protein will miss those patients with significant concentrations of inactive protein. In this study, 1 of the 19 MPS II individuals (5%) had IDS protein concentrations that were within the reference interval; therefore, MPS II would not be identified in this patient from a single IDS protein assay. We previously reported that the lysosomal storage of mucopolysaccharides leads to an increase in the number and size of lysosomal vacuoles (32). Furthermore, we reported alterations to other lysosomal protein concentrations such as LAMP-1 (21), LAMP-2 (33), and saposin C (34) in several LSDs, including MPS II. Multiplexing of the IDS protein assay with another 10 lysosomal proteins has been reported (35). Thus, we anticipate that alterations in multiple lysosomal proteins (a protein profile) will enable differentiation of patients with MPS II and other LSDs from the control population even if their mutant protein concentrations are within reference intervals. This technology could provide a platform for the further development of a newbornscreening program for LSDs.

We thank Emma Parkinson-Lawrence for production of the IDS affinity column and polyclonal antibody, Alison Whittle for the polyclonal antibody purification, and Debbie Lang for europium labeling of the polyclonal antibody. This work was supported by the National Health and Medical Research Council of Australia.

Received October 10, 2005; accepted February 2, 2006.

Previously published online at DOI: 10.1373/clinchem.2005.061838

References

(1.) Neufeld EF, Muenzer J. The mucopolysaccharidoses. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic bases of inherited diseases, 8th ed. New York: McGraw-Hill, 2001:3421-52.

(2.) Stenson PD, Ball EV, Mort M, Phillips AD, Shiel JA, Thomas NS, et al. Human Gene Mutation Database (HGMD): 2003 update. Hum Mutat 2003;21:577-81.

(3.) Villani GR, Balzano N, Grosso M, Salvatore F, Izzo P, Di Natale P. Mucopolysaccharidosis type II: identification of six novel mutations in Italian patients. Hum Mutat 1997;10:71-5.

(4.) Froissart R, Moreira da Silva I, Guffon N, Bozon D, Maire I. Mucopolysaccharidosis type II--genotype/phenotype aspects. Acta Paediatr Suppl 2002;91:82-7.

(5.) Parkinson EJ, Muller V, Hopwood JJ, Brooks DA. Iduronate-2-sulphatase protein detection in plasma from mucopolysaccharidosis type II patients. Mol Genet Metab 2004;81:58-64.

(6.) Vellodi A, Young E, Cooper A, Lidchi V, Winchester B, Wraith JE. Long-term follow-up following bone marrow transplantation for Hunter disease. J Inherit Metab Dis 1999;22:638-48.

(7.) Wenstrup RJ, Bailey L, Grabowski GA, Moskovitz J, Oestreich AE, Wu W, et al. Gaucher disease: alendronate disodium improves bone mineral density in adults receiving enzyme therapy. Blood 2004;104:1253-7.

(8.) Wilcox WR, Banikazemi M, Guffon N, Waldek S, Lee P, Linthorst GE, et al. Long-term safety and efficacy of enzyme replacement therapy for Fabry disease. Am J Hum Genet 2004;75:65-74.

(9.) Wraith JE, Clarke LA, Beck M, Kolodny EH, Pastores GM, Muenzer J, et al. Enzyme replacement therapy for mucopolysaccharidosis I: a randomized, double-blinded, placebo-controlled, multinational study of recombinant human [alpha]-L-iduronidase (laronidase). J Pediatr 2004;144:581-8.

(10.) Harmatz P, Ketteridge D, Giugliani R, Guffon N, Teles EL, Miranda MC, et al. Direct comparison of measures of endurance, mobility, and joint function during enzyme-replacement therapy of mucopo lysaccharidosis VI (Maroteaux-Larry syndrome): results after 48 weeks in a phase 2 open-label clinical study of recombinant human IV-acetylgalactosamine 4-sulfatase. Pediatrics 2005;115: e681-9.

(11.) Muenzer J, Lamsa JC, Garcia A, Dacosta J, Garcia J, Treco DA. Enzyme replacement therapy in mucopolysaccharidosis type II (Hunter syndrome): a preliminary report. Acta Paediatr Suppl 2002;91:98-9.

(12.) Fuller M, Brooks DA, Evangelista M, Hein LK, Hopwood JJ, Meikle PJ. Prediction of neuropathology in mucopolysaccharidosis I patients. Mol Genet Metab 2005;84:18-24.

(13.) Hein LK, Meikle PJ, Dean CJ, Bockmann MR, Auclair D, Hopwood JJ, et al. Development of an assay for the detection of mucopolysaccharidosis type VI patients using dried blood-spots. Clin Chim Acta 2005;353:67-74.

(14.) Fuller M, Lovejoy M, Brooks DA, Harkin ML, Hopwood JJ, Meikle PJ. Immunoquantification of [alpha-galactosidase: evaluation for the diagnosis of Fabry disease. Clin Chem 2004;50:1979-85.

(15.) Umapathysivam K, Hopwood JJ, Meikle PJ. Determination of acid [alpha]-glucosidase activity in blood spots as a diagnostic test for Pompe disease. Clin Chem 2001;47:1378-83.

(16.) Keulemans JL, Sinigerska I, Garritsen VH, Huijmans JG, Voznyi YV, van Diggelen OP, et al. Prenatal diagnosis of the Hunter syndrome and the introduction of a new fluorimetric enzyme assay. Prenat Diagn 2002;22:1016-21.

(17.) Bielicki J, Freeman C, Clements PR, Hopwood JJ. Human liver iduronate-2-sulphatase. Purification, characterization, and catalytic properties. Biochem J 1990;271:75-86.

(18.) Voznyi YV, Keulemans JL, van Diggelen OP. A fluorimetric enzyme assay for the diagnosis of MPS II (Hunter disease). J Inherit Metab Dis 2001;24:675-80.

(19.) Chamoles NA, Blanco MB, Gaggioli D, Casentini C. Hurler-like phenotype: enzymatic diagnosis in dried blood spots on filter paper. Clin Chem 2001;47:2098-102.

(20.) Bielicki J, Hopwood JJ, Wilson PJ, Anson DS. Recombinant human iduronate-2-sulphatase: correction of mucopolysaccharidosis-type II fibroblasts and characterization of the purified enzyme. Biochem J 1993;289(Pt 1):241-6.

(21.) Meikle PJ, Brooks DA, Ravenscroft EM, Yan M, Williams RE, Jaunzems AE, et al. Diagnosis of lysosomal storage disorders: evaluation of lysosome-associated membrane protein LAMP-1 as a diagnostic marker. Clin Chem 1997;43:1325-35.

(22.) Anson DS, Taylor JA, Bielicki J, Harper GS, Peters C, Gibson GJ, et al. Correction of human mucopolysaccharidosis type-VI fibroblasts with recombinant IV-acetylgalactosamine-4-sulphatase. Biochem J 1992;284(Pt 3):789-94.

(23.) Hopwood JJ. [alpha]-L-Iduronidase, [beta]-D-glucuronidase, and 2-sulfo-Liduronate 2-sulfatase: preparation and characterization of radioactive substrates from heparin. Carbohydr Res 1979;69:203-16.

(24.) Meikle PJ, Fietz MJ, Hopwood JJ. Diagnosis of lysosomal storage disorders: current techniques and future directions. Expert Rev Mol Diagn 2004;4:677-91.

(25.) Villani GR, Daniele A, Balzano N, Di Natale P. Expression of five iduronate-2-sulfatase site-directed mutations. Biochim Biophys Acta 2000;1501:71-80.

(26.) Bonuccelli G, Di Natale P, Corsolini F, Villani G, Regis S, Filocamo M. The effect of four mutations on the expression of iduronate-2-sulfatase in mucopolysaccharidosis type II. Biochim Biophys Acta 2001;1537:233-8.

(27.) Hopwood JJ, Ballabio A. Multiple sulfatase deficiency and the nature of the sulfatase family. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic bases of inherited diseases, 8th ed. New York: McGraw-Hill, 2001:3725-32.

(28.) Hanson SR, Best MD, Wong CH. Sulfatases: structure, mechanism, biological activity, inhibition, and synthetic utility. Angew Chem Int Ed Engl 2004;43:5736-63.

(29.) Butler JE. Solid supports in enzyme-linked immunosorbent assay and other solid-phase immunoassays. Methods 2000;22:4-23.

(30.) Meikle PJ, Hopwood JJ, Clague AE, Carey WF. Prevalence of lysosomal storage disorders. JAMA 1999;281:249-54.

(31.) Hadd AG, Brown JT, Andruss BF, Ye F, WalkerPeach CR. Adoption of array technologies into the clinical laboratory. Expert Rev Mol Diagn 2005;5:409-20.

(32.) Karageorgos LE, Isaac EL, Brooks DA, Ravenscroft EM, Davey R, Hopwood JJ, et al. Lysosomal biogenesis in lysosomal storage disorders. Exp Cell Res 1997;234:85-97.

(33.) Hua CT, Hopwood JJ, Carlsson SR, Harris RJ, Meikle PJ. Evaluation of the lysosome-associated membrane protein LAMP-2 as a marker for lysosomal storage disorders. Clin Chem 1998;44: 2094-102.

(34.) Chang MH, Bindloss CA, Grabowski GA, Qi X, Winchester B, Hopwood JJ, et al. Saposins A, B, C, and D in plasma of patients with lysosomal storage disorders. Clin Chem 2000;46:167-74.

(35.) Meikle PJ, Dean CJ, Grasby D, Bockmann MR, Whittle AM, Lang DL, et al. Newborn screening for lysosomal storage disorders: evaluation of protein profiling [Abstract]. J Inherit Metab Dis 2005;28(Suppl 1):14.

CAROLINE J. DEAN, [1] MICHELLE R. BOCKMANN, [1] JOHN J. HOPWOOD, [1,2] DOUG A. BROOKS, [1,2] and PETER J. MEIKLE [1,2] *

[1] Lysosomal Diseases Research Unit, Department of Genetic Medicine, Children, Youth and Women's Health Service, North Adelaide, South Australia, Australia.

[2] Department of Paediatrics, University of Adelaide, South Australia, Australia.

[3] Nonstandard abbreviations: MPS II, mucopolysaccharidosis II; LSD, lysosomal storage disorder; IDS, iduronate-2-sulfatase; 4MU sulfate, 4-methylumbelliferyl sulfate; DELFIA, dissociation-enhanced lanthanide fluorescence immune-assay; and BSA, bovine serum albumin.

* Address correspondence to this author at: Lysosomal Diseases Research Unit, Department of Genetic Medicine, Children Youth and Women's Health Service, 72 King William Road, North Adelaide, South Australia, 5006, Australia. Fax 61-8-8161-7100; e-mail peter.meikle@adelaide.edu.au.
COPYRIGHT 2006 American Association for Clinical Chemistry, Inc.
No portion of this article can be reproduced without the express written permission from the copyright holder.
Copyright 2006 Gale, Cengage Learning. All rights reserved.

Article Details
Printer friendly Cite/link Email Feedback
Title Annotation:Molecular Diagnostics and Genetics
Author:Dean, Caroline J.; Bockmann, Michelle R.; Hopwood, John J.; Brooks, Doug A.; Meikle, Peter J.
Publication:Clinical Chemistry
Date:Apr 1, 2006
Words:4563
Previous Article:Quantification of mRNA in whole blood by assessing recovery of RNA and efficiency of cDNA synthesis.
Next Article:Estimation and application of biological variation of urinary [delta]-aminolevulinic acid and porphobilinogen in healthy individuals and in patients...
Topics:

Terms of use | Privacy policy | Copyright © 2020 Farlex, Inc. | Feedback | For webmasters