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Development of a fluorometric microtiter plate based enzyme assay for MPS IVA (Morquio type A) using dried blood spots

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ABSTRACT

Mucopolysaccharidosis type IVA or Morquio type-A disease is a hereditary lysosomal storage disorder caused by deficient activity of the lysosomal enzyme N-acetylgalactosamine-6-sulfate sulfatase (GALNS). The disease is caused by lysosomal accumulation of unprocessed glycosaminoglycans (GAGs) that manifests with severe to mild skeletal and cardiopulmonary abnormalities. We have developed a modified microtiter plate-based enzyme activity assay using dried blood spots and a fluorescent substrate for measuring specific GALNS activity to identify patients with MPS IVA.

No MeSH data available.


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A) Comparison of GALNS enzyme activity in normal control (circles) and MPS IVA (squares) dried blood spots. B) Specificity of assay for GALNS activity in DBS from affected MPS IVA compared to other known LSDs and normal controls. C) Stability of GALNS activity in dried-blood spots was tested by incubating DBS (n = 10) at 30 °C (circles) & 37 °C (squares) for 6 h, 16 h and 30 h. D) Enzyme activity was measured in ten DBS stored long-term (7 days) at − 20 °C, 4 °C & 22 °C.
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f0005: A) Comparison of GALNS enzyme activity in normal control (circles) and MPS IVA (squares) dried blood spots. B) Specificity of assay for GALNS activity in DBS from affected MPS IVA compared to other known LSDs and normal controls. C) Stability of GALNS activity in dried-blood spots was tested by incubating DBS (n = 10) at 30 °C (circles) & 37 °C (squares) for 6 h, 16 h and 30 h. D) Enzyme activity was measured in ten DBS stored long-term (7 days) at − 20 °C, 4 °C & 22 °C.

Mentions: We then determined relative GALNS enzyme activity levels in the DBS from known MPS IVA patients (n = 13) and normal control DBS (n = 75). The DBS from normal individuals (gray circles) from the Duke repository had enzyme activities in the range of 15.0–33.8 pmol/punch/h, with calculated mean ± standard deviation values of 21.4 ± 3.97 pmol/punch/h. The MPS IVA-affected DBS (black squares) had GALNS activity ranging from 7.4 to 10.2 pmol/punch/h, with a mean value of 8.94 ± 0.97 (mean ± std. dev.). GALNS-deficient, MPS-IVA DBS values were clearly distinguishable from the normal control spots as shown in Fig. 1A (p < 0.0001, Tukey's two-tailed t-test).


Development of a fluorometric microtiter plate based enzyme assay for MPS IVA (Morquio type A) using dried blood spots
A) Comparison of GALNS enzyme activity in normal control (circles) and MPS IVA (squares) dried blood spots. B) Specificity of assay for GALNS activity in DBS from affected MPS IVA compared to other known LSDs and normal controls. C) Stability of GALNS activity in dried-blood spots was tested by incubating DBS (n = 10) at 30 °C (circles) & 37 °C (squares) for 6 h, 16 h and 30 h. D) Enzyme activity was measured in ten DBS stored long-term (7 days) at − 20 °C, 4 °C & 22 °C.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5121358&req=5

f0005: A) Comparison of GALNS enzyme activity in normal control (circles) and MPS IVA (squares) dried blood spots. B) Specificity of assay for GALNS activity in DBS from affected MPS IVA compared to other known LSDs and normal controls. C) Stability of GALNS activity in dried-blood spots was tested by incubating DBS (n = 10) at 30 °C (circles) & 37 °C (squares) for 6 h, 16 h and 30 h. D) Enzyme activity was measured in ten DBS stored long-term (7 days) at − 20 °C, 4 °C & 22 °C.
Mentions: We then determined relative GALNS enzyme activity levels in the DBS from known MPS IVA patients (n = 13) and normal control DBS (n = 75). The DBS from normal individuals (gray circles) from the Duke repository had enzyme activities in the range of 15.0–33.8 pmol/punch/h, with calculated mean ± standard deviation values of 21.4 ± 3.97 pmol/punch/h. The MPS IVA-affected DBS (black squares) had GALNS activity ranging from 7.4 to 10.2 pmol/punch/h, with a mean value of 8.94 ± 0.97 (mean ± std. dev.). GALNS-deficient, MPS-IVA DBS values were clearly distinguishable from the normal control spots as shown in Fig. 1A (p < 0.0001, Tukey's two-tailed t-test).

View Article: PubMed Central - PubMed

ABSTRACT

Mucopolysaccharidosis type IVA or Morquio type-A disease is a hereditary lysosomal storage disorder caused by deficient activity of the lysosomal enzyme N-acetylgalactosamine-6-sulfate sulfatase (GALNS). The disease is caused by lysosomal accumulation of unprocessed glycosaminoglycans (GAGs) that manifests with severe to mild skeletal and cardiopulmonary abnormalities. We have developed a modified microtiter plate-based enzyme activity assay using dried blood spots and a fluorescent substrate for measuring specific GALNS activity to identify patients with MPS IVA.

No MeSH data available.


Related in: MedlinePlus