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Open AccessArticle

Optimization of Enzyme Essays to Enhance Reliability of Activity Measurements in Leukocyte Lysates for the Diagnosis of Metachromatic Leukodystrophy and Gangliosidoses

Department of Pediatric Neurology, University Children’s Hospital Tübingen, Hoppe-Seyler-Straße 1, 72076 Tübingen, Germany
*
Author to whom correspondence should be addressed.
S.S. and N.H. contributed equally to this work.
Cells 2020, 9(12), 2553; https://doi.org/10.3390/cells9122553
Received: 30 October 2020 / Revised: 23 November 2020 / Accepted: 25 November 2020 / Published: 28 November 2020
(This article belongs to the Special Issue Lysosomal Storage Disorders)
(1) Lysosomal storage diseases are rare inherited disorders with no standardized or commercially available tests for biochemical diagnosis. We present factors influencing the quality of enzyme assays for metachromatic leukodystrophy (MLD) and gangliosidoses (GM1; GM2 variants B and 0) and validate the reliability and stability of testing in a retrospective analysis of 725 samples. (2) Patient leukocytes were isolated from ethylene-diamine-tetra-acetic acid (EDTA) blood and separated for subpopulation experiments using density gradient centrifugation or magnetic cell separation. Enzyme activities in whole leukocyte lysate and leukocyte subpopulations were determined. (3) The enzyme activities in leukocyte subpopulations differed significantly. Compared to lymphocytes, the respective enzyme activities were 2.31–4.57-fold higher in monocytes and 1.64–2.81-fold higher in granulocytes. During sample preparation, a considerable amount of the lysosomal enzymes was released from granulocytes. Nevertheless, with the sample preparation method used here, total leukocyte count proved to be more accurate than total protein amount as a reference unit for enzyme activities. Subsequent analysis of 725 individuals showed clear discrimination of enzyme activities in patient samples (48 MLD; 21 gangliosidoses), with a sensitivity of 100% and specificity of 98–99%. View Full-Text
Keywords: lysosomal storage disease; sphingolipidoses; metachromatic leukodystrophy; gangliosidoses lysosomal storage disease; sphingolipidoses; metachromatic leukodystrophy; gangliosidoses
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MDPI and ACS Style

Strobel, S.; Hesse, N.; Santhanakumaran, V.; Groeschel, S.; Bruchelt, G.; Krägeloh-Mann, I.; Böhringer, J. Optimization of Enzyme Essays to Enhance Reliability of Activity Measurements in Leukocyte Lysates for the Diagnosis of Metachromatic Leukodystrophy and Gangliosidoses. Cells 2020, 9, 2553. https://doi.org/10.3390/cells9122553

AMA Style

Strobel S, Hesse N, Santhanakumaran V, Groeschel S, Bruchelt G, Krägeloh-Mann I, Böhringer J. Optimization of Enzyme Essays to Enhance Reliability of Activity Measurements in Leukocyte Lysates for the Diagnosis of Metachromatic Leukodystrophy and Gangliosidoses. Cells. 2020; 9(12):2553. https://doi.org/10.3390/cells9122553

Chicago/Turabian Style

Strobel, Sebastian; Hesse, Naomi; Santhanakumaran, Vidiyaah; Groeschel, Samuel; Bruchelt, Gernot; Krägeloh-Mann, Ingeborg; Böhringer, Judith. 2020. "Optimization of Enzyme Essays to Enhance Reliability of Activity Measurements in Leukocyte Lysates for the Diagnosis of Metachromatic Leukodystrophy and Gangliosidoses" Cells 9, no. 12: 2553. https://doi.org/10.3390/cells9122553

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