Immunocapture and Microplate-Based Activity and Quantity Measurement of Pyruvate Dehydrogenase in Human Peripheral Blood Mononuclear Cells

Pyruvate dehydrogenase (PDH) activity is altered in many human disorders. Current methods require tissue samples and yield inconsistent results. We describe a modified method for measuring PDH activity from isolated human peripheral blood mononuclear cells (PBMCs). RESULTS/METHODOLOGY: We found that...

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Published inBioanalysis Vol. 7; no. 5; pp. 583 - 592
Main Authors Liu, Xiaowen, Pervez, Hira, Andersen, Lars W, Uber, Amy, Montissol, Sophia, Patel, Parth, Donnino, Michael W
Format Journal Article
LanguageEnglish
Published England 01.03.2015
Subjects
Humans
Leukocytes, Mononuclear - immunology
Pyruvate Dehydrogenase Complex - immunology
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Summary:Pyruvate dehydrogenase (PDH) activity is altered in many human disorders. Current methods require tissue samples and yield inconsistent results. We describe a modified method for measuring PDH activity from isolated human peripheral blood mononuclear cells (PBMCs). RESULTS/METHODOLOGY: We found that PDH activity and quantity can be successfully measured in human PBMCs. Freeze-thaw cycles cannot efficiently disrupt the mitochondrial membrane. Processing time of up to 20 h does not affect PDH activity with proteinase inhibitor addition and a detergent concentration of 3.3% showed maximum yield. Sample protein concentration is correlated to PDH activity and quantity in human PBMCs from healthy subjects. Measuring PDH activity from PBMCs is a novel, easy and less invasive way to further understand the role of PDH in human disease.
ISSN:1757-6180
1757-6199
DOI:10.4155/bio.14.302