Revised assays for the investigation of congenital lactic acidosis using 14C keto acids, eliminating problems associated with spontaneous decarboxylation.

Abstract

Improved methods using 14C keto acids for the investigation of patients with congenital lactic acidoses are described. The addition of rat serum to assay media reduces the spontaneous decarboxylation of [1-14C] and [2-14C] pyruvate and alpha-[1-14C]ketoglutarate to low levels. A study of the stability of pyruvate dehydrogenase in fibroblasts has shown that the activity is rapidly lost when cell membranes are broken unless homogenisation is done gently at -15 degrees C. Under these conditions broken cell preparations may be stored for up to 3 hours without loss of activity. Freezing and thawing results in unpredictable changes in pyruvate dehydrogenase activity. A quality control solution containing pyruvate dehydrogenase activity has been prepared which is stable for at least 6 months (coefficient of variation = 7.7%). Normal values for pyruvate dehydrogenase in fibroblasts range from 0.59 to 1.26 nmol . min-1 . mg-1 protein (mean = 0.98, n = 8) and pyruvate dehydrogenase deficient fibroblasts can be detected with confidence.

• Authors: K Hyland, J V Leonard
• Journal: Clinica chimica acta; international journal of clinical chemistry (Clin Chim Acta) Vol. 133 Issue 2 Pg. 177-87 (Sep 30 1983) ISSN: 0009-8981 [Print] Netherlands
• PMID: 6414741 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Carbon Radioisotopes
• Keto Acids
• Lactates
• Pyruvate Dehydrogenase Complex

Topics

• Acidosis (congenital, metabolism)
• Animals
• Blood
• Carbon Radioisotopes
• Cell Line
• Decarboxylation
• Drug Stability
• Fibroblasts (enzymology)
• Humans
• Keto Acids (metabolism)
• Lactates
• Pyruvate Dehydrogenase Complex (metabolism)
• Pyruvate Dehydrogenase Complex Deficiency Disease
• Rats