By collecting released 14CO2 following the enzymatic decarboxylation of radiolabeled lactate, picomoles of the latter can be precisely, easily, and reproducibly measured in small biological fluids. This radioactive [14C]lactate microassay does not require a neutralization step, nor does it require chemical extractions and partioning procedures, ion exchange, or pyruvate derivatization. Under our specified conditions this simple reaction goes to completion in 90 min. Using this assay in porous adipose cells, with the cell number logarithmically less than that found in other literature methods, the measured glycolytic flux rates were consistent with those previously reported. In these studies, glycolysis was initiated with [U-14C]glucose 6-phosphate. This microradioactive lactate assay is useful when dealing with minute tissue samples and/or microliter aliquots of biological fluids.