A simple screening procedure for the detection of glucose-phosphate isomerase (GPI), phosphofructokinase (PFK), aldolase (AL) and glyceraldehyde-3-phosphate dehydrogenase (GAPD) deficiencies in blood, is described. These enzymes catalyze the second, third, fourth, and sixth reactions in the Embden-Meyerhof pathway. The procedure is based on the conversion of glucose-6-phosphate to 1,3-diphosphoglycerate (1,3-DPG) which is catalyzed by the sequential action of the GPI, PFK, AL and GAPD. The presence of the enzyme activities is visually estimated by the reduction of NAD+ (non-fluorescent) to NADH (fluorescent) which occurs when 1,3-DPG is formed. Absence of fluorescence indicates the deficiency of anyone of the four enzymes, which are specified by using separately the PFK, AL and GAPD respective substrates.

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