Aldolase Assay

Method: Based on Boyer's modification of the hydrazine assay (Jagannathan et al. 1956) in which 3-phosphoglyceraldehyde reacts with hydrazine to form a hydrazone which absorbs at 240 nm. The advantages of the system are its simplicity and specificity. One unit is described as a change in absorbance of 1.00 per minute at 25°C and pH 7.5 under the specified conditions. Alternate methods for the assay of aldolase have been described by Rutter (1961). The coupled a-glycerophosphate dehydrogenase assay (Richards et al. 1961) has been used.


  • 0.0001 M EDTA, pH 7.5 containing 0.0035 M hydrazine sulfate
  • Substrate: 0.012 M Fructose-1,6-bisphosphate pH 7.5


Immediately before use, dilute in water to a concentration of 0.5-2.0 units per ml.



Set spectrophotometer at 240 nm and 25°C.

Pipette into cuvettes as follows:

  Blank Test
Fructose-1,6-bisphosphate 1.0 ml 1.0 ml
Hydrazine sulfate 2.0 ml 2.0 ml
Distilled water 0.1 ml ----
Record ΔA240 for 10 minutes    
Add enzyme ---- 0.1 ml
Record ΔA240 for 10 minutes    

Subtract ΔA240/min of the blank from ΔA240/min of the test.



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