Histochemical technique for the demonstration of phosphofructokinase activity in heart and skeletal muscles.
A histochemical multi-step technique for the demonstration of phosphofructokinase activity in tissue sections is described. With this technique a semipermeable membrane is interposed between the incubating solution and the tissue sections preventing diffusion of the non-structurally bound enzyme into the medium during incubation. In the histochemical system the enzyme converts the substrate D-fructose-6-phosphate to D-fructose-1,6-diphosphate, which in turn is hydrolyzed by exogenous and endogenous fructose diphosphate aldolase to dihydroxyacetone phosphate and D-glyceral-dehyde-3-phosphate. The dihydroxyacetone phosphate is reversibly converted into D-glyceraldehyde-3-phosphate by exogenous and endogenous triosephosphate isomerase. Next the D-glyceraldehyde-3-phosphate is oxidized by exogenous and endogenous glyceraldehyde-3-phosphate dehydrogenase into 1,3-diphospho-D-glycerate. Concomitantly the electrons are transported via NAD+, phenazine methosulphate and menadione to nitro-BT. Sodium azide and amytal are incorporated to block electron transfer to the cytochromes.
D000693:Anaerobiosis; D000818:Animals; D002417:Cattle; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D006651:Histocytochemistry; D006801:Humans; D009132:Muscles / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009580:Nitroblue Tetrazolium / Q000378:metabolism; D010732:Phosphofructokinase-1 / Q000378:metabolism*; D011690:Purkinje Fibers / Q000201:enzymology; D051381:Rats; D014305:Triose-Phosphate Isomerase / Q000378:metabolism
D009243:NAD; D009580:Nitroblue Tetrazolium; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010732:Phosphofructokinase-1; D014305:Triose-Phosphate Isomerase