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SIGNIFICANCE OF COUPLING RATE IN HISTOCHEMICAL AZO DYE METHODS FOR ENZYMES

V. DEFENDI 1 and A. G. E. PEARSE 1

1 Postgraduate Medical School of London, W. 12

1) In order to estimate the significance of the coupling rate in histochemical localization of enzymes a series of naphthols has been tested, using a streaming method with photoelectric recording. The naphthols were those used in histochemical methods (as esters) and the diazonium salt employed (Fast Blue B salt) was chosen as being the one most often used in histochemistry.

2) It was observed that the usual straight line relationship between the bimolecular velocity constant of coupling and pH (Conant and Peterson, 1930) became invalid at certain "critical points." For beta-naphthol the "critical point" lay between pH 7.4 and 7.0 and for agr-naphthol between pH 5.3 and 5.0.

3) Parallel histochemical studies indicated that the "critical point" and therefore the coupling rate, was likely to be important in the case of the more soluble naphthols, but of less importance with the relatively insoluble ones, in producing accurate histochemical localization of enzyme.


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