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Estimating the initial reaction velocity of a soluble dehydrogenase in situ

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Summary

The initial reaction velocities (v i ) of lactate dehydrogenase in single hepatocytes were determined, by microdensitometry or computer-assisted image analysis, in sections of unfixed mouse liver incubated at 37°C on substrate-containing agarose gel films. They were found to fit the equations v i =2.82°A and v i =v+2°A, where v and °A are, respectively, gradients (or steady-state linear velocities) and the intercepts on the absorbance axis of the linear regression lines of the absorbance (A) on incubation time plots for incubation times between 1 and 3 min. Both equations were independent of section thickness between 4 and 14 μm. The observed and calculated values of v i , agreed within 11.5% (n = 71). The validity of the equations for v i was confirmed by showing that the calculated v i was proportional to the thickness of the section and hence the amount of enzyme present. Thus, v i can be determined from measurements of either °A alone or v and °A.

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Nakae, Y., Stoward, P.J. Estimating the initial reaction velocity of a soluble dehydrogenase in situ . Histochem J 25, 199–205 (1993). https://doi.org/10.1007/BF00163815

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