Temperature regulation of microtiter plates for enzyme assays

To facilitate the use of microtiter plates as vessels for enzyme assays, an incubator has been designed to maintain the wells of the microtiter plates at the appropriate temperature. The temperature variation within a single well was +/- 0.02 degrees (standard error of the mean) at 25 degrees C and +/- 0.02 degrees at 37 degrees C. The temperature variation was the same for internal and peripheral wells within the plates, although the internal wells were approximately 0.14 degrees C warmer than the outer wells at 25 degrees C and 0.68 degrees C cooler at 37 degrees C. The overall uncertainty (95% confidence interval) of the well temperature in a typical plate was +/- 0.4 degrees C at 25 degrees C and +/- 0.7 degrees C at 37 degrees C. This uncertainty is realistic for routine enzyme determinations, as opposed to precise studies. The incubator was designed to allow access to the plates from above so that additions could be made during the incubation. To demonstrate the suitability of microtiter plates and the incubator for enzyme determinations, this method was used to measure the activity of myeloperoxidase and alpha-naphthylbutyryl esterase.