Determination of proteins in the presence of imidazole buffers

Imidazole buffers were found to interfere with the determination of soluble proteins using Lowry's classical method. The influence of the constituent elements of the buffer on the calibration line was studied statistically. By combining the data corresponding to different experimental sequences, interserial calibration curves for different concentrations of imidazole buffer (10-30 mM) were obtained. The absorbance-buffer volume dependence curves produced a good fit to second-order polynomials. The accuracy of protein determination in a medium with imidazole buffer, using appropriate calibration curves, were tested by comparison with the technique of multiple standard addition and by means of recovery studies. These experiments were performed on chick brain homogenate samples. Other important aspects of validation, such as sensitivity and accuracy, were also studied.