Abstract
The fluorescent antibody method is widely used as a routine procedure for the histochemical examination of enzymes. In the present paper, the antiserum used in such procedures was applied to quantitative immunoelectrophoresis for the microquantification of enzymes. Bovine liver tissue in amounts corresponding to that available from liver biopsies was used to perform quantitative determinations of arginase which is the key enzyme in the urea cycle.
Antiserum was obtained by sensitization of a rabbit using standard crystalline bovine liver arginase, which had been obtained from the Boehlinger Mannheim Company and purified by disc electrophoresis. This antiserum was mixed with 1% agarose, the supporting medium, and then quantitative immunoelectrophoresis was performed using 5μl of the supernatant of bovine liver homogenate.
The area enclosed within the precipitation lines that had been produced as a result of electrophoresis was confirmed to be proportional to the enzymatic activity. Furthermore, based upon the knowledge that the standard arginase had an enzymatic activity of 70 units per mg, the enzymatic activity units were calculated for the area enclosed within the precipitation lines.
It was thus demonstrated that microquantification is possible. In addition, the DNA content within the precipitate that had been produced during the preparation of the sample solution was measured and used as the index of the cell count. This value was used to calculate the enzymatic activity per mg DNA.
