Automatic assay of urinary protein using Coomassie Brilliant Blue G-250

Abstract

An automatic method for the protein assay using Coomassie Brilliant Blue G-250 was developed and applied to the assay of urinary proteins. In developing the automatic system, the adhesion of protein-bound dye to the walls of the flow cell and tubes was found to be the most troublesome problem, by which the baseline was shifted upwardly to give positive errors. For the purpose of preventing such adhesion, the concentration of CBB was reduced to half of that used in the manual method, glass tubes and glass coils were changed to those made of Kel-F material, and the flow cell was coated with fluorine resin. As a result, the staining with protein-bound dye was nearly completely eliminated. The final system showed satisfactory ability in performance, namely, the value of a coefficient variation for the reproducibility within run was 1.3%, that for the carry over was 0–1.1%, and the recovery was 98.8%. The calibration curve was linear in a range of 0–1000 μg/ml, and 80 samples could be processed in 1 h. Thus, the present method may serve as an efficient automatic protein analyzer for routine clinical tests of urine samples.