A simple and sensitive method for rapid semiquantitive determination of urinary protein was devised. Five μl of the urine to be measured is dropped on the 2cm(2) filter paper, and dried at room temperature. The paper is immersed in a mixture of ethanol, acetic acid and distillated water (35:10:55) which contains 0.2% Coomassie Brilliant Blue R-250 (CBB), and washed with water until the waste water becomes colorless. The paper is again immersed into a mixture but without CBB, after which the paper is rinsed with water again. The protein concentration in the urine sample is determined by comparing the hue of the urine spot with a standard violet hue chart, as well as by the color intensity of the extract of the spot with 3% hydrogen chloride in ethanol at 595 nm by spectrophotometry. This method was applied to the determination of protein in urine from junior college girls immediatly after arriving at school and after cooking study, from adults in a factory at the time of their regular medical examination, and from high school boys before and after physical exercise. The results were compared with that of the TONEIN-TP method. With the SPOTCBB-DYE method, samples can be kept as colored spots on the filter paper, and the protein in urine can be preserved as a spot on the filter paper and measured at any time.
Coomassie Brilliant Blue R-250