Studies on urinary protein after physical load II.Microdetermination of urinary protein using filter paper and CBB staining

A simple and sensitive method for rapid semiquantitative and quantitative determinations of urinary protein was devised, in which protein absorbed and fixed on filter paper containing sulfosalicylic acid was reacted with Coomassie Brilliant Blue G-250 (CBB). For the semiquantitative method, filter paper (NO. 50, Toyo) was immersed in a 20% sulfosalicylic acid solution and dried. Dried filter paper was immersed in aurine, dried again immersed TONEIN (Othuka Assay Laboratories) and compared with standard filter paper by the naked eye. For the quantitative method, filter paper was immersed in a mixture of ethanol, acetic acid and distilled water (35 : 10 : 55) containing 0.2% CBB, and washed water untill the waste water became colorless. The paper was again immersed into the mixture but without CBB, after which the paper was rinsed with water again. The protein concentration in the urine sample was determined by comparing the hue of the urine with a standard violet hue chart by the naked eye, or the color intensity of the extract of 1c㎡ of filter paper with 3% hydrogen chloride in ethanol was determined at 590 nm.