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J.Health Sci., 52(6), 718-723, 2006
Improved Recovery of Human Urinary Protein for Electrophoresis
Tetsuya Fujimoto,a Makiko Miya,a Maiko Machida,a
Shigeru Takechi,b Shigeo Kakinoki,c Koichi Kanda,d and
Akikazu Nomura*, a
aDepartment of Pathophysiology, Hokkaido Pharmaceutical University, School of Pharmacy, 7-1 Katsuraoka, Otaru, Hokkaido 047-0264, Japan,
bDepartment of Cardiovascular Medicine, Date Red Cross Hospital, 81 Suenaga, Date, Hokkaido 052-8511, Japan,
cDepartment of Cardiovascular Medicine, Otaru Kyokai Hospital, 6-15 Suminoe-1-chome, Otaru, Hokkaido 047-8510, Japan, and
dDepartment of Cardiovascular Medicine, Sapporo Kosei Hospital, 5 Higashi-8-chome, Kita-3-jo, Chuoku, Sapporo, Hokkaido 060-0033, Japan
Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) is frequently used to analyze urinary proteins, but appropriate urine sample preparation is essential, because urinary proteins are present in small concentrations, and urine contains high concentrations of salts and metabolic wastes. The aim of this study was to determine the optimum method for urine sample preparation. The effects on urinary protein recovery of pretreating the ultrafiltration apparatus with various agents (Triton X-100, Tween20, PEG compound, and SDS) were studied; Triton X-100 was found to be the most effective agent. Ultrafiltration and acetone precipitation were compared as sample preparation methods for SDS-PAGE. Recovery of urinary protein using ultrafiltration (89.5 ± 8.1%) was better than that achieved by using acetone precipitation (75.1 ± 12.2%) (p < 0.01). Integrated densitometric values for five protein bands (62, 52, 39, 31, and 6 kDa) were higher for ultrafiltration samples than for acetone samples (42350 ± 2568 vs. 37010 ± 725, 34665 ± 1519 vs. 32355 ± 2278, 10755 ± 1683 vs. 8870 ± 941, 65920 ± 3354 vs. 60680 ± 3651, and 1355 ± 160 vs. 1130 ± 211, respectively) (p < 0.05), although differences for the other nine bands were not significant. In conclusion, ultrafiltration is a suitable method for preparing urine samples for SDS-PAGE.
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