개에서 thin layer chromatography 박층크로마토그라피 및 ELISA를 이용한 요중의 cortisol : creatinine비 측정

Determination of urinary cortisol : creatinine ratios by sequential thin layer chromatography and ELISA in dogs

This study was conducted to evaluate the ELISA kit for measuring the level of cortisol in the urine. The CV of within-run variation and day to day variation were 0.4~2.8 and 1.8~5.7, respectively. The minimum limitation of measurement was 1ng/ml. The cross reaction was high ($CR_{50}(%)=11.4{\sim}43.2$) in prednisolone, 11-deoxycortisol, 21-deoxycortisol and predinosone. There was low and no cross reaction in other steroid. To develop the ELISA kit we measured the cortisol level in diluted urine with PBS (procedure I), extracted urine with methylene chloride (procedure II) and extracted methylene chloride-extracted urine from thin-layer chromatography (procedure III). The CV value of procedure I, II, III was 9.4~28.3%, 7.2~8.9% and 2.5~5.7%, respectively. There was significant difference between procedure I with II, and pro-cedure I with III(p < 0.01), but no difference between procedure II with III significantly(p < 0.01). The mean UCCR of urine collected through am 8 to 10 was $9.5{\pm}7.6$(0.14~28.0) in 12-month-old dog(n = 47). In this study we can measure the cortisol level in extracted urine with methylene chloride and sequential thin-layer chromatography accurately using ELISA kit.