三氯醋酸沉淀法测定食蟹猴体内^(125)I-rhIL-12的含量

Trichloroacetic acid precipitation method for determination of ^(125)I-rhIL-12 content in vivo of cynomolgus monkeys

目的建立三氯醋酸(Trichloroacetic acid,TCA)沉淀法测定食蟹猴体内125I-rhIL-12的含量,并进行验证及初步应用。方法采用Na125I标记结合TCA沉淀蛋白的方法测定食蟹猴体内125I-rhIL-12的含量,对建立的方法进行线性、精密度及回收率验证;将2μg/kg125I-rhIL-12单剂量皮下注射食蟹猴,分别于注射后4、8、24、32、48、72、96、120、144和168 h收集尿和粪,用建立的方法测定放射性,计算排出放射性占注入放射性的百分比。结果125I-rhIL-12的效价与未标记对照品效价无显著差异;在0.02～200 ng-当量范围内,不同组织/体液中125I-rhIL-12的加入量与总放射性及TCA沉淀放射性之间线性关系良好,变异系数(CV)多小于20%;125I-rhIL-12原形主要在TCA沉淀部分。125I-rhIL-12经皮下注射食蟹猴后,放射性主要经尿排泄,少量经粪排泄,皮下注射后168 h,可排泄完全。结论125I与rhIL-12在体内结合稳定,TCA沉淀法可用于测定食蟹猴体内125I-rhIL-12的含量。

Objective To preliminarily apply and verify trichloroacetic acid（TCA） precipitation method for determination of 125I-rhIL-12 content in vivo of cynomolgus monkeys.Methods The 125I-rhIL-12 contents in vivo of cynomolgus monkeys were by Na 125I and TCA precipitation.The developed method was verified for linearity,precision and recovery rate.Cynomolgus monkeys were injected s.c.with a single dose of 2 μg / kg 125I-rhIL-12,of which urine and stools were collected 4,8,24,32,48,72,96,120,144 and 168 h after injection,and determined for radioactivity by the developed method to calculate the percentage of output radioactivity in input radioactivity.Results The titer of 125I-rhIL-12 showed no significant difference with that unlabeled.The 125I-rhIL-12 contents in various tissues / body liquids,at a range of 0.02 ～ 200 ng-equivalent / g,showed good linear relationship to total radioactivity and TCA precipitation radioactivity,with most of CV values of less than 20%.The 125I-rhIL-12 mainly existed in TCA precipitate.The radioactivity of 125I-rhIL-12 was mainly excreted by urine,a small part of which by stool,and was completely excreted 168 h after subcutaneous injection.Conclusion 125I was combined stably with rhIL-12 in vivo,and TCA precipitation method was suitable for determination of 125I-rhIL-12 content in vivo of cynomolgus monkeys.