卵清蛋白基因表达放射免疫分析法

A RADIOIMMUNOASSAY METHOD FOR DETERMINATION OF OVALBUMIN GENE EXPRESSION

应用竞争结合原理建立了卵清蛋白基因表达的放射免疫分析法。采用氯胺T法标记卵清蛋白,^(125)Ⅰ-OA比活度达32～43μCi·μg^(-1),冻干后在常温下保存,有效期2～3个月;以(鸡)卵清蛋白作免疫原制备出的抗血清特异性强,与其他生物大分子无交叉反应,亲和常数为3.8×10~9L/M,50％结合率为1:1.6×10~5(最终稀释度);用双抗-PEG作分离剂沉淀免疫复合物,其方法灵敏度为0.17ng,可检范围为0.25～16ng,批内、批间变异系数分别为2.99％和8.96％。该方法能有效地直接检测类卵清蛋白含量,可作为一种特异、灵敏、简便、可靠的分析技术,应用于卵清蛋白基因表达生物基因工程研究。

A radioimmunoassay method for determination of ovalbumin-like protein was established on the principle of competition binding assay.The antigen ovalbumin (chicken) was labelled with 125I by Ch-T method.The specific activity of the iodinated antigen was 32-43μCi/μg,it was freeze-drying,stored under room tempreture and could be used for about three months.The anti-serum against ovalbumin (chicken )had no cross-reaction with the other biomacro-molecules.The affinity constant (K) of antiserum against ovalbumin-like protein was as high as 3.81×109L/M,the final dilution of antiserum was 1 : 160000.In this procedure,second antibody-PEG would be used as a precipitant to isolate the immune compound.The sensitivity was 0.17ng and the range of the determination for samples was 0.25-16 ng.The coefficient variation of intrabatch and in-terbatch were 2.99% and 8.96 % respectively.The method could successfully and directly determine the concentration of ovalbumin-like protein in samples.The radioimmunoassay method described here is specific,sensitive,reliable,simple and convenient for studying gene expression of ovalbumin-like protein.