MBS和戊二醛连接hPTH载体特异抗体及载体抗体的比较研究

Comparing the hPTH and carrier antibodies of MBS connect method with that of glutaraldehyde method

目的:比较间-马来酰亚胺苯甲酸-N-羟基琥珀酰亚胺酯(MBS)和戊二醛连接人甲状旁腺激素(hPTH)载体后抗体的产生效果。方法:MBS法和戊二醛一步法分别将hPTH(1-84)与载体牛血清白蛋白(BSA)连接后作为免疫原免疫家兔;间接ELISA法比较两组兔中特异性hPTH(1-84)抗体和BSA抗体的产生,并使用免疫吸收和亲和层析吸附清除BSA抗体,对清除情况进行比较。结果:MBS组抗hPTH(1-84)抗体效价总体上高于戊二醛组,抗载体BSA抗体效价低于戊二醛组;MBS组在同一时间hPTH(1-84)抗体效价高于BSA抗体,戊二醛组获取的抗血清中BSA抗体效价则明显高于hPTH(1-84)抗体。MBS组经亲和层析吸附BSA抗体被清除完全,戊二醛组经免疫吸收和亲和层析吸附仍有残留BSA抗体。结论:采用MBS法连接半抗原载体制备hPTH(1-84)抗体,降低了载体抗体的影响,提高了hPTH(1-84)抗体的特异性,可应用于hPTH药理作用和代谢途径等研究工作中。

Objective： To compare the antibodies production of m-maleimidobenzoyl-N-hydrozysuccinimide （MBS） with glutaraldehyde that connect human parathyroid hormone （hPTH） , which is in order to provide a advantageous tool to study the pharmacological mechanism and metabolism pathway of hPTH in treatment of osteoporosis. Methods： The hPTH（1-84） -BSA compounds were prepared by MBS method and one-step glutaraldehyde method. The two different compounds were used as immunogens to immunize rabbits. ELISA was used to compare the products of the hPTH（1-84） and carrier antibodies of two connectmethods. The results of the carrier antibodies which were cleaned by immune absorption method and affinity chromatography method were compared. Results： The titer of hPTH （1-84） antibodies of MBS method were more than that of glutaraldehyde method, and the titer of BSA antibodies of MBS method were fewer than that of glutaraldehyde method. The titer of hPTH （ 1-84 ） antibodies of MBS method were more than that of BSA antibodies during the immunization,and the titer of BSA antibodies of glutaraldehyde method were more than that of hPTH （1-84） antibodies. The carrier antibodies of MBS method could be easily cleaned by affinity chromatography. The carrier antibody of glutaraldehyde method could not be cleaned completely by immune absorption or affinity chromatography methods. Conclusions： Using the MBS method to parepare the hPTH poly-antibodies can reduce the influence of carrier antibody and increase the speciality of hPTH （ 1-84 ） antibody. The hPTH poly-antibodies of MBS method can be used to study the pharmacological mechanism and metabolism pathway of hPTH.