生产针对莫能菌素的高特异性单克隆抗体并开发用于检测这种抗生素的微量酶联免疫吸附测定法。

Production of highly specific monoclonal antibodies to monensin and development of a microELISA to detect this antibiotic.

作者信息

Pauillac S, Halmos T, Labrousse H, Antonakis K, Avrameas S

机构信息

CNRS URA 359, Département d'Immunologie, Institut Pasteur, Paris, France.

出版信息

J Immunol Methods. 1993 Sep 15;164(2):165-73. doi: 10.1016/0022-1759(93)90309-u.

DOI:10.1016/0022-1759(93)90309-u

PMID:8370924

Abstract

Monensin, a polyether antibiotic of molecular weight 671 Da, was converted into a hemisuccinate and covalently linked to bovine serum albumin via the mixed anhydride method. Using this immunogen, polyclonal anti-monensin antibodies were raised in rabbits and monoclonal antibodies were prepared from mice. The specificity of the anti-monensin antibodies was examined by using several structural analogues as the immunogen and by performing direct binding and competitive microELISA assays on Terasaki plates. Rabbit polyclonal antibodies had a dissociation constant (KD) of 5.5 x 10(-8) M for monensin and reacted with nigericin, an antibiotic structurally related to monensin. In contrast, a mouse monoclonal antibody, 2H8, reacted only with monensin and had a much lower KD = 3 x 10(-8) M for monensin. Monoclonal antibody 2H8 was used to develop a competitive microELISA able to detect as little as 5 ng/ml of monensin in solution which corresponds to 75 pg or 110 fmol of this hapten per Terasaki well.

摘要

莫能菌素是一种分子量为671道尔顿的聚醚抗生素，它被转化为半琥珀酸酯，并通过混合酸酐法与牛血清白蛋白共价连接。使用这种免疫原，在兔子体内产生了多克隆抗莫能菌素抗体，并从小鼠体内制备了单克隆抗体。通过使用几种结构类似物作为免疫原，并在Terasaki板上进行直接结合和竞争性微酶联免疫吸附测定，对抗莫能菌素抗体的特异性进行了检测。兔多克隆抗体对莫能菌素的解离常数（KD）为5.5×10⁻⁸ M，并与尼日利亚菌素发生反应，尼日利亚菌素是一种与莫能菌素结构相关的抗生素。相比之下，小鼠单克隆抗体2H8仅与莫能菌素发生反应，对莫能菌素的KD值低得多，为3×10⁻⁸ M。单克隆抗体2H8被用于开发一种竞争性微酶联免疫吸附测定法，该方法能够检测溶液中低至5 ng/ml的莫能菌素，这相当于每个Terasaki孔中有75 pg或110 fmol的这种半抗原。