THE USE OF MALEIMIDOCAPROYLOXYSUCCINIMIDE TO PREPARE MALARIAL PEPTIDE CARRIER IMMUNOGENS - IMMUNOGENICITY OF THE LINKING REGION
THE USE OF MALEIMIDOCAPROYLOXYSUCCINIMIDE TO PREPARE MALARIAL PEPTIDE CARRIER IMMUNOGENS - IMMUNOGENICITY OF THE LINKING REGION
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DOI：
10.1016/0022-1759(89)90224-x
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发表时间：
1989-10-24
影响因子：
2.2
通讯作者：
SAUL, A
中科院分区：
文献类型：
--
作者：
JONES, GL;EDMUNDSON, HM;SAUL, A
使用双功能试剂马来酰亚胺己酰氧基琥珀酰亚胺（MCS）将添加N末端半胱氨酸合成的疟疾肽缀合至纯化的白喉类毒素（DT）蛋白。肽与载体的摩尔比通过减法巯基滴定确定，并通过十二烷基硫酸钠（SDS）电泳确认。对于DT偶联物免疫动物血清的酶联免疫吸附（ELISA）分析，使用MCS以及基于戊二醛的偶联程序将肽偶联至牛血清白蛋白（BSA）。Western印迹分析表明，DT和BSA加合物都被针对天然序列中肽表位的单克隆抗体（Mab）识别。用DT-肽缀合物免疫的动物产生针对偶联试剂（MCS）的抗体以及白喉类毒素和肽特异性抗体。通过将血清与可溶性MCS同系物（硫代琥珀酰亚胺己酰胺（TSC））预孵育，可在很大程度上消除MCS特异性。
Malarial peptides synthesized with an added N terminal cysteine were conjugated to purified diphthria toxoid (DT) protein using the bifunctional reagent maleimidocaproyloxysuccinimide (MCS). The molar ratio of peptide to carrier was determined by subtractive sulphydryl titration and confirmed by sodium dodecyl sulphate (SDS) electrophoresis. For enzyme-linked immunoabsorbent (ELISA) analysis of sera from animals immunized with the DT conjugates, peptides were conjugated to bovine serum albumin (BSA) using MCS as well as a glutaraldehyde based coupling procedure. Western blotting analysis shows that both DT and BSA adducts were recognized by monoclonal antibodies (Mabs) directed against the peptide epitopes in the native sequences. Animals immunized with the DT-peptide conjugates produced antibodies to the coupling reagent (MCS) as well as diphtheria toxoid and peptide specific antibodies. This MCS specificity could be largely abolished by pre-incubation of sera with a soluble MCS homologoue, a thiosuccinimidocaproylamide (TSC).