Werno Anja M, Lewis John G, George Peter M, Murdoch David R
Microbiology Unit, Canterbury Health Laboratories, Christchurch, New Zealand.
Hybridoma (Larchmt). 2012 Jun;31(3):168-75. doi: 10.1089/hyb.2011.0111.
In this study four monoclonal antibodies (MAbs) and two polyclonal antisera were raised against pneumococcal surface adhesion A (psaA), a 37 kDa cell wall protein, and were characterized. The MAbs were purified, isotyped, and epitope mapped with peptide microarrays. All monoclonals and polyclonals underwent testing in immunodot blot, Western blot, and ELISA assays to assess their specificity. All monoclonal antibodies belonged to isotype IgG1 κ. Peptide microarray mapping identified two likely epitopes, which could not be confirmed using synthetic peptides of the identified amino acid sequence. All four MAbs detected the 53 pneumococcal serotypes tested in the immunodot blot and only reacted with Streptococcus pseudopneumoniae in cross-reactivity studies by Western blot. The polyclonal antisera also recognized all tested pneumococcal serotypes and cross-reacted with S. pseudopneumoniae and Streptococcus oralis by Western blot. The MAbs cross-reacted with S. pseudopneumoniae in the ELISA. Both polyclonal antisera cross-reacted with all isolates tested in the ELISA. These antisera should be suitable to establish a diagnostic ELISA platform for pneumococcal antigen detection with polyclonal antisera as the capture antibody. The specificity of the four MAbs appears to be high as they only recognized S. pneumoniae and S. pseudopneumoniae. However, further testing of closely related streptococcal species is necessary to confirm this finding.
在本研究中，针对肺炎球菌表面黏附素A（PsaA，一种37 kDa的细胞壁蛋白）制备了四种单克隆抗体（MAb）和两种多克隆抗血清，并对其进行了表征。对单克隆抗体进行了纯化、分型，并使用肽微阵列绘制了表位图谱。所有单克隆抗体和多克隆抗体都在免疫斑点印迹、蛋白质印迹和酶联免疫吸附测定（ELISA）中进行了测试，以评估其特异性。所有单克隆抗体均属于IgG1κ同种型。肽微阵列图谱鉴定出两个可能的表位，但使用鉴定出的氨基酸序列的合成肽无法证实。所有四种单克隆抗体在免疫斑点印迹中均检测到了所测试的53种肺炎球菌血清型，并且在蛋白质印迹交叉反应研究中仅与伪肺炎链球菌发生反应。多克隆抗血清也识别所有测试的肺炎球菌血清型，并在蛋白质印迹中与伪肺炎链球菌和口腔链球菌发生交叉反应。单克隆抗体在ELISA中与伪肺炎链球菌发生交叉反应。两种多克隆抗血清在ELISA中与所有测试的分离株均发生交叉反应。这些抗血清应适合建立一个以多克隆抗血清作为捕获抗体的用于肺炎球菌抗原检测的诊断ELISA平台。这四种单克隆抗体的特异性似乎很高，因为它们仅识别肺炎链球菌和伪肺炎链球菌。然而，需要对密切相关的链球菌进行进一步测试以证实这一发现。
