A rapid pneumococcal serotyping system based on monoclonal antibodies and PCR
A rapid pneumococcal serotyping system based on monoclonal antibodies and PCR
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DOI：
10.1099/jmm.0.47549-0
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发表时间：
2008-02-01
影响因子：
3
通讯作者：
Nahm, M. H.
中科院分区：
文献类型：
--
作者：
Yu, J.;Carvalho, M. da G. S.;Nahm, M. H.
肺炎链球菌表达至少91种不同的多糖（PS）胶囊，目前可用的血清分型方法是繁琐的。我们一直在开发一种快速的肺炎球菌血清分型检测方法基于肺炎球菌裂解物抑制24种不同抗荚膜抗体与包被有24种不同PS的乳胶珠结合的能力，（血清型1、3、4、5、6A、6B、7F、9N、9V、14、18 C、19 A、19 F、23 F、2、8、10 A、11 A、12 F、15 B、17 F、20、22 F和33 F）。由于用于10种血清型（2、8、10A、11A、12F、15B、17F、20、22F和33F）的多克隆抗体具有有限的血清型特异性，我们用用于10种血清型的单克隆抗体代替它们。为了将血清型覆盖范围扩展到24种血清型之外，我们对另外5种血清型（15 A、15 C、16 F、35 B和38）的多重PCR进行了调整，以便与制备用于多珠测定的肺炎球菌裂解物一起使用。然后，我们用来自疾病控制和预防中心的157个临床分离株验证了联合检测，发现新的联合检测产生的结果与quellung反应一致。该联合检测方法具有稳健性，可用于快速鉴定大多数肺炎球菌的血清型（接近90%）。此外，试验验证研究表明血清型11A中存在血清学亚型。
Streptococcus pneumoniae expresses at least 91 different polysaccharide (PS) capsules and the currently available serotyping methods are tedious to perform. We have been developing a rapid pneumococcal serotyping assay (named the 'multibead assay') based on the capacity of pneumococcal lysates to inhibit the ability of 24 different anti-capsule antibodies to bind to latex beads coated with 24 different PSs (serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9N, 9V, 14, 18C, 19A, 19F, 23F, 2, 8, 10A, 11A, 12F, 15B, 17F, 20, 22F and 33F). Because the polyclonal antibodies used for 10 serotypes (2, 8, 10A, 11A, 12F, 15B, 17F, 20, 22F and 33F) had limited serotype specificity, we replaced them with monoclonal antibodies for the 10 serotypes. To extend the serotype coverage beyond the 24 serotypes, we have adapted multiplexed PCR for five additional serotypes (15A, 15C, 16F, 35B and 38) to be useful with the pneumococcal lysates prepared for the multibead assay. We then validated the combined assay with 157 clinical isolates from the Centers for Disease Control and Prevention and found that the new combined assay produced results that are concordant with the quellung reaction. The combined assay is robust and could be used to rapidly identify the serotypes of the majority of pneumococci (similar to 90%). In addition, the assay validation study suggests the presence of serological subtypes within serotype 11A.