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首页> 外文期刊>Infection and immunity >Platelet receptors for the Streptococcus sanguis adhesin and aggregation-associated antigens are distinguished by anti-idiotypical monoclonal antibodies.
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Platelet receptors for the Streptococcus sanguis adhesin and aggregation-associated antigens are distinguished by anti-idiotypical monoclonal antibodies.

机译：血链球菌粘附素和聚集相关抗原的血小板受体通过抗独特型单克隆抗体来区分。

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Platelets aggregate in response to an adhesin and the platelet aggregation-associated protein (PAAP) expressed on the cell surfaces of certain strains of Streptococcus sanguis. We sought to identify the corresponding PAAP receptor and accessory adhesin binding sites on platelets. Since the adhesion(s) of S. sanguis for platelets has not been characterized, an anti-idiotype (anti-id) murine monoclonal antibody (MAb2) strategy was developed. First, MAb1s that distinguished the adhesin and PAAP antigens on the surface of S. sanguis I 133-79 were selected. Fab fragments of MAb1.2 (immunoglobulin G2b [IgG2b]; 70 pmol) reacted with 5 x 10(7) cells of S. sanguis to completely inhibit the aggregation of human platelets in plasma. Under similar conditions, MAb1.1 (IgG1) inhibited the adhesion of S. sanguis cells to platelets by a maximum of 34%, with a comparatively small effect on platelet aggregation. Together, these two MAb1s inhibited S. sanguis-platelet adhesion by 63%. In Western immunoblots, both MAb1s reacted with S. sanguis 133-79 87- and 150-kDa surface proteins and MAb1.2 also reacted with purified type I collagen. The hybridomas producing MAb1.1 and MAb1.2 were then injected into BALB/c mice. Enlarged spleens were harvested, and a panel of MAb2 hybridomas was prepared. To identify anti-ids against the specific MAb1s, the MAb2 panel was screened by enzyme-linked immunosorbent assay for reaction with rabbit polyclonal IgG antibodies against the 87- and 150-kDa antigens. The reactions between the specific rabbit antibodies and anti-ids were inhibited by the 87- and 150-kDa antigens. When preincubated with platelets, MAb2.1 (counterpart of MAb1.1) inhibited adhesion to platelets maximally by 46% and MAb2.2 (anti-MAb1.2) inhibited adhesion to platelets maximally by 35%. Together, both MAb2s inhibited the adhesion of S. sanguis to platelets by 81%. MAb2.2 also inhibited induction of platelet aggregation. MAb2.2 immunoprecipitated a biotinylated platelet membrane antigen of 170 kDa (unreduced); MAb2.1 precipitated membrane antigens of 175- and 230-kDa (unreduced). Therefore, platelet binding sites and the receptor for the S. sanguis adhesin and PAAP, respectively, are distinguished by the anti-id MAb2s.
机译：血小板对黏附素和某些血链球菌菌株的细胞表面表达的血小板聚集相关蛋白（PAAP）产生反应。我们试图鉴定血小板上相应的PAAP受体和辅助粘附素结合位点。由于尚无桑氏葡萄球菌对血小板的粘附特性，因此开发了一种抗独特型（anti-id）鼠单克隆抗体（MAb2）策略。首先，选择能够区分血红链霉菌I 133-79表面的粘附素和PAAP抗原的MAb1。 MAb1.2的Fab片段（免疫球蛋白G2b [IgG2b]; 70 pmol）与血红葡萄球菌的5 x 10（7）细胞反应，以完全抑制血浆中人血小板的聚集。在相似的条件下，MAb1.1（IgG1）最多可抑制血红葡萄球菌与血小板的黏附力达34％，对血小板聚集的影响相对较小。这两个MAb1共同抑制血丝链霉菌血小板粘附的作用达63％。在Western免疫印迹中，两种MAb1s均与血红链霉菌133-79 87-和150-kDa表面蛋白反应，MAb1.2也与纯化的I型胶原蛋白反应。然后将产生MAb1.1和MAb1.2的杂交瘤注射到BALB / c小鼠中。收获扩大的脾脏，并制备一组MAb2杂交瘤。为了鉴定针对特定MAb1的抗ID，通过酶联免疫吸附试验筛选MAb2面板，以与针对87 kDa和150 kDa抗原的兔多克隆IgG抗体反应。特异性兔抗体和抗-ids之间的反应被87-和150-kDa抗原抑制。与血小板预温育时，MAb2.1（MAb1.1的对应物）最大程度地抑制了46％的血小板粘附，而MAb2.2（抗MAb1.2）最大程度地抑制了35％的血小板粘附。两种MAb2共同抑制血红链球菌对血小板的黏附作用达81％。 MAb2.2也抑制了血小板聚集的诱导。 MAb2.2免疫沉淀170 kDa的生物素化血小板膜抗原（未还原）; MAb2.1沉淀了175-kDa和230-kDa的膜抗原（未还原）。因此，抗SID MAb2s分别区分了血红细胞粘附素和PAAP的血小板结合位点和受体。

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