Thatcher E F, Gershwin L J
Department of Veterinary Microbiology and Immunology, School of Veterinary Medicine, University of California, Davis 95616.
Vet Immunol Immunopathol. 1988 Feb;18(1):53-66. doi: 10.1016/0165-2427(88)90036-0.
Monoclonal antibodies were produced against serum-derived bovine immunoglobulin E (IgE). Culture supernatants of hybridomas were initially screened by enzyme-linked immunosorbent assay (ELISA). Supernatant-derived antibodies were concentrated and further characterized using ELISA, reverse cutaneous anaphylaxis, immunohistochemical staining, and immunoblotting of IgE-containing samples separated by SDS-polyacrylamide gel electrophoresis (PAGE). Eight monoclonal antibodies showed specificity for bovine epsilon immunoglobulin heavy chain. Two antibodies (E2 and E32) reacted in immunoblots of SDS-PAGE of serum IgE under reducing conditions. Additionally, E2, E5, and E32 detected epsilon chain in serum separated by SDS-PAGE and then renatured. Antigen-specific IgE was detected in Western blots by E5 and E32. Immunoperoxidase staining of IgE-containing cells in mesenteric lymph node sections was detected with E5, E21 and E32. All eight antibodies produced positive reverse cutaneous anaphylaxis reactions in calf skin. All functioned well in ELISA as a plate-sensitizing reagent for quantitation of total IgE; E5 and E32 worked well as a primary antibody in antigen-specific IgE assays. These antibodies will be useful in research applications and in diagnostic assays.
制备了抗血清来源的牛免疫球蛋白E（IgE）的单克隆抗体。杂交瘤细胞的培养上清液最初通过酶联免疫吸附测定（ELISA）进行筛选。将上清液来源的抗体浓缩，并使用ELISA、反向皮肤过敏反应、免疫组织化学染色以及对经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）分离的含IgE样品进行免疫印迹分析，进一步对其特性进行鉴定。八种单克隆抗体对牛ε免疫球蛋白重链具有特异性。两种抗体（E2和E32）在还原条件下血清IgE的SDS-PAGE免疫印迹中发生反应。此外，E2、E5和E32在经SDS-PAGE分离然后复性的血清中检测到ε链。E5和E32在Western印迹中检测到抗原特异性IgE。用E5、E21和E32检测肠系膜淋巴结切片中含IgE细胞的免疫过氧化物酶染色。所有八种抗体在小牛皮肤中均产生阳性反向皮肤过敏反应。在ELISA中，所有抗体作为用于定量总IgE的平板致敏试剂均表现良好；E5和E32在抗原特异性IgE测定中作为一抗效果良好。这些抗体将在研究应用和诊断检测中发挥作用。
