Zimmermann U, Love-Homan L, Gessner P, Clark D, Klöck G, Johlin F C, Neil G A
Lehrstuhl für Biotechnologie, Biozentrum, Universität Würzburg, Germany.
Hum Antibodies Hybridomas. 1995;6(2):77-80.
We have generated a human monoclonal antibody with binding specificity for hepatitis C virus (HCV)-specific peptides using peripheral blood lymphocytes isolated from a HCV antibody positive patient. The B-lymphocytes were stimulated with lipopolysaccharide (LPS) for 72 hours prior to the fusion. A recently described high efficiency hypo-osmolar electrofusion technique was employed, allowing generation of a large number of human hybridomas. The hybridomas were screened for human immunoglobulin and HCV-specific peptide binding by EIA. A single HCV-positive clone, JRA1, was detected and sub-cloned. Isotype analysis showed it to secrete an IgM lambda monoclonal antibody. The antibody was positive on both first and second generation HCV antibody analysis. This study confirms that viable pathogen-specific B-cells may be recovered from the peripheral blood. Although such cells are likely to be relatively uncommon in the circulating B-cell pool, they may be successfully immortalized by high efficiency electrofusion techniques. This technique might be valuable for the generation of human monoclonal antibodies with specificity for other human pathogens.
我们使用从一名丙型肝炎病毒（HCV）抗体阳性患者分离的外周血淋巴细胞，制备了一种对HCV特异性肽具有结合特异性的人单克隆抗体。在融合前，用脂多糖（LPS）刺激B淋巴细胞72小时。采用了一种最近描述的高效低渗电融合技术，从而能够产生大量的人杂交瘤。通过酶免疫测定法（EIA）筛选杂交瘤的人免疫球蛋白和HCV特异性肽结合情况。检测到一个HCV阳性克隆JRA1并进行了亚克隆。亚型分析表明它分泌一种IgMλ单克隆抗体。该抗体在第一代和第二代HCV抗体分析中均呈阳性。这项研究证实，可从外周血中回收有活力的病原体特异性B细胞。尽管这类细胞在循环B细胞库中可能相对罕见，但通过高效电融合技术它们可能成功永生化。该技术对于制备对其他人类病原体具有特异性的人单克隆抗体可能具有重要价值。
