Zhou C, Shao J, Wang X, Zhu H, Zhang Y, Xiong W, Shen G
Department of Immunology, Tongji Medical University, Wuhan, PR China.
Hybridoma. 1998 Jun;17(3):275-82. doi: 10.1089/hyb.1998.17.275.
By using human placental acidic isoferritin (PAF) as antigen to immunize BALB/c mice and conventional cell fusion, we have established three mouse hybridoma cell lines that secrete IgG monoclonal antibodies (MAbs) to PAF, termed as Z-2-3, Z-2-5, and Z-3-6, respectively. In ELISA, the MAbs were shown to react specifically with human PAF. We then applied the polymerase chain reaction (PCR) technique to clone variable region genes of the heavy (V(H)) and light (V(L)) chains of these MAbs, and appropriate full-length cDNA clones were obtained and characterized by nucleotide sequence analysis. V(H) and V(L) segments of anti-human PAF MAbs belong to the J558 and Vkappa19 family, respectively. The nucleotide sequence of Z-3-6 in the V(H) segment is highly homologous to that of MAb 18.1.16 except for their diversity minigenes. The light chain sequences of these MAbs show high homology with that of MAb cc92. It is implied that the D segment and the nucleotides inserted at the V(H)-D and D-J splice junctions are mostly responsible for the specificity of Z-3-6, and that the differences between the V(H) and V(L) sequences of these MAbs may determine their different affinity or recognition of different antigenic determinants.
以人胎盘酸性铁蛋白（PAF）作为抗原免疫BALB/c小鼠，并采用常规细胞融合方法，我们建立了三株分泌抗PAF IgG单克隆抗体（MAb）的小鼠杂交瘤细胞系，分别命名为Z-2-3、Z-2-5和Z-3-6。在酶联免疫吸附测定（ELISA）中，这些单克隆抗体显示能与人PAF特异性反应。随后我们应用聚合酶链反应（PCR）技术克隆这些单克隆抗体重链（V(H)）和轻链（V(L)）的可变区基因，获得了合适的全长cDNA克隆，并通过核苷酸序列分析进行了鉴定。抗人PAF单克隆抗体的V(H)和V(L)片段分别属于J558和Vkappa19家族。Z-3-6的V(H)片段核苷酸序列与单克隆抗体18.1.16的高度同源，只是它们的多样性小基因不同。这些单克隆抗体的轻链序列与单克隆抗体cc92的高度同源。这表明D片段以及在V(H)-D和D-J拼接位点插入的核苷酸主要决定了Z-3-6的特异性，并且这些单克隆抗体V(H)和V(L)序列的差异可能决定它们不同的亲和力或对不同抗原决定簇的识别。
