Golsaz-Shirazi Forough, Amiri Mohammad Mehdi, Bahadori Motahareh, Bayat Ali Ahmad, Mohammadi Hamed, Farid Samira, Maddah Mahshid, Khoshnoodi Jalal, Zarnani Amir-Hassan, Jeddi-Tehrani Mahmood, Shokri Fazel
1 Department of Immunology, School of Public Health, Tehran University of Medical Sciences , Tehran, Iran .
2 Monoclonal Antibody Research Center, Avicenna Research Institute , ACECR, Tehran, Iran .
Viral Immunol. 2015 Oct;28(8):425-33. doi: 10.1089/vim.2015.0023.
Hepatitis B virus (HBV) surface antigen (HBsAg) induces a vigorous neutralizing antibody response, which causes effective protection against HBV infection. Little is known about the profile of variable region genes of immunoglobuline heavy (VH) and light (VL) chains rearranged in anti-HBs antibodies, and also the possible association of this profile with specificity pattern of these antibodies to mutant forms of HBsAg.
The present study determined the nucleotide sequence of VH and VL genes of mouse monoclonal antibodies (MAbs) generated against HBsAg.
Hybridoma clones secreting anti-HBsAg MAbs were developed from hyperimmunized Balb/c mice. VH and VL gene sequences of all MAbs were determined by amplifying the genes using a panel of VH and VL family specific primers by reverse transcription polymerase chain reaction. The reactivity pattern of anti-HBs MAbs with different mutant forms of HBsAg was evaluated by enzyme-linked immunosorbent assay, and then the profile of antigen specificity and its association to VH/VL family expression was analyzed.
Twenty-three murine hybridomas producing anti-HBs MAbs were generated. Nucleotide sequence analysis revealed that heavy chains of these MAbs were encoded by IGHV genes from the HV1 (52%), HV6 (22%), HV5 (17%), and HV3 (9%) families in combination with IGHJ2 (57%), HJ1 (26%), and HJ4 (17%). Besides, 56% of MAbs used IGHD1 genes in their VDJ rearrangements. Concerning the IGKV gene, 26% and 22% of clones used KV4 and KV10 gene families, while the rest of the clones used KV8, KV6, KV1, KV12, and KV14 gene families. Besides, the IGKJ2 gene was the most represented KJ gene (43%). No association was found between the specificity pattern of MAbs to mutant forms of HBsAg with their preferential V, D, and J genes usage for most of MAbs.
The data suggest that heavy chains of anti-HBs MAbs preferentially use genes derived from the IGHV1, IGHV6, IGHJ2, and IGHD1 families. In contrast to heavy chains, which predominantly use four families of IGHV genes, light chains use more diverse IGKV gene families.
乙型肝炎病毒（HBV）表面抗原（HBsAg）可诱导强烈的中和抗体反应，从而有效预防HBV感染。关于抗-HBs抗体中免疫球蛋白重链（VH）和轻链（VL）可变区基因重排的情况，以及该情况与这些抗体对HBsAg突变形式的特异性模式之间的可能关联，目前所知甚少。
本研究确定了针对HBsAg产生的小鼠单克隆抗体（MAb）的VH和VL基因的核苷酸序列。
从经超免疫的Balb/c小鼠中培养分泌抗-HBsAg MAb的杂交瘤克隆。使用一组VH和VL家族特异性引物通过逆转录聚合酶链反应扩增基因，从而确定所有MAb的VH和VL基因序列。通过酶联免疫吸附测定评估抗-HBs MAb与不同突变形式HBsAg的反应模式，然后分析抗原特异性概况及其与VH/VL家族表达的关联。
产生了23个分泌抗-HBs MAb的小鼠杂交瘤。核苷酸序列分析显示，这些MAb的重链由来自HV1（52%）、HV6（22%）、HV5（17%）和HV3（9%）家族的IGHV基因编码，并与IGHJ2（57%）、HJ1（26%）和HJ4（17%）组合。此外，56%的MAb在其VDJ重排中使用IGHD1基因。关于IGKV基因，26%和22%的克隆使用KV4和KV10基因家族，其余克隆使用KV8、KV6、KV1、KV12和KV14基因家族。此外，IGKJ2基因是最具代表性的KJ基因（43%）。对于大多数MAb，未发现其对HBsAg突变形式的特异性模式与其优先使用的V、D和J基因之间存在关联。
数据表明，抗-HBs MAb的重链优先使用源自IGHV1、IGHV6、IGHJ2和IGHD1家族的基因。与主要使用四个IGHV基因家族的重链不同，轻链使用更多样化的IGKV基因家族。
