Production and characterization of monoclonal antibodies specific to lactotriaosylceramide.
Production and characterization of monoclonal antibodies specific to lactotriaosylceramide.
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DOI：
10.1093/glycob/cwq117
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发表时间：
2010-12
期刊：
影响因子：
4.3
通讯作者：
H. Nozaki;M. Yanagida;K. Koide;Kazusa Shiotani;M. Kinoshita;Y. Kobayashi;S. Watarai;Kazuo Nakamura;A. Suzuki;T. Ariga;Y. Kushi
中科院分区：
文献类型：
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作者：
H. Nozaki;M. Yanagida;K. Koide;Kazusa Shiotani;M. Kinoshita;Y. Kobayashi;S. Watarai;Kazuo Nakamura;A. Suzuki;T. Ariga;Y. Kushi
用乳糖三己糖神经酰胺（Lc（3）Cer）免疫BALB/c小鼠，建立了抗N-乙酰氨基葡萄糖β1- 3半乳糖（GlcNAcβ1-3Gal）单克隆抗体的杂交瘤细胞株。这些获得的对Lc（3）Cer特异的杂交瘤细胞是双重免疫球蛋白（IG）产生细胞，其分泌IgM和IgG分子作为抗体。已建立的mAb不仅能够与Lc（3）Cer反应，而且能够与GlcNAcβ1-3-末端鞘糖脂（GSL）反应，尽管存在分支或乳糖胺链长度以及具有末端GlcNAc残基的人转铁蛋白。通过逆转录-聚合酶链反应（RT-PCR）对克隆的IgM和IgG抗体的可变区进行比较，证实可变区决定抗体的特异性，其余氨基酸均为保守氨基酸，并证实这两种抗体均由J558和Vκ-21家族基因编码。此外，我们还利用这些单克隆抗体分析了具有GlcNAcβ1-3表位的GSL在急性白血病细胞系和小鼠胚胎组织中的表达情况，并比较了其抗原分布情况。这些单克隆抗体可用于研究GlcNAcβ1- 3Gal末端GSL在组织中的精确表达分布以及检测携带末端GlcNAcβ1-3Gal碳水化合物结构的GSL。
We have established hybridoma cell lines producing monoclonal antibodies (mAbs) directed to N-acetylglucosaminylβ1-3galactose (GlcNAcβ1-3Gal) residue by immunizing BALB/c mice with lactotriaosylceramide (Lc(3)Cer). These obtained hybridoma cells, specific to Lc(3)Cer, were dual immunoglobulin (Ig)-producing cells which secreted both IgM and IgG molecules as antibodies. The established mAbs are able to react with not only Lc(3)Cer but also GlcNAcβ1-3-terminal glycosphingolipids (GSLs) despite branching or lactosamine chain lengths and human transferrin with terminal GlcNAc residues. Comparison of the variable regions of the cloned IgM and IgG by reversed transcription-polymerase chain reaction analysis confirmed that the variable regions determine the specificity, the other amino acids are conserved, and these mAbs are encoded by J558 and Vκ-21family genes. Furthermore, we have analyzed the expression of GSLs with GlcNAcβ1-3 epitope in acute leukemia cell lines and mouse fetal tissues using these mAbs, in which antigens were distributed comparatively. These mAbs are useful for studying the precise distribution of GlcNAcβ1-3Gal-terminating GSL expression in tissues as well as for detecting GSLs carrying terminal GlcNAcβ1-3Gal carbohydrate structure.