Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Central 6, 1-1-1 Higashi, Tsukuba, 305-8566, Japan.
Multi-Material Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 2266-98 Anagahora, Shimoshidami, Moriyama-ku, Nagoya, 463-8560, Japan.
Sci Rep. 2023 Aug 22;13(1):13644. doi: 10.1038/s41598-023-40707-3.
We previously generated three types of anti-glycan monoclonal IgM antibodies that react with certain structures on the glycans of glycosphingolipids and glycoproteins. As the nucleotide sequences for the variable regions of these IgM antibodies showed homology with those of anti-DNA antibodies deposited in public databases, we analyzed the reactivity of the anti-glycan IgM antibodies to DNA by ELISA. We found that anti-α2,6-sialyl LacNAc IgM in the supernatant of a hybridoma culture cross-reacted with DNA, and after purification of the IgM by zirconia column chromatography, the highly purified IgM showed increased cross-reactivity to DNA. As most of the contaminating bovine serum proteins in the culture supernatant were removed by the purification process, it is likely that a part of the removed components influences antibody reactivity to DNA. Purified anti-DNA antibodies prepared from lupus model NZB/W F1 and MRL/lpr mouse sera and normal human serum were then analyzed, and similar results showing increased reactivity to DNA were obtained. Furthermore, ELISA using these purified antibodies and various carbohydrate antigens showed that the antigen-binding specificity of these antibodies was altered by the purification process from serum-containing antibody preparations. Our results indicate that mammalian serum contains components that strongly influence antibody reactivity to carbohydrate antigens, including DNA.
我们先前生成了三种抗糖单克隆 IgM 抗体，它们与糖脂和糖蛋白糖链上的某些结构反应。由于这些 IgM 抗体的可变区核苷酸序列与公共数据库中保存的抗 DNA 抗体具有同源性，我们通过 ELISA 分析了抗糖 IgM 抗体对 DNA 的反应性。我们发现杂交瘤培养上清液中的抗-α2,6-唾液酸 LacNAc IgM 与 DNA 发生交叉反应，经氧化锆柱层析纯化 IgM 后，高度纯化的 IgM 对 DNA 的交叉反应性增强。由于纯化过程中去除了大部分污染的牛血清蛋白，因此很可能有一部分去除的成分影响了抗体对 DNA 的反应性。然后分析了从小鼠狼疮模型 NZB/W F1 和 MRL/lpr 血清和正常人血清中制备的纯化抗 DNA 抗体，得到了类似的结果，表明对 DNA 的反应性增强。此外，使用这些纯化抗体和各种碳水化合物抗原进行的 ELISA 分析表明，这些抗体的抗原结合特异性在从含血清的抗体制剂中进行纯化的过程中发生了改变。我们的结果表明，哺乳动物血清中含有强烈影响抗碳水化合物抗原（包括 DNA）反应性的成分。
