Institute of Burn Research, Southwest Hospital, State Key Laboratory of Trauma, Burns and Combined Injury, The Third Military Medical University, Gao Tanyan Main Street 30th, 400038, Shapingba District, Chongqing, People's Republic of China.
Protein J. 2010 Oct;29(7):475-80. doi: 10.1007/s10930-010-9275-z.
The objective is to prevent and treat injuries caused by lipopolysaccharide (LPS) from gram negative bacteria in animals and humans, we produced antibodies against LPS from egg yolk. LPS from E. coli (O111:B4) mixed with Freund's Adjuvant was used as the immunogen to immunize Roman hens. Immunized eggs were collected, and immunoglobulin Y (IgY) was purified using a water solution, salt precipitation and gel chromatography. The molecular weight and purity were determined by SDS-PAGE, the antibody titer by noncompetitive enzyme-linked immunosorbent assay (ELISA) and antibody activity against LPS by the mortality of mice intraperitoneally injected with LPS or LPS-IgY solutions. IgY against LPS showed two protein bands at 68 and 26 kDa on the gel; the antibody titer was almost 1:25,600. After incubation with LPS, IgY decreased the mortality of mice challenged with LPS. This study provided an efficient way to produce high-titer egg yolk antibodies, which could attenuate lethal effects of LPS, by immunizing hens. Furthermore, the LPS antibody was purified well using a water solution, salting-out and gel chromatography.
目的是预防和治疗动物和人类由革兰氏阴性菌脂多糖（LPS）引起的损伤，我们从蛋黄中产生了针对 LPS 的抗体。用大肠杆菌（O111:B4）的 LPS 与弗氏佐剂混合作为免疫原，对罗曼鸡进行免疫。收集免疫蛋，用水溶液、盐沉淀和凝胶层析法纯化免疫球蛋白 Y（IgY）。通过 SDS-PAGE 确定分子量和纯度，通过非竞争性酶联免疫吸附试验（ELISA）确定抗体效价，通过 LPS 或 LPS-IgY 溶液腹腔注射小鼠的死亡率确定抗体活性。针对 LPS 的 IgY 在凝胶上显示出 68 和 26 kDa 的两条蛋白带；抗体效价接近 1:25600。与 LPS 孵育后，IgY 降低了 LPS 攻击小鼠的死亡率。这项研究通过免疫母鸡为生产高效价蛋黄抗体提供了一种有效的方法，这种抗体可以减轻 LPS 的致命作用。此外，通过水溶液、盐析和凝胶层析法，LPS 抗体得到了很好的纯化。
