Tokuda M, Shimizu J, Sugiyama N, Kiryu T, Matsuoka K, Sasaki O, Fukuda K, Hatase O, Monden H
Department of Physiology, Kagawa Medical School, Kagawa, Japan.
Acta Otolaryngol Suppl. 1996;523:182-4.
Tonsils were obtained by tonsillectomy from a 33-year-old woman with IgA nephropathy. Hetero-hybridoma cells of human tonsillar B-lymphocytes with mouse myeloma cells (NS-1) were made, and cultured in HAT medium supplemented with 10% fetus bovine serum and hybridoma cloning factor. The culture medium was analyzed by Western blot analysis using anti-human IgA antibody, and both IgA1 and IgA2 were demonstrated to be produced. The specimens of the biopsied kidney tissue of IgA nephropathy were washed with 0.02 M citrate buffer (pH 3.2) to remove deposited IgA from glomerulus. The specimens were then incubated with the culture media of hybridoma cells, and immuno-fluorescence analysis using FITC-conjugated anti-human IgA antibody was performed. IgA deposit was efficiently removed by washing with citrate buffer and was recovered after incubation with the culture medium of hybridoma cells. Direct evidence of binding of IgA produced by tonsillar B-lymphocytes to the glomerular mesangium of IgA nephropathy was demonstrated.
扁桃体取自一名患有IgA肾病的33岁女性患者，通过扁桃体切除术获得。将人扁桃体B淋巴细胞与小鼠骨髓瘤细胞（NS-1）制成异源杂交瘤细胞，并在补充有10%胎牛血清和杂交瘤克隆因子的HAT培养基中培养。使用抗人IgA抗体通过蛋白质印迹分析对培养基进行分析，结果表明同时产生了IgA1和IgA2。IgA肾病活检肾组织标本用0.02M柠檬酸盐缓冲液（pH 3.2）洗涤，以去除肾小球中沉积的IgA。然后将标本与杂交瘤细胞的培养基一起孵育，并使用异硫氰酸荧光素（FITC）偶联的抗人IgA抗体进行免疫荧光分析。用柠檬酸盐缓冲液洗涤可有效去除IgA沉积物，与杂交瘤细胞培养基孵育后可回收。这证明了扁桃体B淋巴细胞产生的IgA与IgA肾病肾小球系膜结合的直接证据。
