Immunofluorescent detection of erythrocyte sialoglycoprotein antigens on murine erythroid cells.
Immunofluorescent detection of erythrocyte sialoglycoprotein antigens on murine erythroid cells.
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DOI：
10.1083/jcb.93.3.591
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发表时间：
1982-06
期刊：
影响因子：
--
通讯作者：
Palade GE
中科院分区：
文献类型：
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作者：
Sarris AH;Palade GE
从鼠（DBA/2）红细胞血影分离的唾液酸糖蛋白级分（参见配套文章，Sarris和Palade，1982，J. Cell. 93：583-590）用于在兔中产生抗体。通过免疫-IgG（血清）-[125 I]蛋白A覆盖，发现抗体与四种唾液酸糖蛋白单体呈阳性反应（gp-2.1、gp-2.2、gp- 3.1和gp-3.2），在红细胞血影中检测到唾液酸糖蛋白（gp-2.1和gp-3.1），在溶解的鬼的SDS聚丙烯酰胺凝胶电泳图中，在gp-3.1周围尾随弥散组分（可能是大糖脂），并且在该踪迹下隐藏有少量唾液酸糖蛋白。将从免疫和非免疫兔血清中分离的IgG与异硫氰酸四甲基罗丹明结合，并通过荧光显微镜观察在小鼠寿命期间已知相互继承的三种不同红系上同源抗原的分布。在成人的外周血中，抗体只识别成熟的红细胞;它们不与血小板、单核细胞或不同类型的粒细胞发生交叉反应。在成年贫血小鼠的脾脏中，抗体与原成红细胞反应较弱，与所有类型的成红细胞反应强烈。在去核成红细胞中，抗原优先分离在新生网织红细胞的细胞膜上。在10天的胚胎中，抗原已经存在于原始有核红细胞（由卵黄囊的血胰岛产生）上，在14天的胎儿中，在所有肝成红细胞和衍生的无核红细胞上都发现了抗原。在二甲基亚砜诱导前后，Friend红白血病细胞也出现了阳性免疫反应。所有病例的非免疫血清或非免疫IgG均呈阴性反应。这些抗体具有物种特异性：它们不与人或大鼠红细胞发生交叉反应。
A sialoglycoprotein fraction isolated from murine (DBA/2) erythrocytic ghosts (see companion article, Sarris and Palade, 1982, J. Cell. Biol. 93:583-590) was used to raise antibodies in rabbits. By immune-IgG (serum)-[125I] protein A overlays, the antibodies were found to react positively with the four sialoglycoprotein monomers (gp-2.1, gp-2.2, gp- 3.1, and gp-3.2) of the original fraction, with the sialoglycoproteins detected in erythrocytic ghosts (gp-2.1 and gp-3.1), with a diffuse component (probably a macroglycolipid) trailing around gp-3.1 in SDS polyacrylamide gel electrophoretograms of solubilized ghosts, and with a minor sialoglycoprotein hidden under this trail. IgG's isolated from immune and nonimmune rabbit sera were conjugated to tetramethylrhodamine isothiocyanate and used to survey, by fluorescence microscopy, the distribution of the cognate antigens on the three different erythroid lines known to succeed each other during the life span of the mouse. In the peripheral blood of the adult, the antibodies recognized only mature erythrocytes; they did not crossreact with either platelets, monocytes, or different types of granulocytes. In the spleen of adult anemic mice, the antibodies reacted weakly with proerythroblasts and strongly with all types of erythroblasts. In enucleating erythroblasts, antigens were preferentially segregated on the cell membrane of the nascent reticulocyte. In the 10-day-old embryo, antigens were already present on the primitive nucleated erythrocytes (produced by the blood islets of the yolk sack), and in the 14-d fetus they were found on all hepatic erythroblasts and derived non-nucleated erythrocytes. A positive immunoreaction was also obtained on Friend erythroleukemic cells, before or after induction by dimethyl sulfoxide. Nonimmune serum, or nonimmune IgGs gave negative reactions in all cases. The antibodies were species-specific: they did not crossreact with either human or rat erythrocytes.