Murai J, Naka K, Shimojo N, Katakami T, Nakagishi M, Kuroki T, Okuda K
Department of Blood Transfusion, Osaka City University Medical School, Japan.
Clin Chim Acta. 1994 Apr;226(1):21-8. doi: 10.1016/0009-8981(94)90099-x.
We have developed a method for the quantitative analysis of Lewis antigens on human red blood cells (RBC) using immunofluorescence labeling and flow cytometry. Initially, Lewis a and Lewis b (Le(a) and Le(b)) antigens were labeled with monoclonal anti-Le(a) or anti-Le(b) antibodies followed by labeling with the fluorescein isothiocyanate (FITC)-conjugated second antibody. This method was not sensitive enough to identify the Lewis antigens on RBC, although the FITC method is very commonly used for antigens on white blood cells. Next, we selected the enhanced labeling technique using the avidin-biotin procedure. Biotinylated anti-mouse IgM was used for the second label and the reaction with R-phycoerythrin (RPE)-conjugated streptavidin followed to produce the fluorescence. The method was found to be effective for our objectives. From the results analyzed by the enhanced labeling technique, differences were not found in either the levels of the antigen-positive percentage and the peak mean channel of Le(a) antigens on RBC in the groups of blood type O and A (in ABO system). On the other hand, both the levels of Le(b) antigens on RBC were higher in the groups of blood type O than in those of blood type A. We found both Le(a) and Le(b) antigens on RBC from a few blood type O subjects. We conclude that enhanced labeling and flow cytometry constitute a useful technique for the determination of Lewis antigens on RBC and that this method enables the precise quantification of such antigens.
我们已经开发出一种利用免疫荧光标记和流式细胞术对人红细胞（RBC）上的Lewis抗原进行定量分析的方法。最初，用抗Le(a)或抗Le(b)单克隆抗体标记Lewis a和Lewis b（Le(a)和Le(b)）抗原，随后用异硫氰酸荧光素（FITC）偶联的二抗进行标记。尽管FITC方法常用于白细胞抗原的检测，但该方法对鉴定RBC上的Lewis抗原不够灵敏。接下来，我们选择了使用抗生物素蛋白-生物素程序的增强标记技术。生物素化的抗小鼠IgM用于二次标记，然后与藻红蛋白（RPE）偶联的抗生物素蛋白反应产生荧光。结果发现该方法对我们的目标有效。根据通过增强标记技术分析的结果，在O型血和A型血组（ABO血型系统）中，RBC上Le(a)抗原的抗原阳性百分比水平和峰值平均通道均未发现差异。另一方面，O型血组RBC上Le(b)抗原的水平均高于A型血组。我们在少数O型血受试者的RBC上发现了Le(a)和Le(b)抗原。我们得出结论，增强标记和流式细胞术是测定RBC上Lewis抗原的一种有用技术，并且该方法能够精确量化此类抗原。
