α-Gal表达的无创快速半定量检测

A Prompt Method to Quantitative Assay of α-Gal on Pig Cell Surface without Injury

目的 建立无创快速半定量检测猪血细胞表面α- Gal表达的方法。方法 取版纳微型猪近交系(BMI)及四川白猪(SWP)新鲜血,肝素抗凝,180 0 r/ m in离心,取白细胞富集层,利用激光共聚焦(confocal)及流式细胞术(FCM)检测猪血细胞与FITC- BSIB4的结合。结果 以人B型血细胞为阴性对照,小鼠骨髓瘤SP2 / 0细胞为阳性对照,两种中国猪种各血细胞α- Gal表达均为阳性,BMI粒细胞表面α- Gal约为9.16×10 5个位点/细胞,单个核细胞1.37×10 5个位点/细胞;SWP粒细胞1.16×10 6 个位点/细胞,单个核细胞2 .4 5×10 5个位点/细胞。两种中国猪种同一种类细胞α- Gal无明显差异(P>0 .0 5 ) ,同一猪种不同个体相同细胞α- Gal表达不尽相同;同一个体不同血细胞α- Gal表达强度为:血小板>粒细胞>单个核细胞>红细胞。结论 本研究建立的无创快速半定量检测猪细胞表面α- Gal的方法可用于现场猪种的大规模筛查。

Objective To develop a method for quantitative assay of α-Gal on pig cell surface. Methods After centrifugation of the anticoagulated blood samples, the white blood cell layer was collected for addition of FITC-BSIB4 and analysis using FCM as well as Laser Scan Confocal Microscope(Bio-Rad). Results With human blood group B as negative control, and mouse myeloma SP2/0 cells as positive control. the peripheral blood cells of two Chinese pigs were all positive, there were about 9.16×105 alphα-Galactosyl epitopes expressed on the cell-membrane of granulocytes 1.37×105 on the monocytes of Banna Minipig Inbred Line (BMI), and 1.16×106 on the granulocyte and 2.45×105 on the monocytes of Sichuan White Pig (SWP). The levels of alphα-Galactosyl expression on the surface of blood cells in BMI and SWP differed in various cells, the sequence from high to lower expression levels being on the: platelets, granulocytes, monocytes, erthrocytes in each species of pig. Conclusion The method is useful for quantitative assay of α-Gal on pig cell surface, and the assay may be used for α-Gal for on-site large-scaled screening of living pigs.