摘要
目的建立全血三色法流式细胞术测定血小板P -选择素 ,反映体内活化血小板的表达 ,并且与常规法 (富血小板血浆法 ,PRP法 )比较该法。 方法在该法及PRP法中 ,加入三种荧光标记的单克隆抗体FITC -CD42a、PE - 6 2P及CY -CD45 ,室温避光反应 2 0min ,1%多聚甲醛固定 10min ,用PBS洗 2次 ,上机分析。 结果该法测定P -选择素的表达比常规法明显降低 (P <0 .0 5 )。 结论全血三色法测定全血中P -选择素表达特异性高 。
Objective P-selectin is known to be marker of platelet activation. We developed the technique of three-color flow cytometry determine P-selectin on the platelets whole blood and compared with the PRP method. Methods Blood was obtained from an antecubital vein through a 5ml syringe without whole blood a applying tourniquent. The first 2ml of blood was discarded. Then a 1ml blood sample was collected in 0.1ml of 3.8% sodium citrate. Within 3 min 10μl blood was added to 5μl FITC-CD42a (to label platelets), 5μl PE-CD62P (to label activation marker P-selectin) and 5μl CY-CD45 (to label WBC). After carefully mixing and 20 min of incubation at ambient temperature in the dark, the sample was fixed in formaldehyde (1%) for 10 min and washed with PBS two times. Finally the smaple was diluted to 200μl with PBS for flow cytometry. Results The results showed that the expression of P-selectin in whole blood was lower than that of the PRP. Conclusion This method would reflect very closely the physiological status of platelet activation in vivo.
出处
《上海第二医科大学学报》
CSCD
2001年第4期328-331,共4页
Acta Universitatis Medicinalis Secondae Shanghai
基金
上海市技术发展基金项目计划 (98- 4 4 1 90 63)
