荧光猝灭法测定冷沉淀凝血因子中的纤维蛋白原含量  

Determination of fibrinogen content in cryoprecipitated antihemophilic factor by fluorescence quenching method

作　　者：兰静 冯娜[1] 张文娟[1] 丁谨 彭鹏[1] 刘娜 曹鑫 徐华[1] LAN Jing;FENG Na;ZHANG Wenjuan;DING Jin;PENG Peng;LIU Na;CAO Xin;XU Hua(Shaanxi Blood Center,Xian 710000,China)

机构地区：[1]陕西省血液中心,陕西西安710000

出　　处：《中国输血杂志》2022年第12期1208-1212,共5页Chinese Journal of Blood Transfusion

摘　　要：目的 建立冷沉淀凝血因子中纤维蛋白原含量测定的新方法,实现纤维蛋白原含量的简单、快速、准确测定。方法 纤维蛋白原(Fib)与羊抗人纤维蛋白原(抗-Fib)能够发生特异性的结合,进而形成抗原抗体复合物。当溶液中有Fib存在时,标记有异硫氰酸荧光素(FITC)的羊抗人纤维蛋白原(FITC-抗-Fib)因免疫复合物的形成导致FITC的荧光发生猝灭。在一定浓度范围时,FITC-抗-Fib的荧光猝灭程度与Fib浓度(cFib)呈正相关。结果 当cFib为(0.007 8~0.560 0)g/L时,FITC-抗-Fib的荧光猝灭程度[(I0-I)/I0]与ln(cFib)的线性关系为(I0-I)/I0=15.53ln(cFib)+80.79 (R2=0.99);利用本法测定Fib,回收率为(96.77~102.43)%;利用本法分别测定高、中、低三个浓度的Fib,得到的日内变异系数分别为0.31%、0.56%与0.49%,日间变异系数分别为3.81%、3.06%与4.13%;利用荧光猝灭法测定冷沉淀凝血因子中的Fib含量,测得的结果与使用凝固法测得的结果无显著性差异(t=-0.075,P>0.05)。结论 本文利用纤维蛋白原与FITC-抗Fib的特异性结合成功建立了1种冷沉淀凝血因子中Fib含量测定的荧光分析新方法,该方法简单、快速,测得的结果准确、可靠。Objective To establish a new method for the determination of fibrinogen content in cryoprecipitated antihemophilic factor. Methods Fibrinogen(Fib) could bind with sheep anti-human fibrinogen(anti-Fib) specifically and further form antigen-antibody complex. When the Fib was present in the solution, the fluorescence of fluorescein isothiocyanate(FITC) labeled on the anti-Fib(FITC-anti-Fib) was quenched due to the formation of immune complex. The fluorescence quenching degree of FITC-anti-Fib was positively correlated with Fib concentration(cFib) in a certain concentration range. Results The linear relationship between fluorescence quenching degree [(I0-I)/I0] of FITC-anti-Fib and ln(cFib) was(I0-I)/I0=15.53 ln(cFib)+80.79(R~2=0.99) when the cFibwas in the range of(0.007 8-0.560 0) g/L. The recovery of Fib was(96.77-102.43) %. When the method was applied to determine Fib at high, medium, and low concentrations, the obtained intra-day variation coefficients were 0.31%, 0.56%, and 0.49%, respectively, and the inter-day variation coefficients were 3.81%, 3.06%, and 4.13%, respectively. There was no significant difference between the results measured by fluorescence quenching method and coagulation method(t=-0.075, P>0.05). Conclusion In this work, a new fluorescence method for the determination of Fib in cryoprecipitated antihemophilic factor was successfully established based on the specific combination of fib and FITC-anti-Fib. The method is simple and rapid. The obtained results were accurate and reliable by using this method to determine Fib.

关 键 词：异硫氰酸荧光素标记的羊抗人纤维原 荧光猝灭 冷沉淀凝血因子 纤维蛋白原含量 

分 类 号：R446.1[医药卫生—诊断学]