红细胞血型系统IgG型不规则抗体固相凝集法检测试剂盒的研发  

Anovel solid-phase agglutination reagent for detecting IgG irregular alloantibodies of red cell blood groups

作　　者：骆宏[1] 罗广平[1] 廖志坚[1] 王贞[1] 王嘉励[1] 张润青[1] 莫春妍[1] LUO Hong;LUO Guangping;LIAO Zhijian;WANG Zhen;WANG Jiali;ZHANG Runqing;MO Chunyan(Guangzhou Blood Center,the Key Medical Laboratory of Guangzhou,Guangzhou 510095,China)

机构地区：[1]广州血液中心广州市医学(血液安全)重点实验室,广东广州510095

出　　处：《中国输血杂志》2021年第3期232-235,共4页Chinese Journal of Blood Transfusion

基　　金：广州市科创委科技计划项目(201804010254)。

摘　　要：目的研发基于固相凝集技术的红细胞血型系统IgG型不规则抗体检测试剂盒并评估其性能。方法采用单克隆抗人-RBC抗体包被96孔微板,按100μL/孔加入红细胞悬液形成细胞单层,经过裂解后形成红细胞膜抗原分子单层,加入保护液冷冻干燥后制备成检测用的反应微板;用梯度倍比稀释的多克隆羊抗球蛋白试剂(IgG+C3d/4)与IgG抗-D致敏的O型红细胞反应,筛选构建最佳指示系统;用不同批次的试剂盒检测低效价的IgG抗-D和抗-E,评估试剂盒在保存期内的抗原稳定性;与微柱凝胶抗球蛋白卡、进口同类试剂盒检测不同效价的抗体以测试3者的灵敏度;与进口同类试剂盒对350例血样标本进行检测,以明确2者的检测效能是否一致。结果 20μg/mL的抗人-RBC抗体溶液能够很好的固定红细胞膜抗原分子单层;稀释16倍的多克隆羊抗球蛋白试剂与致敏红细胞构建的指示系统指示效果最佳;试剂盒中冻干的红细胞膜抗原在6个月的保存期内保持稳定;本研究的试剂盒检测灵敏度高于Capture-R Ready Screen试剂盒和抗球蛋白卡;试剂盒与Capture-R Ready Screen对血样标本检测的阳性一致性百分率为98.0%,阴性一致性百分率为99.66%,总一致性百分率为99.43%。结论成功研发了基于固相凝集技术的用于红细胞IgG型同种免疫性不规则抗体检测的试剂盒,该试剂盒具有灵敏度好、保存期长、性能稳定等优势,与进口同类试剂盒等效。Objective To develop a novel solid-phase agglutination reagent for detecting IgG irregular antibodies of red cell blood groups and evaluate its performance. Methods Monoclonal anti-RBC antibody was coated on the bottom of the microwell strips, then RBCs were bonded to the antibody and formed the monolayer by dispensing 100 μL RBC suspesion to microwell strips.RBC antigen membrane monolayer was formed by lysing RBC layer with ddH2O,then the drying medium was added to the strips and dried under reduce pressure in a vacuum dryer, thus the dried reagent microplate was obtained.Serial diluted solutions of polyclonal sheep anti-human globulin(IgG+C3 d/4)was used to react with IgG anti-D sensitized O group RBCs to select out the best indicator.Stability of membrane antigen was tested by detecting IgG anti-D and anti-E with the lowest titer by different batches of regeats. Sensitivity of the novel reagent, Capture-R Ready Screen and microcolumn gel card was carried out by detecting irregular antibodies with different titer.350 plasma samples were tested by the novel reagent and Capture-R Ready Screen to evaluate their detection ability.Results Anti-RBC solution with concentration of 20 μg/mL could fix the membrane monolayer very well on the bottom of microstrips. Sixteentimes dilution of polyclonal sheep anti-human globulin and anti-D sensitized RBCs were selected out as the best indicator.Antigen reactivity of dried RBC membrane was not weakened during the 6-monthstorage period.Sensitivity of the novel reagent was higher than Capture-R Ready Screen and microcolumn gel card. The positive consistence ratio of the novel reagent and Capture-R Ready Screen was 98.0%, the negative consistence ratio was 99.66%, and the total consistence ratio was 99.43%. Conclusion A novel solid-phase agglutination reagent with higher sensitivity and longer storage time has been developed successfully and it has an equal detection ability compared with Capture-R Ready Screen for detecting irregular alloantibodies of red cell blood groups.

关 键 词：不规则抗体 固相凝集技术 试剂盒 

分 类 号：R457.11[医药卫生—治疗学]