DNA工作站在HLA测序分型中的应用

目的 建立从全血样本中高通量提取基因组DNA的方法,以应用于HLA常规测序分型.方法 采用DNA工作站及2 ml深孔板,从400份全血样本中提取基因组DNA.用紫外分光光度仪测定其浓度和A<,260>/A<,280>值,并采用琼脂糖电泳检测DNA的完整性.扩增产物经纯化后作为测序模板,产物经酒精/EDTA/NaOAc法纯化,用ABI Prism<'TM>3730测序仪电泳检测.相关的分析软件分析HLA基因型.结果 使用400μL全血中400份样本的DNA产量平均为(3.217±0.715)μg,A<,260>/A<,280>值平均为1.710±0.103;琼脂糖电泳表明DNA的分子量均大于15×10<'3>;HLA-A、HLA-B和HLA-DRB1座位序列碱基峰高均在2000以上,测序评分在75分以上.48份HLA-A、B和DRB1基因型已知的室内质控样本,经本文的方法提取DNA并进行HLA基因分型,经盲检质控检测,所得到的结果与已知的HLA基因型相一致.结论 采用本方法所提取的DNA能稳定、可靠地应用于骨髓捐献者样本的HLA-A、B和DRB1座位测序分型,具有快速、高通量化的特点和广泛的应用前景.     

A rapid ABO and RhD test demonstrates high fidelity to blood bank testing for RhD typing

Background

The rapid provision of blood products is life-saving for patients with massive hemorrhage. Ideally, RhD-negative blood products would be supplied to a woman of childbearing potential whose Rh type is unknown due to the risk of D-alloimmunization and the potential for hemolytic disease of the fetus and newborn to occur if RhD-positive blood products are transfused. Therefore, there is a need for a test that rapidly determines her RhD type. This study compared the RhD type determined using a rapid ABO and RhD test to the RhD type determined by an immunohematology reference laboratory.

Methods

After receiving ethics review board approval, 200 random, unique, deidentified patient samples that had undergone routine pretransfusion testing in an immunohematology reference laboratory using column agglutination technology were collected and tested using a rapid ABO and RhD test (Eldoncard Home kit 2511). The RhD typing results from these two methods were compared to determine the accuracy of the rapid ABO and RhD test.

Results

The rapid ABO and RhD test produced results that were concordant with the transfusion service's results in 199/200 (99.5%) of cases, with a negative predictive value of 98.2% and 99.3% sensitivity. The single outlier was likely an RhD variant due to its serological characteristics.

Discussion

These data indicate that this rapid ABO and RhD test could be used for the rapid determination of a patient's RhD type, perhaps even in the emergency department, which could guide the selection of blood products provided during their resuscitation.     

抗-D单克隆抗体的研制及其应用

目的 研制用于Rh(D)分型的抗－D单克隆抗体。方法 以EB病毒（EBV）转化能分泌抗－D抗体的Rh阴性供者的B淋巴细胞，用酶处理RhD^ 、O型红细胞富集抗－D分泌细胞，并用有限稀释法克隆。采用一步酶法，于96孔微量板中分别用阳性细胞培养上清液和人源多克隆抗－D血清及进口的单克隆抗－D血清平行检测1000名献血员的Rh(D)血型。结果 建立了3株能分泌抗－D抗体的人B淋巴细胞株，其中1株分泌的为IgM，2株为IgG；抗体效价在64－128之间；3种试剂检测Rh(D）血型的符合率为100%，3名献血员均为RhD^-。结论 抗－D单克隆抗体可用于Rh(D)分型。