PDF(Pubmed)
Abstract：
Patients who carry Rhesus (RH) blood group variants may develop Rh alloantibodies requiring matched red blood cell transfusions. Serologic reagents for Rh variants often fail to specifically identify variant Rh antigens and are in limited supply. Therefore, red blood cell genotyping assays are essential for managing transfusions in patients with clinically relevant Rh variants. Well-characterized DNA reference reagents are needed to ensure quality and accuracy of the molecular tests. Eight lyophilized DNA reference reagents, representing 21 polymorphisms in RHD and RHCE, were produced from an existing repository of immortalized B-lymphoblastoid cell lines at the Center for Biologics Evaluation and Research/US Food and Drug Administration. The material was validated through an international collaborative study involving 17 laboratories that evaluated each DNA candidate using molecular assays to characterize RHD and RHCE alleles, including commercial platforms and laboratory-developed testing, such as Sanger sequencing, next-generation sequencing, and third-generation sequencing. The genotyping results showed 99.4% agreement with the expected results for the target RH polymorphisms and 87.9% for RH allele agreement. Most of the discordant RH alleles results were explained by a limited polymorphism coverage in some genotyping methods. Results of stability and accelerated degradation studies support the suitability of these reagents for use as reference standards. The collaborative study results demonstrate the qualification of these eight DNA reagents for use as reference standards for RH blood group genotyping assay development and analytical validation.
摘要：
携带RH血型变异的患者可能会产生Rh同种抗体，需要进行匹配的红细胞输血。Rh变体的血清学试剂通常不能特异性地鉴定变体Rh抗原并且供应有限。因此,红细胞基因分型检测对于治疗具有临床相关Rh变异的患者输血至关重要.需要充分表征的DNA参考试剂以确保分子测试的质量和准确性。八种冻干DNA参考试剂，代表RHD和RHCE中的21个多态性，是从CBER/美国FDA的永生化B淋巴母细胞细胞系的现有储存库产生的。该材料通过一项国际合作研究进行了验证，该研究涉及17个实验室，这些实验室使用分子测定法评估了每个DNA候选基因，以表征RHD和RHCE等位基因，包括商业平台和实验室开发的测试，如Sanger测序。下一代测序，和第三代测序。基因分型结果显示，目标RH多态性与预期结果符合99.4％，RH等位基因符合87.9％。大多数不一致的RH等位基因结果是由某些基因分型方法中有限的多态性覆盖率解释的。稳定性和加速降解研究的结果支持这些试剂用作参考标准的适用性。合作研究结果证明了这八种DNA试剂用作RH血型基因分型测定开发和分析验证的参考标准的资格。
