Blood Center of Zhejiang Province, Jianye Road 789, Hangzhou, 310052, Zhejiang, China.
Key Laboratory of Blood Safety Research, Zhejiang Province, Jianye Road 789, Hangzhou, 310052, Zhejiang, China.
J Transl Med. 2019 Aug 5;17(1):249. doi: 10.1186/s12967-019-2002-4.
Alloantibodies against human platelet antigens (HPAs) and human leukocyte antigen (HLA) are implicated in several immune-mediated platelet disorders. Detection of these antibodies is crucial in the diagnosis and management of these disorders. The aim of this study was to establish a novel method to simultaneously detect HPA-1, HPA-2, HPA-3, HPA-5 and HLA antibodies with Luminex microbeads technology.
Monoclonal antibodies specific for platelet glycoproteins and HLA class I molecules were separately coupled to the Luminex microbeads. We validated specificity of the Luminex platform using the following antibodies: anti-HPA-1a, anti-HPA-2b, anti-HPA-3a, anti-HPA-5a, and anti-HLA positive samples. Sensitivity was evaluated by a serial dilution (from neat to 1/1024) using the following antibodies: anti-HPA-1a, anti-HPA-3a standard sera, and anti-HPA-5a positive serum. Serum samples were collected from 36 neonatal alloimmune thrombocytopenia (NAIT) patients suspected of having HPA or HLA antibodies and 8 samples from ISBT platelet workshop were tested using the Luminex assay.
The Luminex assay detected all antibodies tested from the known samples. The sensitivities of the Luminex assay detecting anti-HPA-1a, anti-HPA-3a, and anti-HPA-5a were 1:512, 1:64, and 1:128, respectively. The sensitivity of Luminex assay was higher than monoclonal antibody immobilization of platelet antigen method (MAIPA). No cross-reactivity was observed in the samples containing multi-platelet antibodies or mixture antibodies against HPA and HLA. The results of 44 samples with platelet disorders were consistent with those of the same samples processed with the MAIPA assay.
Luminex microbeads coupled with monoclonal antibodies could be successfully used to detect HPA and HLA antibodies simultaneously, especially with high sensitivity in detecting HPA antibodies.
针对人类血小板抗原（HPAs）和人类白细胞抗原（HLA）的同种抗体与几种免疫介导的血小板疾病有关。这些抗体的检测对这些疾病的诊断和治疗至关重要。本研究旨在建立一种新的方法，使用Luminex 微珠技术同时检测 HPA-1、HPA-2、HPA-3、HPA-5 和 HLA 抗体。
分别将针对血小板糖蛋白和 HLA Ⅰ类分子的单克隆抗体偶联到 Luminex 微珠上。我们使用以下抗体验证了 Luminex 平台的特异性：抗-HPA-1a、抗-HPA-2b、抗-HPA-3a、抗-HPA-5a 和抗-HLA 阳性样本。通过使用以下抗体进行系列稀释（从 1:1024 到 1:1024）评估敏感性：抗-HPA-1a、抗-HPA-3a 标准血清和抗-HPA-5a 阳性血清。使用该 Luminex 检测法对 36 例疑似具有 HPA 或 HLA 抗体的新生儿同种免疫性血小板减少症（NAIT）患者的血清样本和 8 例来自 ISBT 血小板工作组的样本进行了检测。
Luminex 检测法检测到了已知样本中的所有抗体。Luminex 检测法检测抗-HPA-1a、抗-HPA-3a 和抗-HPA-5a 的灵敏度分别为 1:512、1:64 和 1:128。Luminex 检测法的灵敏度高于单克隆抗体血小板抗原固定化法（MAIPA）。在含有多血小板抗体或针对 HPA 和 HLA 的混合抗体的样本中未观察到交叉反应。44 例血小板疾病样本的结果与用 MAIPA 检测法处理的相同样本的结果一致。
Luminex 微珠与单克隆抗体偶联可成功用于同时检测 HPA 和 HLA 抗体，特别是在检测 HPA 抗体方面具有高灵敏度。
