Department of Pathology, Faculty of Medicine, Prince of Songkla University, Songkhla, Thailand.
Department of Pathology, Maha Chakri Sirindhorn Medical Center, Faculty of Medicine, Srinakharinwirot University, Nakhon Nayok, Thailand.
Lab Med. 2021 May 4;52(3):232-239. doi: 10.1093/labmed/lmaa065.
To validate a novel rapid molecular testing method for differentiation of homozygous hemoglobin (Hb)E and HbE/β 0-thalassemia genotypes using multiplex melt curve combined with high-resolution melt (HRM) analysis in a single test tube.
All 10 genotypes contained (β N/β N; n = 95), (β N/β 3.5-kb; n = 71), (β N/β 45-kb; n = 28), (β N/β E; n = 10), (β E/β 3.5-kb; n = 6), (β E/β 45-kb; n = 4), (β E/β 41/42; n = 28), (β E/β 17; n = 9), (β E/β IVSI#1; n = 6), and (β E/β E; n = 76) were recruited for validation. A proposed strategy for rapid differentiation of β 0-thalassemia/HbE disease and homozygous Hb E in specimens with HbE greater than 80% and variable HbF levels was demonstrated.
In the validation method, all genotypes showed 100% concordance, compared with the conventional reverse dot blot (RDB) and gap-polymerase chain reaction (PCR) methods.
Our newly developed method could be useful in routine laboratory settings. The method is rapid, simple, and cost effective; does not require a post-PCR step; and can be applied in routine settings.
验证一种新的快速分子检测方法，用于在单个试管中使用多重熔解曲线结合高分辨率熔解（HRM）分析区分纯合血红蛋白（Hb）E 和 HbE/β⁰-地中海贫血基因型。
所有 10 种基因型均包含（βⁿ/βⁿ；n = 95）、（βⁿ/β 3.5-kb；n = 71）、（βⁿ/β 45-kb；n = 28）、（βⁿ/β E；n = 10）、（β E/β 3.5-kb；n = 6）、（β E/β 45-kb；n = 4）、（β E/β 41/42；n = 28）、（β E/β 17；n = 9）、（β E/β IVSI#1；n = 6）和（β E/β E；n = 76）。提出了一种快速区分 HbE 大于 80%和可变 HbF 水平标本中β⁰-地中海贫血/HbE 疾病和纯合 Hb E 的策略。
在验证方法中，与传统的反向斑点印迹（RDB）和Gap-聚合酶链反应（PCR）方法相比，所有基因型均显示 100%一致性。
我们新开发的方法可在常规实验室环境中使用。该方法快速、简单且具有成本效益；不需要 PCR 后处理；并且可以在常规环境中应用。
