Guidelines for the Diagnosis and Monitoring of Paroxysmal Nocturnal Hemoglobinuria and Related Disorders by Flow Cytometry
Guidelines for the Diagnosis and Monitoring of Paroxysmal Nocturnal Hemoglobinuria and Related Disorders by Flow Cytometry
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DOI：
10.1002/cyto.b.20525
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发表时间：
2010-07-01
影响因子：
3.4
通讯作者：
Richards, Stephen J.
中科院分区：
文献类型：
--
作者：
Borowitz, Michael J.;Craig, Fiona E.;Richards, Stephen J.
背景：阵发性睡眠性血红蛋白尿症 (PNH) 是一种罕见的造血干细胞疾病，其特征是 PIGA 基因体细胞突变，导致通过糖磷脂酰肌醇 (GPI) 锚与细胞膜连接的蛋白质缺乏。虽然流式细胞术是识别 GPI 连接蛋白缺陷细胞的首选方法，因此对于诊断 PNH 是必要的，但迄今为止，尚未尝试标准化用于识别这些细胞的方法。方法：在本文件中，我们提出了一项共识工作，描述了检测 PNH 细胞的流式细胞术程序。结果：我们讨论了临床适应症并提供了有关数据解释和报告的建议，但主要关注对分析重要的分析程序。我们区分常规分析（定义为识别 1% 或更多的异常群体）和高灵敏度分析（其中检测到的 PNH 细胞少至 0.01%）。详细介绍了这两种程序所需的抗体组和门控策略。我们讨论了评估白细胞和红细胞中 PNH 群体的方法以及测量每种细胞的相对优势。我们提出了验证更复杂的高灵敏度技术所需的步骤，包括需要仔细滴定试剂和确定正常人群的背景率，并讨论可能影响解释的技术陷阱。结论：本文件既能让有兴趣开始 PNH 检测的实验室建立有效的程序，又能让经验丰富的实验室改进其技术。 (C) 2010 临床细胞计数学会
Background: Paroxysmal nocturnal hemoglobinuria (PNH) is a rare hematopoietic stem cell disorder characterized by a somatic mutation in the PIGA gene, leading to a deficiency of proteins linked to the cell membrane via glycophosphatidylinositol (GPI) anchors. While flow cytometry is the method of choice for identifying cells deficient in GPI-linked proteins and is, therefore, necessary for the diagnosis of PNH, to date there has not been an attempt to standardize the methodology used to identify these cells.Methods: In this document, we present a consensus effort that describes flow cytometric procedures for detecting PNH cells.Results: We discuss clinical indications and offer recommendations on data interpretation and reporting but mostly focus on analytical procedures important for analysis. We distinguish between routine analysis (defined as identifying an abnormal population of 1% or more) and high-sensitivity analysis (in which as few as 0.01% PNH cells are detected). Antibody panels and gating strategies necessary for both procedures are presented in detail. We discuss methods for assessing PNH populations in both white blood cells and red blood cells and the relative advantages of measuring each. We present steps needed to validate the more elaborate high-sensitivity techniques, including the need for careful titration of reagents and determination of background rates in normal populations, and discuss technical pitfalls that might affect interpretation.Conclusions: This document should both enable laboratories interested in beginning PNH testing to establish a valid procedure and allow experienced laboratories to improve their techniques. (C) 2010 Clinical Cytometry Society