延迟测量伊文思蓝 5′-马来酰亚胺（EMA）结合不会影响遗传性球形红细胞增多症诊断试验的结果。

Delay in the measurement of eosin-5′-maleimide (EMA) binding does not affect the test result for the diagnosis of hereditary spherocytosis.

机构信息

Department of Laboratory Diagnostics and Clinical Immunology of Developmental Age, Medical University of Warsaw Marszalkowska 24, 00-576 Warsaw, Poland.

出版信息

Clin Chem Lab Med. 2013 Apr;51(4):817-23. doi: 10.1515/cclm-2012-0240.

DOI:10.1515/cclm-2012-0240

PMID:23023797

Abstract

BACKGROUND

The eosin-5′-maleimide (EMA) binding test is a flow cytometric test widely used to detect hereditary spherocytosis (HS). EMA binds to plasma membrane proteins of red blood cells (RBCs), mainly to band 3 protein. The mean fluorescence of EMA-stained RBCs in HS patients is lower when compared with control RBCs due to the decreased amount of target proteins. EMA dye in aqueous solution is sensitive to light and high temperature. Its fluorescence can decrease when exposed to light or ambient temperatures higher than 4°C. The aim of the study was to evaluate the stability of fluorescence readings of EMA-labeled RBCs over a period of 24 h.

METHODS

The EMA test was performed in peripheral blood from 35 patients with microcytic anemia (five with HS, and 30 without HS). Peripheral blood samples were stained immediately after blood collection and analyzed using a flow cytometer at three time points: 0, after 1 and 24 h of storage at 4°C in the darkness. The results are presented as the percentage of normal control RBCs fluorescence. Flow cytometric studies were performed with Cytomics FC500 (Beckman Coulter, USA).

RESULTS

In HS patients the mean result of the test reached 66.72%±9.26% of normal controls, and in non-HS patients the EMA result was 99.48%±5.03% of normal control cells. The results of patients with HS were 66.72%±9.26%, 66.90%±10.24% and 67.86%±11.31% at 0 h, and after 1 and 24 h of storage, respectively. The results obtained from non-HS patients at time 0, after 1 and 24 h of storage reached 99.48%±5.03%, 99.49%±5.34% and 99.78%±6.13%, respectively. There was no difference between the results from each time point in samples from patients with or without HS.

CONCLUSIONS

Results of the EMA binding test do not depend on storage time of stained samples when stored at 4°C up to 24 h after staining.

摘要

背景

曙红 5′-马来酰亚胺（EMA）结合试验是一种广泛用于检测遗传性球形红细胞增多症（HS）的流式细胞术检测。EMA 结合到红细胞（RBC）的质膜蛋白上，主要结合到带 3 蛋白。与对照 RBC 相比，HS 患者的 EMA 染色 RBC 的平均荧光强度较低，这是由于靶蛋白的量减少。水溶液中的 EMA 染料对光和高温敏感。当暴露在光下或环境温度高于 4°C 时，其荧光会降低。本研究的目的是评估 EMA 标记的 RBC 荧光读数在 24 小时内的稳定性。

方法

对 35 例小细胞性贫血患者（5 例 HS，30 例非 HS）的外周血进行 EMA 检测。血液采集后立即对血液样本进行染色，并在三个时间点使用流式细胞仪进行分析：0 小时、储存 1 小时和储存 24 小时，温度为 4°C 避光。结果以正常对照 RBC 荧光的百分比表示。流式细胞术研究使用 Cytomics FC500（贝克曼库尔特，美国）进行。

结果

HS 患者的平均检测结果为正常对照的 66.72%±9.26%，非 HS 患者的 EMA 结果为正常对照细胞的 99.48%±5.03%。HS 患者的结果分别为 0 小时、1 小时和 24 小时储存后的 66.72%±9.26%、66.90%±10.24%和 67.86%±11.31%。非 HS 患者在 0 小时、1 小时和 24 小时储存后获得的结果分别为 99.48%±5.03%、99.49%±5.34%和 99.78%±6.13%。HS 或非 HS 患者样本中各时间点的结果之间无差异。