Carbone Teresa, Gilio Michele, Padula Maria Carmela, Tramontano Giuseppina, D'Angelo Salvatore, Pafundi Vito
IReL, Rheumatology Department of Lucania, San Carlo Hospital, Potenza, Italy; Immunopathology Laboratory, San Carlo Hospital, Potenza, Italy.
IReL, Rheumatology Department of Lucania, San Carlo Hospital, Potenza, Italy.
J Immunol Methods. 2018 May;456:67-71. doi: 10.1016/j.jim.2018.02.014. Epub 2018 Feb 19.
Indirect Immunofluorescence (IIF) is widely considered the Gold Standard for Antinuclear Antibody (ANA) screening. However, the high inter-reader variability remains the major disadvantage associated with ANA testing and the main reason for the increasing demand of the computer-aided immunofluorescence microscope. Previous studies proposed the quantification of the fluorescence intensity as an alternative for the classical end-point titer evaluation. However, the different distribution of bright/dark light linked to the nature of the self-antigen and its location in the cells result in different mean fluorescence intensities. The aim of the present study was to correlate Fluorescence Index (F.I.) with end-point titers for each well-defined ANA pattern.
Routine serum samples were screened for ANA testing on HEp-2000 cells using Immuno Concepts Image Navigator System, and positive samples were serially diluted to assign the end-point titer. A comparison between F.I. and end-point titers related to 10 different staining patterns was made.
According to our analysis, good technical performance of F.I. (97% sensitivity and 94% specificity) was found. A significant correlation between quantitative reading of F.I. and end-point titer groups was observed using Spearman's test and regression analysis. A conversion scale of F.I. in end-point titers for each recognized ANA-pattern was obtained.
The Image Navigator offers the opportunity to improve worldwide harmonization of ANA test results. In particular, digital F.I. allows quantifying ANA titers by using just one sample dilution. It could represent a valuable support for the routine laboratory and an effective tool to reduce inter- and intra-laboratory variability.
间接免疫荧光法（IIF）被广泛认为是抗核抗体（ANA）筛查的金标准。然而，不同阅片者之间的高变异性仍然是ANA检测的主要缺点，也是计算机辅助免疫荧光显微镜需求增加的主要原因。先前的研究提出将荧光强度定量作为经典终点滴度评估的替代方法。然而，与自身抗原的性质及其在细胞中的位置相关的亮/暗光的不同分布导致了不同的平均荧光强度。本研究的目的是将荧光指数（F.I.）与每种明确的ANA模式的终点滴度进行关联。
使用免疫概念图像导航系统在HEp-2000细胞上对常规血清样本进行ANA检测筛查，对阳性样本进行系列稀释以确定终点滴度。对与10种不同染色模式相关的F.I.和终点滴度进行了比较。
根据我们的分析，发现F.I.具有良好的技术性能（敏感性97%，特异性94%）。使用Spearman检验和回归分析观察到F.I.的定量读数与终点滴度组之间存在显著相关性。获得了每种公认的ANA模式的F.I.在终点滴度中的转换量表。
图像导航系统为改善全球ANA检测结果的一致性提供了机会。特别是，数字F.I.允许仅通过一次样本稀释就对ANA滴度进行定量。它可以为常规实验室提供有价值的支持，并成为减少实验室间和实验室内变异性的有效工具。
