Fang Rui, Wey Andrew, Bobbili Naveen K, Leke Rose F G, Taylor Diane Wallace, Chen John J
Office of Biostatistics and Quantitative Health Sciences, University of Hawaii John A. Burns School of Medicine, Honolulu, HI, 96813, USA.
Department of Tropical Medicine, Medical Microbiology and Pharmacology, University of Hawaii John A. Burns School of Medicine, 651 Ilalo Street, BSB 211, Honolulu, HI, 96813, USA.
Malar J. 2017 Jul 17;16(1):287. doi: 10.1186/s12936-017-1933-6.
Antibodies play an important role in immunity to malaria. Recent studies show that antibodies to multiple antigens, as well as, the overall breadth of the response are associated with protection from malaria. Yet, the variability and reliability of antibody measurements against a combination of malarial antigens using multiplex assays have not been well characterized.
A normalization procedure for reducing between-plate variation using replicates of pooled positive and negative controls was investigated. Sixty test samples (30 from malaria-positive and 30 malaria-negative individuals), together with five pooled positive-controls and two pooled negative-controls, were screened for antibody levels to 9 malarial antigens, including merozoite antigens (AMA1, EBA175, MSP1, MSP2, MSP3, MSP11, Pf41), sporozoite CSP, and pregnancy-associated VAR2CSA. The antibody levels were measured in triplicate on each of 3 plates, and the experiments were replicated on two different days by the same technician. The performance of the proposed normalization procedure was evaluated with the pooled controls for the test samples on both the linear and natural-log scales.
Compared with data on the linear scale, the natural-log transformed data were less skewed and reduced the mean-variance relationship. The proposed normalization procedure using pooled controls on the natural-log scale significantly reduced between-plate variation.
For malaria-related research that measure antibodies to multiple antigens with multiplex assays, the natural-log transformation is recommended for data analysis and use of the normalization procedure with multiple pooled controls can improve the precision of antibody measurements.
抗体在疟疾免疫中发挥重要作用。近期研究表明，针对多种抗原的抗体以及总体反应广度与预防疟疾相关。然而，使用多重检测法检测针对多种疟疾抗原组合的抗体时，其变异性和可靠性尚未得到充分表征。
研究了一种使用合并的阳性和阴性对照复制品来减少板间差异的标准化程序。对60个测试样本（30个来自疟疾阳性个体，30个来自疟疾阴性个体）以及5个合并的阳性对照和2个合并的阴性对照进行筛查，检测其针对9种疟疾抗原的抗体水平，这些抗原包括裂殖子抗原（AMA1、EBA175、MSP1、MSP2、MSP3、MSP11、Pf41）、子孢子CSP以及妊娠相关的VAR2CSA。在3个板上对抗体水平进行一式三份测量，并且由同一名技术人员在两个不同日期重复进行实验。使用测试样本的合并对照在线性和自然对数尺度上评估所提出的标准化程序的性能。
与线性尺度上的数据相比，自然对数转换后的数据偏态性较小，并且减少了均值 - 方差关系。在自然对数尺度上使用合并对照的所提出的标准化程序显著降低了板间差异。
对于使用多重检测法测量针对多种抗原的抗体的疟疾相关研究，建议在数据分析时进行自然对数转换，并且使用多个合并对照的标准化程序可以提高抗体测量的精度。
