Fluorescence intensity normalisation: correcting for time effects in large-scale flow cytometric analysis.
Fluorescence intensity normalisation: correcting for time effects in large-scale flow cytometric analysis.
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DOI：
10.1155/2009/476106
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发表时间：
2009
影响因子：
--
通讯作者：
Plagnol V
中科院分区：
文献类型：
--
作者：
Dendrou CA;Fung E;Esposito L;Todd JA;Wicker LS;Plagnol V
解释全基因组关联研究结果的下一步是将分子数量性状与疾病相关等位基因联系起来。为此，研究人员将疾病风险等位基因与基因表达数量性状位点（eQTL）联系起来。然而，mRNA水平的基因表达只是一个中间性状，流式细胞术分析可以使用新鲜获得的样品同时提供多个细胞亚群中更多下游和生物学上有价值的蛋白质水平信息。由于流式细胞术的通量目前是有限的，实验可能需要跨越几周或几个月，以获得足够的样本量来证明遗传关联。因此，需要标准化方法来控制技术变异性，并在较长时间内比较流式细胞术数据。我们展示了使用标准化荧光球如何提高CD 4+记忆T细胞上细胞表面CD 25-APC平均荧光强度表型的可重复性。我们研究了两种类型的归一化珠：广谱和光谱匹配。最后，我们提出了两种替代的归一化程序，可用于在没有归一化珠。
A next step to interpret the findings generated by genome-wide association studies is to associate molecular quantitative traits with disease-associated alleles. To this end, researchers are linking disease risk alleles with gene expression quantitative trait loci (eQTL). However, gene expression at the mRNA level is only an intermediate trait and flow cytometry analysis can provide more downstream and biologically valuable protein level information in multiple cell subsets simultaneously using freshly obtained samples. Because the throughput of flow cytometry is currently limited, experiments may need to span over several weeks or months to obtain a sufficient sample size to demonstrate genetic association. Therefore, normalisation methods are needed to control for technical variability and compare flow cytometry data over an extended period of time. We show how the use of normalising fluorospheres improves the repeatability of a cell surface CD25-APC mean fluorescence intensity phenotype on CD4+ memory T cells. We investigate two types of normalising beads: broad spectrum and spectrum matched. Lastly, we propose two alternative normalisation procedures that are usable in the absence of normalising beads.