Harriman William D, Collarini Ellen J, Cromer Remy G, Dutta April, Strandh Magnus, Zhang Fen, Kauvar Lawrence M
Trellis Bioscience, 2-B Corporate Dr., South San Francisco, CA 94080, United States.
J Immunol Methods. 2009 Feb 28;341(1-2):127-34. doi: 10.1016/j.jim.2008.11.010. Epub 2008 Dec 11.
Micron scale latex beads are well established as highly biocompatible reagents. Imbibing two fluorescent dyes into the interior of the beads enables the creation of a family of combinatorially colored labels. Previous use of such beads, in flow cytometry for example, has focused on beads of approximately 5 microm diameter. We show here that 280 nm combinatorially labeled particles can be used to create ELISA-style assays in 200 microm scale virtual wells, using digital microscopy as the readout. The utility of this technique is illustrated by profiling the secreted cytokine footprints of peripheral blood mononuclear cells in a multiparametric version of the popular Elispot assay. Doing so reveals noncanonical classes of T lymphocytes. We further show that the secreting cell type can be concurrently identified by surface staining with a cell type specific antibody conjugated to the same multiplexed beads.
微米级乳胶珠作为高度生物相容性试剂已得到广泛认可。将两种荧光染料吸入珠子内部能够创建一系列组合染色标签。此前此类珠子的应用，例如在流式细胞术中，主要集中在直径约为5微米的珠子上。我们在此表明，280纳米的组合标记颗粒可用于在200微米规模的虚拟孔中创建类似酶联免疫吸附测定（ELISA）的检测方法，并使用数字显微镜进行读数。通过在流行的酶联免疫斑点试验（Elispot assay）的多参数版本中分析外周血单核细胞分泌的细胞因子足迹，说明了该技术的实用性。这样做揭示了非典型类别的T淋巴细胞。我们进一步表明，可以通过用与相同多重珠子偶联的细胞类型特异性抗体进行表面染色，同时鉴定分泌细胞类型。
