Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, State University of New York at Stony Brook, Stony Brook, New York 11794, United States.
Anal Chem. 2022 Mar 8;94(9):3922-3929. doi: 10.1021/acs.analchem.1c04970. Epub 2022 Feb 25.
Highly multiplexed analysis of biospecimens significantly advances the understanding of biological basics of diseases, but these techniques are limited by the number of multiplexity and the speed of processing. Here, we present a rapid multiplex method for quantitative detection of protein markers on brain sections with the cellular resolution. This spatial multiplex in situ tagging (MIST) technology is built upon a MIST microarray that contains millions of small microbeads carrying barcoded oligonucleotides. Using antibodies tagged with UV cleavable oligonucleotides, the distribution of protein markers on a tissue slice could be "printed" on the MIST microarray with high fidelity. The performance of this technology in detection sensitivity, resolution, and signal-to-noise level has been fully characterized by detecting brain cell markers. We showcase the codetection of 31 proteins simultaneously within 2 h, which is about 10 times faster than the other immunofluorescence-based approaches of similar multiplexity. A full set of computational toolkits was developed to segment the small regions and identify the regional differences across the entire mouse brain. This technique enables us to rapidly and conveniently detect dozens of biomarkers on a tissue specimen, and it can find broad applications in clinical pathology and disease mechanistic studies.
高多重分析生物标本显著提高了对疾病生物学基础的理解，但这些技术受到多重性数量和处理速度的限制。在这里，我们提出了一种快速多重方法，用于以细胞分辨率定量检测脑切片上的蛋白质标记物。这种空间多重原位标记 (MIST) 技术建立在一个 MIST 微阵列上，该微阵列包含数百万个带有条形码寡核苷酸的小微珠。使用带有 UV 可切割寡核苷酸标记的抗体，可以高度保真地将组织切片上的蛋白质标记物“打印”在 MIST 微阵列上。通过检测脑细胞标记物，充分表征了该技术在检测灵敏度、分辨率和信噪比水平上的性能。我们展示了在 2 小时内同时对 31 种蛋白质进行共检测，这比其他具有类似多重性的基于免疫荧光的方法快约 10 倍。还开发了一整套计算工具包，用于分割小区域并识别整个小鼠大脑的区域差异。该技术使我们能够快速方便地在组织标本上检测数十种生物标志物，它可以在临床病理学和疾病机制研究中找到广泛的应用。
