Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.
Division of Nephrology and Hypertension, Department of Medicine, Stony Brook School of Medicine, Stony Brook, NY 11794, USA.
Biosensors (Basel). 2023 Aug 27;13(9):852. doi: 10.3390/bios13090852.
High-multiplex detection of protein biomarkers across tissue regions has been an attractive spatial biology approach due to significant advantages over traditional immunohistochemistry (IHC) methods. Different from most methods, spatial multiplex in situ tagging (MIST) transfers the spatial protein expression information to an ultrahigh-density, large-scale MIST array. This technique has been optimized to reach single-cell resolution by adoption of smaller array units and 30% 8-arm PEG polymer as transfer medium. Tissue cell nuclei stained with lamin B have been clearly visualized on the MIST arrays and are colocalized with detection of nine mouse brain markers. Pseudocells defined at 10 μm in size have been used to fully profile tissue regions including cells and the intercellular space. We showcased the versatility of our technology by successfully detecting 20 marker proteins in kidney samples with the addition of five minutes atop the duration of standard immunohistochemistry protocols. Spatial MIST is amenable to iterative staining and detection on the same tissue samples. When 25 proteins were co-detected on 1 mouse brain section for each round and 5 rounds were executed, an ultrahigh multiplexity of 125 proteins was obtained for each pseudocell. With its unique abilities, this single-cell spatial MIST technology has the potential to become an important method in advanced diagnosis of complex diseases.
高多重检测蛋白质生物标志物在组织区域一直是一个有吸引力的空间生物学方法，因为它与传统的免疫组织化学（IHC）方法相比具有显著的优势。与大多数方法不同，空间多重原位标记（MIST）将空间蛋白质表达信息转移到超高密度、大规模的 MIST 阵列。通过采用较小的阵列单元和 30%的 8 臂 PEG 聚合物作为转移介质，该技术已经得到优化，达到单细胞分辨率。用 lamin B 染色的组织细胞核在 MIST 阵列上清晰可见，并与 9 种小鼠大脑标志物的检测相吻合。已使用 10 μm 大小的伪细胞来充分描绘组织区域，包括细胞和细胞间空间。我们通过在标准免疫组织化学方案的基础上增加 5 分钟，成功地在肾脏样本中检测到 20 种标记蛋白，展示了我们技术的多功能性。空间 MIST 适用于在相同的组织样本上进行迭代染色和检测。当在每一轮和 5 轮中对 1 个小鼠大脑切片共检测 25 种蛋白质时，每个伪细胞获得了高达 125 种蛋白质的超高多重性。凭借其独特的能力，这种单细胞空间 MIST 技术有可能成为复杂疾病的先进诊断的重要方法。
