CITE-seq多组学剖析弥合了单细胞分析中的RNA-蛋白距离，但主要应用于液体活检。将CITE-seq应用于临床相关的实体活检中，以表征健康组织和肿瘤微环境，是单细胞转化研究的下一重要步骤。在本研究中，根据细胞转录组特征标志物对细胞种群进行分层，用于细胞类型特异性的ridge图，从而实现了对阳性抗体信号的识别和手动阈值的设置。接下来，我们通过考虑解离效率、捕获的细胞类型多样性和恢复的表面蛋白组学来比较五种皮肤解离方案。为了评估酶解对免疫细胞群体的转录组和表位表达的影响，我们分析了带有和不带有解离的外周血单个核细胞（PBMCs）。为了进一步评估RNA-蛋白距离，我们对经过阈值处理的蛋白值进行共检测和相关性分析。最后，在一个概念验证研究中，我们使用对健康皮肤、原发性和转移性黑色素瘤队列中选定表面标志物的蛋白丰度分析，发现转移性黑色素瘤细胞上CD56表面标志物的表达，并通过多重免疫组织化学进一步证实。这项工作为临床相关的实体组织活检的生物标记物发现提供了实用指南。©2023.Springer Nature Limited.

Multi-omics profiling by CITE-seq bridges the RNA-protein gap in single-cell analysis but has been largely applied to liquid biopsies. Applying CITE-seq to clinically relevant solid biopsies to characterize healthy tissue and the tumor microenvironment is an essential next step in single-cell translational studies. In this study, gating of cell populations based on their transcriptome signatures for use in cell type-specific ridge plots allowed identification of positive antibody signals and setting of manual thresholds. Next, we compare five skin dissociation protocols by taking into account dissociation efficiency, captured cell type heterogeneity and recovered surface proteome. To assess the effect of enzymatic digestion on transcriptome and epitope expression in immune cell populations, we analyze peripheral blood mononuclear cells (PBMCs) with and without dissociation. To further assess the RNA-protein gap, RNA-protein we perform codetection and correlation analyses on thresholded protein values. Finally, in a proof-of-concept study, using protein abundance analysis on selected surface markers in a cohort of healthy skin, primary, and metastatic melanoma we identify CD56 surface marker expression on metastatic melanoma cells, which was further confirmed by multiplex immunohistochemistry. This work provides practical guidelines for processing and analysis of clinically relevant solid tissue biopsies for biomarker discovery.© 2023. Springer Nature Limited.