Lermi Nejla Ozirmak, Ayala Max Molina, Ruiz Sharia Hernandez, Lu Wei, Khan Khaja, Serrano Alejandra, Lubo Idania, Hamana Leticia, Tomczak Katarzyna, Barnes Sean, Dou Jinzhuang, Liang Qingnan, Raso Maria Gabriela, Tang Ximing, Jiang Mei, Sanchez-Espiridion Beatriz, Weissferdt Annikka, Heymach John, Zhang Jianjun, Sepesi Boris, Cascone Tina, Tsao Anne, Altan Mehmet, Mehran Reza, Gibbons Don, Wistuba Ignacio, Haymaker Cara, Chen Ken, Solis Soto Luisa M
Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
Authors contributed equally to this study.
Res Sq. 2025 Jan 17:rs.3.rs-5656204. doi: 10.21203/rs.3.rs-5656204/v1.
Imaging-based spatial transcriptomics (ST) is evolving rapidly as a pivotal technology in studying the biology of tumors and their associated microenvironments. However, the strengths of the commercially available ST platforms in studying spatial biology have not been systematically evaluated using rigorously controlled experiments. In this study, we used serial 5-μm sections of formalin-fixed, paraffin-embedded surgically resected lung adenocarcinoma and pleural mesothelioma tumor samples in tissue microarrays to compare the performance of the single cell ST platforms CosMx, MERFISH, and Xenium (uni/multi-modal) platforms in reference to bulk RNA sequencing, multiplex immunofluorescence, GeoMx Digital Spatial Profiler, and hematoxylin and eosin staining data for the same samples. In addition to objective assessment of automatic cell segmentation and phenotyping, we performed pixel-resolution manual evaluation of phenotyping to carry out pathologically meaningful comparison between ST platforms. Our study detailed the intricate differences between the ST platforms, revealed the importance of parameters such as tissue age and probe design in determining the data quality, and suggested reliable workflows for accurate spatial profiling and molecular discovery.
基于成像的空间转录组学（ST）作为研究肿瘤生物学及其相关微环境的关键技术正在迅速发展。然而，市售ST平台在研究空间生物学方面的优势尚未通过严格控制的实验进行系统评估。在本研究中，我们使用组织微阵列中福尔马林固定、石蜡包埋的手术切除肺腺癌和胸膜间皮瘤肿瘤样本的连续5μm切片，参照相同样本的批量RNA测序、多重免疫荧光、GeoMx数字空间分析器以及苏木精和伊红染色数据，比较单细胞ST平台CosMx、MERFISH和Xenium（单/多模态）平台的性能。除了对自动细胞分割和表型分析进行客观评估外，我们还进行了像素分辨率的手动表型评估，以便在ST平台之间进行具有病理意义的比较。我们的研究详细阐述了ST平台之间的复杂差异，揭示了组织年龄和探针设计等参数在确定数据质量方面的重要性，并提出了用于精确空间分析和分子发现的可靠工作流程。
