Transcriptome analysis of archived tumors by Visium, GeoMx DSP, and Chromium reveals patient heterogeneity.

Recent advancements in probe-based, full-transcriptome technologies for FFPE tissues, such as Visium CytAssist, Chromium Flex, and GeoMx DSP, enable analysis of archival samples, facilitating the generation of data from extensive cohorts. However, these methods can be labor-intensive and costly, requiring informed selection based on research objectives. We compare these methods on FFPE tumor samples in Breast, NSCLC and DLBCL showing 1) good-quality, highly reproducible data from all methods; 2) GeoMx data containing cell mixtures despite marker-based preselection; 3) Visium and Chromium outperform GeoMx in discovering tumor heterogeneity and potential drug targets. We recommend the use of Visium and Chromium for high-throughput and discovery projects, while the manually more challenging GeoMx platform with targeted regions remains valuable for specialized questions.