SpatialBench: Comparative cross-platform benchmarking of high-resolution spatial transcriptomics using matched mouse lymphoid tissue

Solano, A. N.; Yip, R. K. H.; Wang, C.; Amann-Zalcenstein, D.; Rajasekhar, P.; Zaman, I.; Motyer, A.; Cmero, M.; Xu, Y.; Pan, Y.; Anttila, C. J. A.; Studniberg, S. I.; Hickey, P. F.; Wang, L.; Sargeant, C. J.; Ling, L.; Chen, Y.; Mishi, R. D.; Ioannidis, L. J.; Good-Jacobson, K. L.; King, H. W.; Rogers, K. L.; Hansen, D. S.; Bowden, R.; Ritchie, M. E.

doi:10.64898/2026.04.29.721531

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bioRxiv · 10.64898/2026.04.29.721531

SpatialBench: Comparative cross-platform benchmarking of high-resolution spatial transcriptomics using matched mouse lymphoid tissue

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Abstract

Spatial transcriptomics (ST) has rapidly expanded with the introduction of multiple high-resolution platforms, yet cross-platform benchmarking remains limited and largely focused on technical performance. Here we present SpatialBench, a matched multi-platform resource comprising Visium HD, Xenium and MERSCOPE data together with single-cell and single-nucleus references from a malaria-challenged wild-type and B cell-specific Tbx21 knockout mouse spleen model. In this system, loss of T-bet in B cells disrupts germinal center (GC) polarization and antibody maturation, providing a biologically grounded benchmark for technology comparison. We leveraged this system to systematically evaluate ST platform performance using technical and biological readouts. Across platforms, immune organization and Tbx21-associated programs were consistently recovered, indicating robustness of major biological signals. Platforms instead differed in the level of biological resolution accessible. Visium HD enabled transcriptome-scale GC characterization and, together with Xenium, resolved dark and light zone organization, whereas GC zonation was not resolved in MERSCOPE, consistent with differences in transcript detection sensitivity. SpatialBench provides a biologically defined reference dataset for evaluation of ST technologies, method development, computational benchmarking, and studies of GC spatial organization in lymphoid tissue.

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Solano, A. N., Yip, R. K. H., Wang, C., Amann-Zalcenstein, D., Rajasekhar, P., Zaman, I., Motyer, A., Cmero, M., Xu, Y., Pan, Y., Anttila, C. J. A., Studniberg, S. I., Hickey, P. F., Wang, L., Sargeant, C. J., Ling, L., Chen, Y., Mishi, R. D., Ioannidis, L. J., Good-Jacobson, K. L., King, H. W., Rogers, K. L., Hansen, D. S., Bowden, R., Ritchie, M. E.. 2026-05-01. SpatialBench: Comparative cross-platform benchmarking of high-resolution spatial transcriptomics using matched mouse lymphoid tissue. https://doi.org/10.64898/2026.04.29.721531

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