Xenium In Situ Gene Expression
Six sequential 10 um fresh frozen sections of one 34-year-old case (Table S1) of human hippocampus were placed onto a Xenium slide and processed at –20°C. The tissue was fixed and permeabilized as described in the Xenium Fixation and Permeabilization Protocol (Demonstrated protocol CG000581). A customized panel of probes targeting 316 genes were added to the tissue (Table S2). The probes were hybridized to the target RNA, ligated, and enzymatically amplified generating multiple copies for each RNA target, as described in Probe Hybridization, Ligation and Amplification user guide (User guide CG000582). The Xenium slides were, then, loaded for imaging and analysis on the Xenium Analyzer instrument, followed by decoding, according to the user guide (User guide CG000584, RRID SCR_023910). Following the analysis guidelines of 10x Genomics, negative controls spatial map was considered and valued according to the expectations as described in https://www.10xgenomics.com/support/software/xenium-onboard-analysis/latest/analysis/xoa-output-analysis-summary (10x Genomics Xenium Onboard Analysis, RRID SCR_026158). Instrument software version 1.4.2.0 and software analysis version 1.4.0.6 were used (10x Genomics Xenium Explorer, RRID SCR_025847).

Xenium In Situ Gene Expression
Six sequential 10 um fresh frozen sections of one 34-year-old case (Table S1) of human hippocampus were placed onto a Xenium slide and processed at –20°C. The tissue was fixed and permeabilized as described in the Xenium Fixation and Permeabilization Protocol (Demonstrated protocol CG000581). A customized panel of probes targeting 316 genes were added to the tissue (Table S2). The probes were hybridized to the target RNA, ligated, and enzymatically amplified generating multiple copies for each RNA target, as described in Probe Hybridization, Ligation and Amplification user guide (User guide CG000582). The Xenium slides were, then, loaded for imaging and analysis on the Xenium Analyzer instrument, followed by decoding, according to the user guide (User guide CG000584, RRID SCR_023910). Following the analysis guidelines of 10x Genomics, negative controls spatial map was considered and valued according to the expectations as described in https://www.10xgenomics.com/support/software/xenium-onboard-analysis/latest/analysis/xoa-output-analysis-summary (10x Genomics Xenium Onboard Analysis, RRID SCR_026158). Instrument software version 1.4.2.0 and software analysis version 1.4.0.6 were used (10x Genomics Xenium Explorer, RRID SCR_025847).

output-XETG00123_0030916_TIS06055-001-015_20240801_192724.zip: Xenium Ranger output for 471 gene custom panel on human tonsil section

output-XETG00123_0030916_TIS06055-001-015_20240801_192724.explorer.tar.gz: Baysor re-segmentation output on the dataset above