visium spatial gene expression reagent v2 kit  (10X Genomics)

Journal: bioRxiv

Article Title: OmicGlaze: Spatial Multi-Omic Mapping of Traumatic Brain Injury

doi: 10.64898/2026.03.30.715462

Figure Lengend Snippet: A A focal blunt injury was induced in the upper right hemisphere of the mouse brain. Serial coronal sections were obtained at the mid-hippocampal level, with adjacent sections allocated for histological analysis (hematoxylin and eosin staining and Nissl staining), spatial RNA sequencing (spatial RNA-seq), and spatial ATAC sequencing (spatial ATAC-seq). B Image of a Nissl-stained section adjacent to the TBI_03 sample used for sequencing, illustrating tissue morphology. The left side corresponds to the uninjured hemisphere, and the right side to the injured hemisphere. C Zoomed-in view of the isocortex on the uninjured side of panel B. Scale bar represents 250 µm. D Zoomed-in view of the isocortex on the injured side of panel B. Scale bar represents 250 µm. E Spatial distribution of the number of detected genes per spot in the Control_01 sample. Each dot represents a 55-micron spot on the Visium slide (11 x 11 mm 2 ). F - H Spatial distribution of the number of detected genes per spot in the TBI_01, TBI_02, and TBI_03 samples. I Spatial region annotation of the TBI_03 sample, highlighting 14 anatomical regions. Abbreviations: CNU, cerebral nuclei; CTX_ISO, isocortex; CTX_OLF, olfactory cortex; CTX_RSP, retrosplenial area; CTX_SP, cortical subplate; HPF_CA_PYR, hippocampal formation cornu ammonis pyramidal layer; HPF_DG_GCL, hippocampal formation dentate gyrus granule cell layer; HPF_other, the polymorphic layer and molecular layer of hippocampus; HY, hypothalamus; LM, leptomeninges; TH, thalamus; VS, ventricular systems. J Unsupervised clustering of the integrated dataset comprising Control_01, TBI_01, TBI_02, and TBI_03 samples, colored by brain regions as defined in panel I. K Heatmap showing the expression levels of the top five marker genes for each of the 14 annotated regions in Control_01. Each row represents one gene and each column under the same color represents one region. Selected marker genes are labeled on the plot: Adora2a (marker for CNU region), Apod (fiber tracts), Satb2 (CTX_ISO region), Dlk1 (HY region), Synop2 (TH region), Hopx ( HPF_other region), Nrep (CTX_RSP region), Ttr (VS region), and Gpr151 (habenula region).

Article Snippet: We used Visium V2 CytAssist Spatial Gene Expression Reagent Kits (10X Genomics, #1000523) to profile mRNAs from tissue slices.

Techniques: Staining, RNA Sequencing, Sequencing, Control, Olfactory, Expressing, Marker, Labeling

Journal: bioRxiv

Article Title: 4D multimodal wound healing atlas reveals organ-level controls of repair phase transitions

doi: 10.64898/2026.01.15.699736

Figure Lengend Snippet: (A) Schematic overview of WoundScape, the organ-scale spatial transcriptomic wound atlas. High-resolution 2 μm Visium HD data were generated for UW, D7PW, D15PW, and D30PW skin, totaling 532,066 spatial barcoded spots. These data were merged with the OWHA omnibus, yielding a comprehensive tetra-modal spatially resolved database encompassing 725,590 total cells and spots organized across all anatomical compartments of the skin. Histological H&E staining was combined with WoundScape spatial profiling to precisely align Visium HD-identified neighborhoods within defined cutaneous wound anatomical regions. (B–E ) Spatial transcriptomic mapping of Banksy neighborhoods (bottom), and corresponding H&E sections (above), in 8 μm Visium HD sections from unwounded ( B ) (UW), ( C ) D4PW, ( D ) D7PW, and ( E ) D30PW. Middle insets show magnified views of local BANKSY neighborhoods clustering (clustering resolution = 0.5). Right insets depict corresponding RCTD-derived metacluster annotations for all discrete spatial domains within each section. Each section represents a technical replicate from the same biological specimen (UW = 113,587 spots; D4PW = 104,247; D7PW = 176,256; D30PW = 137,976). Red arrowheads indicate the initial wound edge in the suprabasal layer. White guidelines mark the initial subcutis wound boundaries. Scale bars represent 500 um or 1 mm as indicated. (F–I) Stacked bar plots showing the proportional metacluster composition within each BANKSY cluster for unwounded skin and wounded mouse skin (D4PW, D7PW, D30PW). Data represents two technical replicates derived from the same biological sample. Statistical similarity of metacluster compositions between replicates was assessed using a chi-square test with Monte Carlo permutation (100,000 simulations). BANKSY clusters that show significant concordance between technical replicates ( P < 0.05) are highlighted in red font. (J) Visium HD localization of Dominant Signalers and Central Orchestrators, including Basal IV, Papillary II, HF I, Myofibroblasts II, and Muscle Progenitor clusters, along BANKSY neighborhoods proximal to the wound bed at D7PW. Red outlines mark wound-associated regions of interest, while yellow guidelines and red arrowheads denote suprabasal and subcutis wound edge boundaries, respectively. Scale bars, 1 mm. (K) Anatomical distance quantifications of wound emergent BANKSY clusters along the anterior-posterior transverse plane at D7PW. The x-axis represents arbitrary spatial units (1 a.u. = 9 mm) corresponding to anterior-posterior distance across the entire tissue section. Vertical redline demarcates the wound center. Data represents two technical replicates from the same biological timepoints. A variance-based localization test was used, and multiple testing correction was applied to p-values using the Benjamini–Hochberg procedure. (p < 0.05 = *, p < 0.01 = **, p < .001 = ***). (L) Schematic illustration summarizing the spatial geometry of the wound edge (anterior and posterior margins) as visualized at D7PW. Red arrowheads indicate typical location of wound boundaries from the adjacent wound bed. (M-N) High-magnification 20x H&E of the D7PW wound edge regions of interest (ROI). Red arrowheads indicate wound boundaries. Scale bars, 500 μm. (O-T) BANKSY spatial clustering of respective Dominant Signalers populations within the posterior wound edge ROI: ( O ) BANKSY cluster positions, ( P ) merged overlay of selected Dominant Signaler populations: ( Q ) Spinous I, ( R ) Proliferative Endothelial Cells, ( S ) Pericyte I, and ( T ) HF I. Each overlay highlights discrete but spatially organized domains at the wound front where reparative signaling networks converge. Scale bars, 100 μm. (U) Stacked bar plots showing fine cell-type composition of BANKSY clusters localized at the D7PW wound front. Data represents two technical replicates from the same biological timepoints. (V) Numbering and classification legend of fine cell types corresponding to panel (U) .

Article Snippet: Spatial Transcriptomics data was generated using 10x Genomics Visium V2 CytAssist Spatial Gene Expression Mouse Transcriptome Assay (#1000445) for FFPE tissue as per user’s guide.

Techniques: Generated, Staining, Derivative Assay

Journal: bioRxiv

Article Title: 4D multimodal wound healing atlas reveals organ-level controls of repair phase transitions

doi: 10.64898/2026.01.15.699736

Figure Lengend Snippet: ( A ) Schematic illustration of cross-tissue wound signaling between epidermal Central Orchestrators and deep tissue population populations. ( B–E ) Visium HD spatial localization of Basal IV keratinocytes (blue) and proliferative endothelial cells (yellow) across the healing time course ( B ) UW, ( C ) D4PW, ( D ) D7PW, ( E ) D30PW. Images represent one biological replicate, with one technical replicate shown for UW. Red arrowheads indicate the suprabasal wound edge; red guidelines denote subcutis wound boundaries. Yellow boxes indicate magnified insets highlighting Basal IV–endothelial interactions at the wound edge. Scale bar, 1 mm. ( F–I ) Quantification of anatomical distance between Basal IV and proliferative endothelial cells along the anterior–posterior wound axis (1 a.u. = 9 mm) at ( F ) UW, ( G ) D4PW, ( H ) D7PW, ( I ) D30PW. Data represent two technical replicates from one biological sample per timepoint. Significance was assessed using a variance-based localization test with Benjamini–Hochberg correction (* p < 0.05; ** p < 0.01; *** p < 0.001). ( J–K ) CellChat-inferred signaling network ( J ) and corresponding ligand–receptor pathways ( K ) transmitted between Basal IV keratinocytes and endothelial cells. Edge width denotes interaction strength. ( L ) Visium HD spatial transcriptomic expression of Sema3c pathway components ( Sema3c , Nrp1 , Nrp2 , Plxna4 ) overlaid on corresponding H&E sections at D7PW. Scale bar, 1 mm. ( M–N ) Single-molecule RNA FISH (smFISH) immunofluorescence showing Sox6 (magenta) and Sema3c (yellow) mRNA transcripts with DAPI (white) in ( M ) UW and ( N ) D4PW skin. Boxed regions highlight an Sox6-high epidermal zone. (Right) zoomed in images of the Sox6-high zone with single-channel panels shown. Wound edge (w.e.) indicated by red arrowheads and subcutis wound boundaries indicated by red lines. Scale bars, 100 µm (full images) or 10 µm (zoomed insets). ( O–P ) Boxplots quantifying mean fluorescence intensity of ( O ) Sox6 and ( P ) Sema3c smFISH signals across timepoints (UW, D4PW, D7PW), comparing unwounded, distal, and proximal wound regions. UW includes three biological replicates, D4PW two biological replicates, and D7PW one biological replicate . Statistical testing performed using a two-sided Wilcoxon test. Variability is represented using the interquartile range (IQR). Statistical significance was determined using a Wilcoxon rank-sum test (p < 0.05 = *, p < 0.01 = **). ( Q ) UMAP visualizations and corresponding pseudotime ordering of UW IFE keratinocyte subclusters from OWHA snRNA-seq. n = two biological replicates. ( R ) Heatmap of top 100 pseudotime-associated genes in UW snRNA-seq keratinocytes ordered by cluster and pseudotime. Arrowheads mark genes enriched in early (purple) vs. late (yellow) pseudotime. ( S ) UMAP visualizations and corresponding pseudotime ordering of D4PW–D7PW IFE keratinocyte subclusters showing branching into re-epithelization (red) and neurovasculogenesis (green) lineage trajectories. n = two biological replicates per timepoint. ( T ) Heatmap of the top 100 pseudotime-associated genes expressed along the neurovasculogenesis pseudotime trajectories in D4PW–D7PW keratinocytes, ordered by cluster and timepoint. Arrows indicate early (purple) and late (yellow) pseudotime gene signatures. ( U–W ) Mean pseudotime expression profiles of ( U ) Sox6 , ( V ) Sema3c , and ( W ) Krt6a along the D4PW–D7PW proliferative pseudotime trajectory. ( X ) Gene Ontology terms among genes upregulated at early pseudotime stages of the neurovasculogenesis trajectory, subdivided into molecular function (MF), biological process (BP), and cellular component (CC) categories.

Article Snippet: Spatial Transcriptomics data was generated using 10x Genomics Visium V2 CytAssist Spatial Gene Expression Mouse Transcriptome Assay (#1000445) for FFPE tissue as per user’s guide.

Techniques: Expressing, Immunofluorescence, Fluorescence

Journal: bioRxiv

Article Title: 4D multimodal wound healing atlas reveals organ-level controls of repair phase transitions

doi: 10.64898/2026.01.15.699736

Figure Lengend Snippet: ( A ) UMAP embedding of the fully integrated cross-species wound-healing atlas (COWA) containing 236,930 single cells. COWA integrates the multimodal OWHA dataset with the human wound healing dataset (GSE241132), generating a total of 40 sequencing runs (28 murine, 12 human). ( B ) Annotation legend displaying all 107 fine-grained subcluster identities represented in COWA. ( C–D ) UMAP projections showing ( C ) human (n=12) and ( D ) mouse (n=28) wound datasets downsampled to 43,406 cells each for comparable visualization of interspecies cell capture differences. ( E ) Sankey plot showcasing proportional representation of major cell types between mouse and human samples across metaclusters, highlighting shared and species-specific tissue compositions. Percentages display proportion of each respective metacluster assigned to integrated cross-species Harmony clusters. ( F ) MILO differential abundance analysis of select IFE keratinocyte, endothelial, pericyte, and Schwann cell subclusters states significantly enriched in mouse (blue) or human (red) samples (FDR = 0.15). Mouse-enriched (>0 LFC) states represent populations preferentially detected in by multimodal murine atlasing. ( G–H ) UMAP feature plots depicting ( G ) mouse Sox6 and ( H ) human SOX6 expression. ( I–J ) UMAP feature plots of (I) mouse Sema3c and (J) human SEMA3C , showing substantially reduced SEMA3C detection in human wound-healing scRNA-seq datasets. ( K–L ) Dot plots of Basal IV–associated markers ( Sox6 , Cdh13 , Sema3c ) and the wound-induced keratinocyte gene Krt16 in IFE keratinocytes from ( K ) the murine OWHA dataset and ( L ) the human wound dataset. Dot size indicates the percentage of expressing cells; color intensity denotes mean expression level. ( M ) Beeswarm plot of MILO differential abundance comparing whole-cell (blue) versus single-nucleus (red) RNA-seq, highlighting modality-specific biases in cell type capture (FDR = 0.15). ( N ) Visium spatial transcriptomic feature plot of D7PW human wound tissue (GSE241132), annotated by regional labels as defined Liu et al. 2024. Red arrowheads mark the wound edge. ( O–P ) Quantification of Basal IV module scores in ( O ) unwounded and ( P ) D7PW human tissue sections (calculated using AddModuleScore() ). Red box highlights wound edge region zoomed-in in the right panel. Red arrowheads denote the wound edge; white dashed lines outline the epidermal-dermal boundary. ( Q–R ) Quantification of SEMA3C pathway signature scores in ( Q ) unwounded and ( R ) D7PW human tissue sections. Red box highlights wound edge region zoomed-in in the right panel. Red arrowheads denote the wound front; white dashed lines indicate the epidermal-dermal boundary. ( S–T ) Mean expression of ( S ) SEMA3C in keratinocytes and ( T ) NRP1 and NRP2 in endothelial cells at the human wound edge across healing timepoints (D0PW, D1PW, D7PW, D30PW). Each dot represents the mean signal per sequencing run. Statistical significance was assessed using a Wilcoxon rank-sum test relative to D0/UW (* p < 0.05; ** p < 0.01). n = 4.

Article Snippet: Spatial Transcriptomics data was generated using 10x Genomics Visium V2 CytAssist Spatial Gene Expression Mouse Transcriptome Assay (#1000445) for FFPE tissue as per user’s guide.

Techniques: Sequencing, Expressing, RNA Sequencing