Fig. 6
From: FixNCut: single-cell genomics through reversible tissue fixation and dissociation
FixNCut protocol tested in human colon biopsies. a Mapping analysis of sequencing reads within a genomic region. b Comparative analysis of the number of detected genes (top) and UMIs (bottom) across various sequencing depths. c Cumulative gene counts analyzed using randomly sampled cells. d Principal component analysis (PCA) and uniform manifold approximation and projection (UMAP) representation of gene expression profile variances prior data integration, and harmony integrated UMAP representation of gene expression profile variances of fresh, fixed, cryopreserved, and fixed/cryopreserved samples. e Linear regression model comparing average gene expression levels of expressed genes across protocols used. The coefficient of determination (R2) computed with Pearson correlation is indicated. f Hierarchical clustering of coefficient of determination (R2) obtained for all pair comparisons across protocols for biological hallmarks, including apoptosis, hypoxia, reactive oxygen species (ROS), cell cycle G2/M checkpoint, unfolded protein response (UPR), and inflammatory response genes. g UMAP visualization of 17,825 fresh, fixed, cryopreserved, and fixed/cryopreserved human colon cells, colored by 21 cell populations. h Comparison of cell population proportions between fresh (n = 5759), fixed (n = 4250), cryo (n = 3489), and fixed/cryo (n = 4327) cells. The bottom figure shows the results of compositional cell analysis using the Bayesian model scCODA. Credible changes and Log2FC are indicated. i Differential gene expression analysis across conditions: fixed vs fresh (top-left), fixed vs cryo (top-right), fixed/cryo vs cryo (bottom-left), and fixed/cryo vs fixed (bottom-right). Significant adjusted p-values (FDR) < 0.05, upregulated (red), and downregulated (blue) genes with Log2FC > |1| are indicated. The top DE genes are included in the plot. j Violin plots for stress-related gene signature score [2, 3, 22] for human colon biopsies across protocols. Statistical analysis between protocols was performed using the Wilcoxon signed-rank test