goat anti human prox1  (R&D Systems)

Journal: bioRxiv

Article Title: Breast Cancer Remodels Lymphatics in Sentinel Lymph Nodes

doi: 10.1101/2024.12.30.630756

Figure Lengend Snippet: a, UMAP plot of LECs from distant LNs (left) or metastatic LNs (right), color coded by cluster. b, Proportion of each LEC subset in distant and metastatic LNs, calculated from nine patients LECs (two-tailed, paired Student’s t-test), *p <0.05. c, Differential abundance testing using miloR. Neighborhoods are colored by their log fold abundance change between distant (blue) and metastatic (red) LNs. Non-differential abundance neighborhoods (FDR > 10%) are colored white. d, Beeswarm plot of the cell subset distribution of log fold change between normal and distant LNs. e, The trajectories of LN LEC differentiation shown in a UMAP plot. Six distinct LEC trajectories (T1-T6) were identified using Monocle single-cell trajectory analysis. f, Immunostaining of cancer cells and LEC markers PROX1 and MARCO in non-metastatic (left) and metastatic LNs (right). Zoomed-in images displaying SCS containing cytokeratin + cancer cells (left) and medullary sinuses (MS) both with and without cancer cells (right). A cancer cell in the MARCO + sinus is indicated by an arrow. Blue, cytokeratin; red, PROX1; green, MARCO. Scale bars, 500 μm. Images are representatives of two individuals with similar results. g, Immunostaining of CD200 + lymphatics and its quantification in metastatic (upper, black background) and non-metastatic (lower, grey background) LNs. CD200 + lymphatics and individual cancer cells within lymphatics are indicated by arrows and arrowheads, respectively. Blue, cytokeratin; green, PROX1; red, CD200. Scale bars, 500 μm. Images are representatives of seven individuals with simar results. SCS: subcapsular sinus; MS: medullary sinus; B: B cell zone. Circles in the bar plots represent biological replicates (mean±SEM, two-tailed, unpaired Student’s t-test). **p <0.005.

Article Snippet: The following primary antibodies were used: goat anti-human PROX1 (R&D Systems AF2727), rabbit anti-human MARCO (Atlas Antibodies, HPA063793), AF488 mouse anti-pan-cytokeratin (Thermo Fisher Scientific, MA5-18156), mouse anti-human MGP (Novus Biologicals, NBP2-45844), and BV421 mouse anti-human CD200 (Biolegend, 329209).

Techniques: Two Tailed Test, Immunostaining

Journal: bioRxiv

Article Title: Breast Cancer Remodels Lymphatics in Sentinel Lymph Nodes

doi: 10.1101/2024.12.30.630756

Figure Lengend Snippet: a, Differential abundance testing using MiloR and a heatmap of differentially expressed genes between differential abundance neighborhoods in LN LECs. The UMAP plot was shown in and is inserted here for clarity. In the heatmap, columns and rows represent neighborhoods and differentially expressed genes, respectively. Expression values for each gene are scaled between 0 and 1. The upper panel of the heatmap shows the neighborhood log fold change. FDR, false discovery rate. b, Heatmap showing the expression of the top DEGs between metastatic and distant LNs for each cell. Bars above the heatmap indicate the tissue and cluster origin of each cell (LNs, clusters). c, Violin plots displaying the top DEG expression between metastatic and distant LNs by cluster, with log-normalized expression value labeled. d, Immunostaining of MGP and its quantification in metastatic and distant LNs. Zoomed-in images show SCS and medullary sinuses containing cancer cells and MGP expression on PROX1 + LECs (arrows). Blue, cytokeratin; red, MGP; green, PROX1. Scale bars, 125 μm. Images are representative of four individuals with similar results. Circles in the bar plots represent biological replicates (mean±SEM, two-tailed, unpaired Student’s t-test). *p <0.05. e, Gene Ontology (GO) enrichment analysis of the top DEGs between metastatic and distant LNs using EnrichR package.

Article Snippet: The following primary antibodies were used: goat anti-human PROX1 (R&D Systems AF2727), rabbit anti-human MARCO (Atlas Antibodies, HPA063793), AF488 mouse anti-pan-cytokeratin (Thermo Fisher Scientific, MA5-18156), mouse anti-human MGP (Novus Biologicals, NBP2-45844), and BV421 mouse anti-human CD200 (Biolegend, 329209).

Techniques: Expressing, Labeling, Immunostaining, Two Tailed Test

Journal: bioRxiv

Article Title: Differential contribution of P73 + Cajal-Retzius cells and Reelin to cortical morphogenesis

doi: 10.1101/2024.10.15.618167

Figure Lengend Snippet: (A) Drawing indicating the regions shown in B-L (top) and color key for the panel shown in B. (B) Immunostaining for Reln and Tomato in the neocortex or hippocampus of P1 control or Reln cKO animals. (C) Immunostaining for Reln in control and cKO showing the decreased Reln expression, in the dorsolateral cortex and hippocampus. (D) Quantification of the fraction of Reln + cells among Tomato + CRs in the dorsal cortex and hippocampus. Each point corresponds to one animal. (E) Quantification of the density of Tomato + CRs in the dorsal cortex MZ of control and cKO mice. Each point corresponds to one animal. (F) Density of Tomato + CRs along the medio-lateral axis of the cortical MZ (n=3 animals per genotype). (G) Quantification of the fluorescence intensity of Reln (arbitrary units) along the HF and cortical MZ. The dashed lines represent each animal considered (n=3 per genotype). (H) Immunostaining for Tbr1and Brn2, or for CTIP2 and Tomato in a cortical column of the presumptive somatosensory cortex of P1 control and Reln cKO . (I) In situ hybridization for Tbr1 and Rorb in a rostral section of the cortex from P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. (J) Immunostaining for CTIP2 in a cortical column of the presumptive somatosensory cortex of P1 control and PGK Cre ;Reln lox/- mice. (K) In situ hybridization for Nr4a2 in the lateral cortex of P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. CLA: claustrum. (L) DAPI staining of nuclei in the rostromedial cortex of P1 control, Reln cKO and PGK Cre ;Reln lox/- mice. Arrows point to cells abnormally positioned in the MZ of mutants. (M) Immunostaining for Foxg1 and Tomato along the medio-lateral axis of the caudal cortex of P1 control or Reln cKO . (N) Immunostaining for CTIP2 and Prox1 in the P1 hippocampus of control and Reln cKO . (O) Immunostaining for CTIP2 and Prox1 or Laminin in the P1 hippocampus of control and PGK Cre ;Reln lox/- mice. The arrow points to the defect in compaction of the CA1 pyramidal layer. HF: hippocampal fissure. Scale bars: 50µm in B, K. 100µm in F, H. 200µm in G, J, L, M. 500µm in I, K.

Article Snippet: The following primary antibodies were used: goat anti-Brn2 (POU3F2, Abcam ab101726 1:1000), rat anti-CTIP2 (BCL11B, Abcam ab18465 1:600), rabbit anti-FOXG1 (Abcam ab18259 1:2000), rabbit anti-Laminin (Sigma-Aldrich L9393 1:600), goat anti-Neuropilin-1 (R&D Systems AF566 1:800), rabbit anti-p73 (Cell signaling 14620 1:250), goat anti-Nurr1 (NR4A2, R&D Systems AF2156 1:200), goat anti-Prox1 (R&D Systems AF2727 1:1000), goat anti-Reelin (R&D Systems AF3820 1:2000), rabbit anti-TBR1 (Abcam ab31940 1:1000).

Techniques: Immunostaining, Control, Expressing, Fluorescence, In Situ Hybridization, Staining

Journal: bioRxiv

Article Title: Differential contribution of P73 + Cajal-Retzius cells and Reelin to cortical morphogenesis

doi: 10.1101/2024.10.15.618167

Figure Lengend Snippet: (A) Drawing indicating the region shown in B-E (top) and region specificity of the markers used (bottom). (B) Immunostaining for CTIP2 on coronal sections of the hippocampus from E16 to P1 in control and Gmnc -/- animals. DG: dentate gyrus anlage. (C) Immunostaining for EdU on sections of the hippocampus at P2 in control and Gmnc mutants following EdU incorporation at E15. SO: stratum oriens , SP: stratum pyramidale . The yellow dashed line separates the regions considered normal and folded in mutants. Quantification of EdU labelled cells in control and mutant (n=3 each), considering normal or folded regions of CA1. (D) Immunostaining for Laminin (green) on coronal sections of the hippocampus in E18. HF: hippocampal fissure, only distinguishable in controls. (E) In situ hybridization for Nr4a2 and Zbtb20 , and immunostaining for Prox1 on coronal sections of the hippocampus at P1 in control and Gmnc -/- animals. Scale bars: 200µm in B, C, E. 50µm in D.

Article Snippet: The following primary antibodies were used: goat anti-Brn2 (POU3F2, Abcam ab101726 1:1000), rat anti-CTIP2 (BCL11B, Abcam ab18465 1:600), rabbit anti-FOXG1 (Abcam ab18259 1:2000), rabbit anti-Laminin (Sigma-Aldrich L9393 1:600), goat anti-Neuropilin-1 (R&D Systems AF566 1:800), rabbit anti-p73 (Cell signaling 14620 1:250), goat anti-Nurr1 (NR4A2, R&D Systems AF2156 1:200), goat anti-Prox1 (R&D Systems AF2727 1:1000), goat anti-Reelin (R&D Systems AF3820 1:2000), rabbit anti-TBR1 (Abcam ab31940 1:1000).

Techniques: Immunostaining, Control, Mutagenesis, In Situ Hybridization