primary antibodies against cd31 pecam 1  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: Bioabsorbable Magnesium Metal Scaffolds Improve Dermal Wound Healing and Tissue Regeneration

doi: 10.64898/2026.03.03.709352

Figure Lengend Snippet: Mg increases capillary lumen density. (a) CD31 (endothelial cells) IHC staining of PBS and Mg wounds at day 7 post-wounding was performed and 20x representative images of wounds are shown. Scale bar = 50 µm. (b) Quantification of lumens per HPF from 4-6 HPFs across the wound bed per wound (40x). (c) Quantification of lumens proximal (within 200 µm from Mg wires) vs distal (200 µm away from Mg wires) at day 7 post-wounding. (d) Tuj1 stained positive nerve bundles in PBS and Mg wounds at day 7 post-wounding. Scale bar = 200 µm. (e) Quantification of % of Tuj1 positive nerve bundles per HPF (40x) at day 7 post-wounding. Bar graphs show mean ± SD. Individual dots represent average measurements from 4-6 HPFs per wound from different animals. * P < .05, ** P < .01, *** P < .001.

Article Snippet: Primary antibodies against CD31 (PECAM-1) for endothelial cells (D8V9E; 1:50; Cell Signaling Technology, Danvers, MA), CD45 for pan-leukocytes (ab10558; 1:5000; Abcam, Cambridge, MA) and F4/80 for pan-macrophages (ab111101; 1:100; Abcam, Cambridge, MA) were detected by EnVision+System-HRP (DAB) kits (Dako North America, Carpinteria, CA) and hematoxylin counterstaining.

Techniques: Immunohistochemistry, Staining

Journal: bioRxiv

Article Title: Bioabsorbable Magnesium Metal Scaffolds Improve Dermal Wound Healing and Tissue Regeneration

doi: 10.64898/2026.03.03.709352

Figure Lengend Snippet: Mg improves tissue regeneration. (a) Pictures of scars of PBS and Mg wounds at day 28 post-wounding. Yellow dashed lines indicate scar area. (b) Mg wires (yellow arrows) in skin explants on micro-CT at day 28. Scale bar = 2.5 mm. (c) Trichrome staining of Mg wound shows basket weave pattern compared to parallel collagen bundles in PBS wound. Scale bar = 50 µm. (d) Scar area measured at day 28. (e) Picrosirius red fluorescence in PBS and Mg wounds at day 28. Scale bar = 100 µm. (f) The scars were divided into 4 parts to represent the middle area and edges, and picrosirius positive area was calculated as percent of region of interest (ROI) area to represent collagen ratio in PBS and Mg wounds at day 28. (g) Epidermal thickness at day 28. (h) Dermal appendages/HPF at day 28. (i) CD31 IHC staining of Mg and PBS wounds at day 28, Scale bar = 50 µm. (j) Vessel density at day 28 determined by CD31 staining. Bar graphs show mean ± SD. Individual dots represent average measurements from 4-6 HPFs per wound from different animals. * P < .05, ** P < .01, *** P < .001.

Article Snippet: Primary antibodies against CD31 (PECAM-1) for endothelial cells (D8V9E; 1:50; Cell Signaling Technology, Danvers, MA), CD45 for pan-leukocytes (ab10558; 1:5000; Abcam, Cambridge, MA) and F4/80 for pan-macrophages (ab111101; 1:100; Abcam, Cambridge, MA) were detected by EnVision+System-HRP (DAB) kits (Dako North America, Carpinteria, CA) and hematoxylin counterstaining.

Techniques: Micro-CT, Staining, Fluorescence, Immunohistochemistry

Journal: Journal of Advanced Research

Article Title: OTUD1 delays wound healing by regulating endothelial function and angiogenesis in diabetic mice

doi: 10.1016/j.jare.2025.04.038

Figure Lengend Snippet: Endothelial OTUD1 is significantly upregulated in diabetic wound tissues. (A) Real-time qPCR analysis for mRNA levels of OTU subfamily members in the skin wound tissues from control and T2DM mice (n = 7). (B) Immunoblotting and densitometric quantification analyses illustrating OTUD1 protein expression in skin wound tissues from both control and T2DM mice (n = 3). (C) Representative immunohistochemical images and (D) quantitative analysis of showing OTUD1-positive cells (brown) in the skin wound tissues of mice. Black arrowheads indicate positive OTUD1 signals (Scale bar = 50 μm; n = 7). (E) Immunoblot and quantitative analysis of OTUD1 protein expression in HUVECs, HaCaT, HDFa, and MPM cells (n = 4). (F) Representative immunofluorescence images displaying the colocalization of CD31 (red) and OTUD1 (green) in mouse skin wound tissues, with white arrowheads showing OTUD1 and CD31 colocalization sites. Tissues were counterstained with DAPI (blue; Scale bar = 50 μm). (G) Immunofluorescence staining and (H) quantitative analysis of OTUD1-positive cells (red) in both control and HG + PA-treated HUVECs for 4 h, counterstained with DAPI (blue; Scale bar = 50 μm). (I) Time-course study of OTUD1 expression in response to HG + PA in HUVECs, including immunoblot analysis and quantitative measurement (n = 3). Data are shown as mean ± SEM. Statistical analyses were performed using a two-tailed unpaired Student's t -test (A, B, D), Welch’s t test (H), and one-way ANOVA analysis followed by Bonferroni post-hoc test (E, I). HG + PA indicates treatment with 50 mM HG and 300 μM PA, unless specified otherwise. Abbreviations: Ctrl, control; T2DM, type 2 diabetes mellitus; OTU, ovarian tumor protease; HUVECs, human umbilical vein endothelial cells; HaCaT, human keratinocytes; HDFa, human dermal fibroblasts-adult; MPMs, mouse primary peritoneal macrophages; DAPI, 4ʹ,6-diamidino-2-phenylindole; HG + PA, high glucose plus palmitic acid; and OTUD1, ovarian tumor deubiquitinase 1.

Article Snippet: After one hour of blocking with rabbit serum (G1209, Servicebio), primary antibodies against CD31 (AF3628, R&D Systems, Minnesota, USA) were incubated for the whole night at 4 °C.

Techniques: Control, Western Blot, Expressing, Immunohistochemical staining, Immunofluorescence, Staining, Two Tailed Test

Journal: Journal of Advanced Research

Article Title: OTUD1 delays wound healing by regulating endothelial function and angiogenesis in diabetic mice

doi: 10.1016/j.jare.2025.04.038

Figure Lengend Snippet: OTUD1 deficiency rescues impaired wound healing by enhancing angiogenesis and fibrosis in T2DM mice. (A) Schematic diagram illustrating the animal experiment procedure. The mice (B) FBG levels and (C) body weight were recorded from weeks 9 to 16 (n = 7). * P < 0.05 vs WT-Sham; ** P < 0.01 vs WT-Sham; *** P < 0.001 vs WT-Sham; ns, no significance. ns (green) indicates that there is no statistical significance between OTUD1 −/− -T2DM and WT-T2DM. (D) Representative wound images and (E) wound closure rates are shown (n = 5). *** P < 0.001 vs WT-Sham; ## P < 0.01 vs WT-T2DM; ns, no significance. (F) H&E staining demonstrated regenerated skin at day 12 across different groups. Scale bar = 50 μm. (G) Quantitative assessments of epidermis thickness in mice (n = 7). (H) Masson's trichrome staining and (I) quantitative analysis of collagen deposition in skin wound tissues at day 12 (Scale bar = 50 μm; n = 7). (J) Representative images and (K) quantification of CD31-positive (brown) neovascularization via immunohistochemical staining at days 3, 7, and 12 (Scale bar = 50 μm; n = 7). Black arrows indicate skin neovascularization. (L-O) Immunoblotting and quantification of OTUD1, VEGFR2, p-eNOS, and eNOS in wound tissue lysates from Sham or T2DM mice with WT or OTUD1 knockout, normalized to GAPDH (n = 4). Data are displayed as mean ± SEM. Statistical analyses were performed using two-way ANOVA analysis followed by Bonferroni post-hoc test (B, C, E) and one-way ANOVA analysis followed by Bonferroni post-hoc test (G, I, K, M-O). Abbreviations: WT, wild-type; OTUD −/− , OTUD1-knockout; HFD, high-fat diet; STZ, streptozotocin; FBG, fasting blood glucose; eNOS, endothelial nitric oxide synthase.

Article Snippet: After one hour of blocking with rabbit serum (G1209, Servicebio), primary antibodies against CD31 (AF3628, R&D Systems, Minnesota, USA) were incubated for the whole night at 4 °C.

Techniques: Staining, Immunohistochemical staining, Western Blot, Knock-Out

Journal: Journal of Advanced Research

Article Title: OTUD1 delays wound healing by regulating endothelial function and angiogenesis in diabetic mice

doi: 10.1016/j.jare.2025.04.038

Figure Lengend Snippet: Pharmacological inhibition of β-catenin reverses OTUD1 deletion-mediated recovery of delayed wound healing in db/db mice. (A) Schematic of the protocol for establishing the OTUD1 deletion mouse model in db/m or db/db mice. The mice (B) FBG levels and (C) body weight of the indicated mice at weeks 10, 12, 14, 16, 18, 20, 22, and 24 (n = 7). *** P < 0.001 vs db/m -AAV-shNC. ns (green) indicates that there is no statistical significance between db/db -AAV-shNC and db/db -AAV-shOTUD1. ns (purple) indicates that there is no statistical significance between db/db -AAV-shOTUD1 and db/db -AAV-shOTUD1-MSAB. (D) Skin wound healing images and (E) statistical analysis of wound closure rate (n = 5). ** P < 0.01 vs db/m -AAV-shNC; *** P < 0.001 vs db/m -AAV-shNC; ## P < 0.01 vs db/db -AAV-shNC; ### P < 0.001 vs db/db -AAV-shNC; && P < 0.01 vs db/db -AAV-shOTUD1; ns, no significance. (F) H&E staining images of skin wound tissues. Scale bar = 50 μm. (G) Representative Masson’s trichrome staining in skin wound tissues. Scale bar = 50 μm. (H) CD31 immunohistochemical staining of neovascularization in skin wound tissues on days 3, 7, and 12 (Scale bar = 50 μm). The black arrows denote skin neovascularization. (I) Representative immunoblotting and (J-M) quantitative analysis of OTUD1, VEGFR2, Nuc-β-catenin, p-eNOS, and eNOS protein levels in skin wound tissues from different groups (n = 4). Data are shown as mean ± SEM. Statistical analyses were performed using two-way ANOVA analysis followed by Bonferroni post-hoc test (B, C, E) and one-way ANOVA analysis followed by Bonferroni post-hoc test (J-M). Abbreviations: MSAB, ethionine sulfoxide β-methyl ester; AAV2/BI30-shOTUD1, adeno-associated virus serotype 2/BI30 carrying shOTUD1 under the CMV promoter.

Article Snippet: After one hour of blocking with rabbit serum (G1209, Servicebio), primary antibodies against CD31 (AF3628, R&D Systems, Minnesota, USA) were incubated for the whole night at 4 °C.

Techniques: Inhibition, Staining, Immunohistochemical staining, Western Blot, Virus