primary antibodies tim  (Sino Biological)

Journal: The FASEB Journal

Article Title: ZXDB Drives Macrophage Inflammatory Programming in Sepsis‐Induced Acute Kidney Injury by Recruiting EIF4A3 to Enhance ACACA Translation

doi: 10.1096/fj.202502962RR

Figure Lengend Snippet: Macrophage‐Specific Deletion of Zxdb Protects Against Sepsis‐Induced Acute Kidney Injury. (A) Generation strategy for myeloid‐specific Zxdb knockout (Mac‐Zxdb‐KO) mice. (B) H&E staining of kidney sections from Sham, WT SI‐AKI, and Mac‐Zxdb‐KO SI‐AKI mice. (C‐D) Serum creatinine (Scr) (C) and blood urea nitrogen (BUN) (D) levels in Sham, WT SI‐AKI, and Mac‐Zxdb‐KO SI‐AKI mice at 24 h post‐CLP. (E) KIM‐1 immunohistochemistry of kidney sections from Sham, WT SI‐AKI, and Mac‐Zxdb‐KO SI‐AKI mice. (F) TUNEL staining of kidney sections from Sham, WT SI‐AKI, and Mac‐Zxdb‐KO SI‐AKI mice. (G) Immunofluorescence of kidney sections stained for F480, CD86, and CD206. (H) Serum IL‐1β, IL‐10, and TNF‐α levels in Sham, WT SI‐AKI, and Mac‐Zxdb‐KO SI‐AKI mice at 24 h post‐CLP. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Sections were blocked and incubated overnight at 4°C using primary antibody against KIM‐1 (AF1817; R&D Systems, 1:200).

Techniques: Knock-Out, Staining, Immunohistochemistry, TUNEL Assay, Immunofluorescence

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Temporal Proteomic and Phosphoproteomic Profiling Deciphers Molecular Dynamics of Acute-to-Chronic Kidney Disease After Ischemia-Reperfusion Injury, With Dock2 Emerging as a Key Regulator

doi: 10.1016/j.mcpro.2026.101509

Figure Lengend Snippet: Ischemia-reperfusion injury elicits time-dependent changes of TECs injury, inflammatory infiltration, and interstitial fibrotic remodeling. A , schematic illustration of the experimental protocol for inducing renal IRI in a mouse model. B , levels of Scr from sham-operated mice and mice sacrificed at 1 h, 1 day, 3 days, 7 days, 28 days after I/R. C and D , protein expression profiling by western blot ( C ) and subsequent statistical analysis ( D ) of Havcr1/Kim1 in renal cortical samples. E , immunohistochemical detection of Havcr1/Kim1 expression in renal cortical sections from sham and I/R-treated mice. The scale bar represents 100 μm. F and G , protein expression profiling by western blot ( F ) and subsequent statistical analysis ( G ) of IL-6 in renal cortical samples. H , immunofluorescent staining for the macrophage marker F4/80 in renal cortical sections. The scale bar represents 40 μm. I , histopathological assessment of renal tissue architecture (H&E) and collagen deposition (Masson’s trichrome) in experimental groups. The scale bar represents 100 μm. Data are presented as mean ± SEM. n.s, not significant; ∗ p < 0.05; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. Havcr, hepatitis A virus cellular receptor 1; IL-6, interleukin-6; IRI, ischemia-reperfusion injury; Kim1, kidney injury molecule-1; Scr, serum creatinine; TEC, tubular epithelial cell.

Article Snippet: Following blocking, the sections were incubated overnight at 4 °C with an anti-Havcr1/Kim1 primary antibody (R&D Systems, cat. AF1817, dilution 1:200), then treated with horseradish peroxidase (HRP)-conjugated secondary antibody at room temperature (RT) for 1 h, and finally developed with 3,3′-diaminobenzidine (DAB) substrate (ZSGB, ZLI-9017).

Techniques: Expressing, Western Blot, Immunohistochemical staining, Staining, Marker, Virus

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Temporal Proteomic and Phosphoproteomic Profiling Deciphers Molecular Dynamics of Acute-to-Chronic Kidney Disease After Ischemia-Reperfusion Injury, With Dock2 Emerging as a Key Regulator

doi: 10.1016/j.mcpro.2026.101509

Figure Lengend Snippet: Dock family proteins are identified, with knockdown of Dock2 demonstrating a significant attenuation of the proinflammatory response of TECs in vitro . A , heatmap of Dock family proteins and phosphosites (ANOVA, p < 0.05). B , intensity plots showing Dock2 ( black line ) and its phosphosite S1704 (Dock2_pS1704, green line ) across the six experimental groups. C and D , protein expression profiling by western blot ( C ) and subsequent statistical analysis ( D ) of Dock2 in renal cortical samples. E , immunofluorescent detection of Dock2 and Havcr1/Kim1 expression and colocalization in renal tissue sections from sham and I/R-treated mice. The scale bar represents 40 μm. F , the knockdown efficiency of Dock2 by siRNA in H/R-treated HK-2 cells was analyzed by western blot. G , the statistical analysis of ( F ). H – J , RT-qPCR showing the effects of Dock2 knockdown on mRNA expression levels of MCP-1, TNF-α, and IL-6. K , western blot analysis was performed to assess the effects of Dock2 knockdown on IKK-β phosphorylation in H/R-treated HK-2 cells. L , densitometric analysis of p-IKKβ from ( K ) was performed with normalization to the respective total protein. M , translocation of NF-κB p65 in HK-2 cells was detected by immunofluorescence. NF-κB p65 ( green ), DAPI ( blue ). The scale bar represents 5 μm. Data are presented as mean ± SEM. n.s, not significant; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. DAPI, 4′,6-diamidino-2-phenylindole; Dock2, dedicator of cytokinesis 2; Havcr, hepatitis A virus cellular receptor 1; H/R, hypoxia/reoxygenation; IKKβ, IκB kinase beta; Kim1, kidney injury molecule-1; IL-6, interleukin-6; MCP-1, monocyte chemoattractant protein-1; RT-qPCR, reverse transcription quantitative PCR; TEC, tubular epithelial cell; TNF-α, tumor necrosis factor-α.

Article Snippet: Following blocking, the sections were incubated overnight at 4 °C with an anti-Havcr1/Kim1 primary antibody (R&D Systems, cat. AF1817, dilution 1:200), then treated with horseradish peroxidase (HRP)-conjugated secondary antibody at room temperature (RT) for 1 h, and finally developed with 3,3′-diaminobenzidine (DAB) substrate (ZSGB, ZLI-9017).

Techniques: Knockdown, In Vitro, Phospho-proteomics, Expressing, Western Blot, Quantitative RT-PCR, Translocation Assay, Immunofluorescence, Virus, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Temporal Proteomic and Phosphoproteomic Profiling Deciphers Molecular Dynamics of Acute-to-Chronic Kidney Disease After Ischemia-Reperfusion Injury, With Dock2 Emerging as a Key Regulator

doi: 10.1016/j.mcpro.2026.101509

Figure Lengend Snippet: Inhibition of Dock2 by CPYPP attenuates tubular injury, inflammatory infiltration and interstitial fibrosis after renal IRI in mice. A , schematic of the experimental design for unilateral I/R in mice treated with CPYPP or vehicle via i.p. injection following I/R induction. B , measurement of Scr levels. C , H&E staining was performed to assess the effect of pharmacological inhibition of Dock2 by CPYPP on renal tissue architecture in mice at 3 days and 28 days post-I/R. Inflammatory cells, tubular casts, and dilated tubules are indicated by solid arrows , hollow arrows , and triangles , respectively. The scale bar represents 100 μm. D , tubular injury score at 3 days post-I/R (quantified from C ). E – I , western blot ( E ) and quantitative analysis ( F – I ) of renal cortical Havcr1/Kim1, IL-6, TNF-α, and MCP-1 expression in sham-operated and 3 days post-I/R mice, with or without CPYPP treatment. J , immunofluorescent staining for the macrophage marker F4/80 in renal cortical sections to assess inflammatory infiltration. The scale bar represents 40 μm. K , the quantitative analysis of the F4/80 immunofluorescence staining from ( J ). L – N , western blot ( L ) and quantitative analysis ( M and N ) of renal cortical FN and α-SMA expression in sham-operated and 28 days post-I/R mice, with or without CPYPP treatment. O , Masson's trichrome staining was used to evaluate the effect of CPYPP on renal collagen deposition at 28 days post-I/R, with quantification data shown in ( P ). The scale bar represents 100 μm. Data are presented as mean ± SEM. n.s, not significant; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. CPYPP, 4-[3′-(2″-chlorophenyl)-2′-propen-1′-ylidene]-1-phenyl-3,5-pyrazolidinedione; Dock2, dedicator of cytokinesis 2; FN, fibronectin; Havcr, hepatitis A virus cellular receptor 1; IL-6, interleukin-6; IRI, ischemia-reperfusion injury; Kim1, kidney injury molecule-1; MCP-1, monocyte chemoattractant protein-1; α-SMA, alpha-smooth muscle actin; Scr, serum creatinine; TNF-α, tumor necrosis factor-α.

Article Snippet: Following blocking, the sections were incubated overnight at 4 °C with an anti-Havcr1/Kim1 primary antibody (R&D Systems, cat. AF1817, dilution 1:200), then treated with horseradish peroxidase (HRP)-conjugated secondary antibody at room temperature (RT) for 1 h, and finally developed with 3,3′-diaminobenzidine (DAB) substrate (ZSGB, ZLI-9017).

Techniques: Inhibition, Injection, Staining, Western Blot, Expressing, Marker, Immunofluorescence, Virus