rabbit derived antibodies  (R&D Systems)

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: CTGF knockdown in Vero cells reduces autophagy and adhesion and promotes short-term suspension adaptation

doi: 10.3389/fbioe.2026.1777187

Figure Lengend Snippet: Vero cells are activated for autophagy by EBSS. (A) Schematic diagram of the EBSS-induced starvation model in Vero cells. (B) LC3 tagged with mCherry and GFP: yellow fluorescence indicates no autophagy (both fluorophores active); red fluorescence indicates autophagy (GFP quenched in acidic conditions, mCherry retained). (C) GFP quenching and mCherry retention under starvation induction, indicating autophagy. (D) RT-qPCR detection of autophagy gene expression at various stages under starvation. (E) Western blot analysis of autophagy marker proteins LC3 and P62 under starvation induction. (F,G) Densitometric analysis confirmed the increased conversion of LC3-I to LC3-II and the gradual decrease of p62 in Vero cells post-autophagy (n = 3 per group; *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001, ns: p > 0.05). (F) Quantitative analysis of LC3-II. (G) Quantitative analysis of P62. (H) Under hunger-induced conditions, TEM (Transmission Electron Microscopy) was used to observe the formation of autophagosomes and mitophagy.

Article Snippet: TRIzol, Evo M-MLV Reverse Transcription Premix Kit, and qRT-PCR Kit were acquired from Hunan Accura Biology Co., Ltd. BCA Protein Quantification Kit and EdU Cell Proliferation Kit with Alexa Fluor 555 were obtained from Dalian Meilun Biotechnology Co., Ltd. RIPA lysis buffer, protease inhibitor, 5× SDS-PAGE protein loading buffer, and DAPI (4′,6-diamidino-2-phenylindole dihydrochloride) staining mounting medium were purchased from Beijing Solarbio Science and Technology Co., Ltd. Rabbit-derived LC3, rabbit-derived P62, mouse-derived β-actin, and mouse-derived GAPDH antibodies were obtained from Hangzhou Hua’an Biotechnology Co., Ltd. Rabbit-derived Bax and Bcl-2 antibodies were purchased from Abmart Medical Technology Co., Ltd. Horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG and HRP-labeled goat anti-mouse IgG were acquired from Beijing Zhongshan Jinqiao Biotechnology Co., Ltd. TRITC Phalloidin was obtained from Yeasen Biotechnology Co., Ltd. Hoechst 33,342 was purchased from AAT Bioquest (Sunnyvale, CA, USA).

Techniques: Fluorescence, Quantitative RT-PCR, Gene Expression, Western Blot, Marker, Transmission Assay, Electron Microscopy

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: CTGF knockdown in Vero cells reduces autophagy and adhesion and promotes short-term suspension adaptation

doi: 10.3389/fbioe.2026.1777187

Figure Lengend Snippet: Knock down of CTGF in Vero Cells reduce Autophagy Capacity. (A) Western blot analysis of autophagy marker proteins P62 and LC3 after CTGF knockdown (shCTGF). (B,C) Densitometric analysis confirmed the decreased conversion of LC3-I to LC3-II and the gradual accumulation of p62 following knockdown. (B) Quantitative analysis of LC3-II. (C) Quantitative analysis of P62. (D) Lysosome formation detected by LysoBrite Red staining after CTGF knockdown (shCTGF) and (E) its quantitative results. (F) Cathepsin B activity detected by Magic Red staining after CTGF knockdown (shCTGF) and (G) its quantitative results. (n = 3 per group; *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001, ns: p > 0.05). (H) Autophagy levels in knockdown cells under starvation conditions were observed by TEM. Green arrows indicate the occurrence of autophagy.

Article Snippet: TRIzol, Evo M-MLV Reverse Transcription Premix Kit, and qRT-PCR Kit were acquired from Hunan Accura Biology Co., Ltd. BCA Protein Quantification Kit and EdU Cell Proliferation Kit with Alexa Fluor 555 were obtained from Dalian Meilun Biotechnology Co., Ltd. RIPA lysis buffer, protease inhibitor, 5× SDS-PAGE protein loading buffer, and DAPI (4′,6-diamidino-2-phenylindole dihydrochloride) staining mounting medium were purchased from Beijing Solarbio Science and Technology Co., Ltd. Rabbit-derived LC3, rabbit-derived P62, mouse-derived β-actin, and mouse-derived GAPDH antibodies were obtained from Hangzhou Hua’an Biotechnology Co., Ltd. Rabbit-derived Bax and Bcl-2 antibodies were purchased from Abmart Medical Technology Co., Ltd. Horseradish peroxidase (HRP)-labeled goat anti-rabbit IgG and HRP-labeled goat anti-mouse IgG were acquired from Beijing Zhongshan Jinqiao Biotechnology Co., Ltd. TRITC Phalloidin was obtained from Yeasen Biotechnology Co., Ltd. Hoechst 33,342 was purchased from AAT Bioquest (Sunnyvale, CA, USA).

Techniques: Knockdown, Western Blot, Marker, Staining, Activity Assay

Journal: bioRxiv

Article Title: Multimodality Molecular Profiling Nominates Targetable Mechanisms in Progressive RV Dysfunction

doi: 10.64898/2026.03.09.710504

Figure Lengend Snippet: RV endothelial and pericyte abundances and colocalization patterns were associated with cardiomyocyte hypoxia signaling and with progressive RV compromise. (A) UMAP visualizing endothelial cells and pericytes in snRNAseq data. (B) Endothelial cell and (C) pericyte relative abundances were similar between mild RVD and control, but elevated in severe RVD. (D) Representative image of RV sections stained with isolectin B4 (IB4) showing capillary density is higher in severe RVD (white arrows: capillaries, yellow: WGA, purple: IB4), quantified as (E) the percent area occupied by endothelial cells. (F) Representative image of RV sections stained with IB4 and platelet derived growth factor receptor beta (PDGFRβ) to mark endothelial cells and pericytes, respectively. Endothelial cell-pericyte localization was altered in severe RVD (white arrows: co-localized endothelial cells and pericytes, red arrow: pericytes not located near endothelial cells, orange: IB4, blue: PDGFRβ) quantified with a colocalization coefficient. (H) Cardiomyocyte HIF1A expression was highest in severe RVD. One way ANOVA with Tukey’s post-hoc, means shown in B-C, E, G.

Article Snippet: Pericyte-endothelial cell colocalization was calculated by staining OCT-embedded RV tissue sections with primary antibody to platelet-derived growth factor receptor beta (PDGFRβ, Cell Signaling Technology, 3169S) at a 1:50 dilution.

Techniques: Control, Staining, Derivative Assay, Expressing

Journal: The American Journal of Pathology

Article Title: Matrix Gla Protein Expression in Pericytes and Myofibroblasts Contributes to Renal Fibrosis

doi: 10.1016/j.ajpath.2025.12.003

Figure Lengend Snippet: Pericytes express Mgp in the kidney. A: Scheme represents Rosa Tomato/+ ;Mgp-Cre mice generation. Cre recombinase expression following the Mgp gene expression deleted the stop cassette and expressed red fluorescent protein (RFP) fluorescence. B: Fluorescence microscopy showing the endogenous RFP expression in tubular interstitial cells of control ( Rosa Tomato/+ ) and Rosa Tomato/+ ;Mgp-Cre kidneys. Nuclei were stained with Hoechst (blue). Bottom panels: Magnified images of yellow dotted boxed areas are presented. C: Mgp target sequence, the sequence of the guide RNA (blue), and single-stranded oligodeoxynucleotide (ssODN) to introduce the desired insertion. The ssODN carries homology arms flanking each side of the target nucleotide with hemagglutinin (HA) sequences ( yellow boxed area ). As shown, the guide RNA anneals on the stop codon (TAG) of Mgp ( red boxed area ), complementary to the protospacer adjacent motif ( green circle ), where co-injected clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) is expected to generate a double-stranded break. D: Left: A representative agarose gel image of genotyping PCR using HA/F3 and HA/R3 primers. Wild-type (WT) Mgp allele produces a 297-bp fragment, and HA- Mgp allele produces a 324-bp fragment. Right: Sanger sequencing analyses confirm the insertion of the HA sequence (yellow) before the stop codon (red). E: Confocal microscopy showing matrix Gla protein (MGP)–expressing cells in a healthy kidney. Tissues were stained using rabbit anti-CD31 or rabbit anti–platelet-derived growth factor receptor (PDGFR)-β with mouse anti-HA antibody, followed by anti-rabbit Alexa 555 (green or white) and anti-mouse Alexa 647 (red) antibodies. Nuclei were stained with Hoechst (blue). Right: Magnified images of yellow dotted boxed areas are presented. Scale bars: 10 μm ( B , bottom panels, and E , left ); 20 μm ( B , top panels ); 2 μm ( E , right ). IRES, internal ribosome entry site.

Article Snippet: The following primary antibodies were used: anti-HA tag antibody (1:50; 2367; Cell Signaling Technology, Danvers, MA), anti-CD31 (1:200; 77699; Cell Signaling Technology), anti–platelet-derived growth factor receptor β (1:50; 3169; Cell Signaling Technology), and anti–α smooth muscle actin (SMA) (1:200; 19245; Cell Signaling Technology).

Techniques: Expressing, Gene Expression, Fluorescence, Microscopy, Control, Staining, Sequencing, Introduce, Injection, CRISPR, Agarose Gel Electrophoresis, Confocal Microscopy, Derivative Assay

Journal: The American Journal of Pathology

Article Title: Matrix Gla Protein Expression in Pericytes and Myofibroblasts Contributes to Renal Fibrosis

doi: 10.1016/j.ajpath.2025.12.003

Figure Lengend Snippet: Loss of matrix Gla protein (MGP) suppresses the extracellular matrix expression and kidney fibrosis. A: Sirius red–stained histologic sections and quantification of stained area confirmed the collagen fibers in wild-type (WT) or Mgp –/– kidneys at 2 weeks after the folic acid (FA) administration. For quantification, 10 fields were examined from each sample. B: Quantitative real-time PCR analysis of fibrotic marker gene expression levels in WT and Mgp –/– kidneys before and after FA (or vehicle) administration in reference to Hprt . C: Confocal microscopy showing pericytes ( arrowheads ) in the WT and Mgp –/– kidneys. Tissues were stained using rabbit anti–platelet-derived growth factor receptor (PDGFR)-β, followed by anti-rabbit Alexa 555 (red). Nuclei were stained with Hoechst (blue). Right: The bar graph represents the number of pericytes over total interstitial cells per field. For each sample, 10 fields were examined. D: Quantitative real-time PCR analysis of Hey1 expression levels in WT and Mgp –/– kidneys before or 2 weeks after the FA administration in reference to Hprt . A – D: Statistical analysis: t -test ( A and C ) or two-way analysis of variance, followed by Bonferroni correction ( B and D ), was used to calculate P . The results are presented as means ± SEM ( A – D ). n = 4 for each group ( A and C ); n = 3 for each group ( B ); n = 3 to 5 for each group ( D ). ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗∗ P < 0.0001. Scale bars: 100 μm ( A , left ); 20 μm ( C ). HPF, high-power field; NS, not significant.

Article Snippet: The following primary antibodies were used: anti-HA tag antibody (1:50; 2367; Cell Signaling Technology, Danvers, MA), anti-CD31 (1:200; 77699; Cell Signaling Technology), anti–platelet-derived growth factor receptor β (1:50; 3169; Cell Signaling Technology), and anti–α smooth muscle actin (SMA) (1:200; 19245; Cell Signaling Technology).

Techniques: Expressing, Staining, Real-time Polymerase Chain Reaction, Marker, Gene Expression, Confocal Microscopy, Derivative Assay