Maraviroc was obtained from Sigma-Aldrich (St. Louis, MO, USA) (for in vitro experiments) or GlaxoSmithKline (Brentford, UK) (for in vivo experiments). Mouse CCL3 was obtained from Peprotech (Rocky Hill, NJ, USA). The following antibodies were used as the primary antibodies for immunohistochemical analyses: goat anti-CCR5 antibody (Santa Cruz Biotechnology, Dallas, TX, USA), rat anti-Ly6G antibody (BD Biosciences, San Jose, CA, USA), rat anti-F4/80 antibody (Serotec, Kidlington, UK), mouse anti-α-SMA antibody (Dako, Glostrup, Denmark), rabbit anti-type I collagen antibody, rabbit anti-CD31 antibody and rabbit anti-EGF antibody (Abcam, Cambridge, UK). The following rat anti-mouse antibodies were used as the primary antibodies for the flow cytometric analysis: anti-CD11b antibody (BD Biosciences), anti-CD25 antibody (BioLegend, San Diego, CA, USA), anti-CD45 antibody (eBioscience, San Diego, CA, USA), anti-F4/80 antibody (eBioscience), anti-Foxp3 antibody (eBioscience), anti-Ly6G antibody (Gr-1) (Tonbo Biosciences, San Diego, CA, USA), anti-MIP-1α antibody (R&D Systems, Minneapolis, MN, USA), anti-CCR5 antibody (BioLegend), and isotype-matched control IgGs for individual rat antibodies (BD Biosciences).
Rabbit anti cd31 antibody
Manufactured by Abcam
Sourced in United Kingdom, United States
Rabbit anti-CD31 antibody is a primary antibody that recognizes the CD31 (PECAM-1) protein. CD31 is a cell adhesion molecule expressed on the surface of endothelial cells and is commonly used as a marker for blood vessel endothelium.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 710 confocal microscope, by Zeiss (3 mentions)
Axio scan z1, by Zeiss (2 mentions)
Rabbit anti rfp antibody, by Tebu-Bio (2 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (2 mentions)
Ripa buffer, by Thermo Fisher Scientific (2 mentions)
Anti foxp3 antibody, by Thermo Fisher Scientific (1 mentions)
Anti cd45 antibody, by Thermo Fisher Scientific (1 mentions)
Maraviroc, by Merck Group (1 mentions)
Anti cd11b antibody, by BD (1 mentions)
Anti f4 80 antibody, by Thermo Fisher Scientific (1 mentions)
Rat anti f4 80 antibody, by Bio-Rad (1 mentions)
Goat anti ccr5 antibody, by Santa Cruz Biotechnology (1 mentions)
Mouse anti α sma antibody, by Agilent Technologies (1 mentions)
Rabbit anti type 1 collagen antibody, by Abcam (1 mentions)
Anti cd25 antibody, by BioLegend (1 mentions)
Anti ccr5 antibody, by BioLegend (1 mentions)
Te2000 u fluorescence microscope system, by Nikon (1 mentions)
Alexa fluor 568 goat anti rabbit antibody, by Thermo Fisher Scientific (1 mentions)
Dm4000 b led fluorescence microscope, by Leica (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Abcam (1 mentions)
17 protocols using rabbit anti cd31 antibody
1
Maraviroc and Mouse Chemokine Analyses
2
Quantifying Tumor Angiogenesis via CD31
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After dewaxing and antigen recovery, the tissue sections were blocked with 5% BSA for 30 minutes at room temperature and stained with rabbit-anti-CD31 antibody (Abcam, Cambridge, UK) at a dilution of 1:20 overnight at 4°C. CD31, a cell marker that is constitutively expressed in the cells lining the endothelium, plays a role in tumor angiogenesis and growth. The next day, the sections were incubated with the goat anti-rabbit IgG diluted secondary antibody (Jackson ImmunoResearch, West Grove, PA, USA) for 1 hour and DAPI for 5 minutes at room temperature in the dark. After washing, photographs were immediately captured with the Nikon TE2000-U fluorescence microscope system (Nikon Instruments Inc., Melville, NY, USA). The results are shown as the mean number of CD31 positive cells ± SD.
3
Immunofluorescent Staining of Xenograft Tumors
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Frozen sections from xenograft tumours were cut as 7 µm thick sections and fixed in 4% paraformaldehyde solution in PBS (ThermoFisher) for 10 min at room temperature. Fixed tumour tissues were washed in TBS and permeabilised with TBS with Triton X-100 (TBS/T, 0.1% v/v) for 30 min at room temperature. Tissues were blocked with 10% normal goat serum (NGS) in 1% BSA in TBS/T for 1 h at room temperature before incubating with rabbit anti-CD31 antibody (Abcam) diluted 1:50 in 1% BSA in TBS/T, overnight at 4 °C. Sections were washed in TBS/T and incubated with Alexafluor 568 goat anti-rabbit antibody (ThermoFisher) at 1:750 dilution in 1% BSA in TBS/T for one hour at room temperature. Sections were washed with TBS and mounted with mounting medium with DAPI (Abcam). Images were taken at 20x magnification with Leica DM4000 B LED Fluorescence microscope.
4
Western Blot Analysis of Tight Junction Proteins
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Cells were washed three times with sterile PBS, and cell lysates were obtained using radioimmunoprecipitation assay (RIPA) buffer (ThermoFisher Scientific) plus protease inhibitor cocktail (Roche). The proteins in the lysates were separated by SDS-PAGE on a 5–20% polyacrylamide gel (Wako) and were then transferred to polyvinylidene fluoride membranes (Merk-Millipore). After blocking with 5% skim milk in TBS containing 0.1% Tween 20 at room temperature for 1 h, the membranes were incubated with a mouse anti-claudin-5 antibody (diluted 1/250; ThermoFisher Scientific), a mouse anti-P-gp antibody (diluted 1/250; GeneTex), a rabbit anti-CD31 antibody (diluted 1/1000; abcam), or a mouse anti-ß-actin antibody (diluted 1/5000; Sigma-Aldrich) at 4 oC overnight, and then with a HRP-conjugated anti-mouse IgG (Cell Signaling Technology) or a HRP-conjugated anti-rabbit IgG (Cell Signaling Technology) at room temperature for 1 h. The bands were detected with ECL Plus western blotting detection reagents (ThermoFisher Scientific), and the signals were visualized with a LAS-4000 imaging system (Fuji Film).
5
Comprehensive Biomaterial Characterization for Cell Studies
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CS was purchased from Aladdin biochemical technology Co., Ltd. (Shanghai, China); Doxorubicin hydrochloride was purchased from Meilun Biotechnology (Dalian, China); BSA (Albumin Bovine V) was purchased from BioFroxx (Einhausen, Germany); Soybean oil for injection was purchased from Yuanye Bio-Technology Co., Ltd. (Shanghai, China); Solutol HS15 was purchased from BASF (Ludwigshafen, Germany); 3-(4,5-Dimethyl-2-thiazolyl)−2,5-diphenyl-2H-tetrazolium bromide (MTT), 4,6-diamidino-2-phenylindole (DAPI), monensin sodium and genistein were purchased from J&K Scientific Ltd.; Mouse antibodies including anti-CD44, FITC anti-CD44 and Rhodamine-labeled secondary antibodies were purchased from Affymetrix eBioscience (San Diego, USA). rabbit anti-CD31 antibody and Alexa Fluor 647 conjugated goat anti-rabbit IgG were purchased from Abcam (Cambridge, UK). Other reagents were analytical or HPLC grade and purchased commercially.
6
Immunofluorescent Staining of FFPE Brain
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Sections of 5-µm FFPE tumor-containing brain tissues were processed after heat-induced antigen retrieval with Tris–EDTA (pH 9.0). Sections were stained with DyLight488-labeled lectintomato together with rabbit anti-CD31 antibody (Abcam, ab 28364; dilution 1:1000) or rabbit anti-RFP antibody (Tebu-Bio, cat. no. 600-901-397; dilution 1:500). The rabbit antibodies were detected with AlexaFluor594-labeled anti-rabbit as secondary antibody. Alternatively, slides were stained with hematoxylin/eosin. Image acquisition was done with a Zeiss Axio Scan Z1 slide scanner.
7
Immunofluorescence Staining of OPCs, ECs, and EPCs
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Cells were fixed at room temperature for 15 min with 4% paraformaldehyde (PFA) and rinsed with 0.01 mol/l PBS. After blocking non-specific antigens with 5% BSA, OPCs were treated with mouse monoclonal anti-A2B5 antibody (1:200, Sigma, USA) and ECs and EPCs were incubated with rabbit anti-CD31 antibody (1:100, Abcam, USA) and rabbit anti-CD133 antibody (1:100, Abcam, USA) overnight at 4 °C, respectively. After washing with PBS, Dylight-488- or Dylight-649-conjugated goat anti-mouse IgG (H + L) secondary antibodies (1:500, Abbkine, USA) were added at room temperature for 2 h. The nucleus was stained with 4′,6-diamidino-2-phenylindole (DAPI, 1:200, Beyotime, China) for 10 min. The images were taken under an Olympus Bx60 fluorescence microscope (Olympus, TKY, Japan).
8
Immunohistochemical Analysis of Tissue Markers
9
Immunofluorescence Staining of Lung Sections
10
Spinal Cord TLR4 Activation Imaging
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Double immunofluorescence analysis was performed to measure the activation and localization of TLR4 after I/R injury. The spinal cord was fixed and sectioned into 10-μm slices with a Leica CM3050 S cryostat. The sections were blocked with 10% bovine serum albumin for 1 h at room temperature and incubated overnight at 4°C with the primary antibodies: mouse anti-TLR4 (1:100, Abcam, Cambridge, UK), rabbit anti-CD31 antibody (1:800, Abcam), rabbit anti-CD13 antibody (1:500, Abcam), rabbit anti-Iba-1 antibody (1:800, Wako, Germany), and rabbit anti-GFAP antibody (1:800, Abcam). After incubation with Alexa 594-conjugated donkey anti-mouse IgG (1:500, Molecular Probes, Eugene, Oregon, USA) and Alexa 488-conjugated donkey anti-rabbit IgG (1:500, Molecular Probes) for 2 h at room temperature, the images were captured using a Leica TCS SP2 (Leica Microsystems, Buffalo Grove, IL, USA) laser scanning microscope and photographed by the attached digital camera to determine the number of immunoreactive cells. Non-specific staining was determined by omitting the primary antibody. The data were expressed as numbers of positive cells/area/spinal section ± standard error mean (SEM).
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