For immunohistochemistry,21 (link) 4-μm thick serial sections were deparaffinized, rehydrated and, after antigen retrieval and nonspecific peroxidase blocking, incubated with mouse monoclonal anti-ICAM-1 (Pierce, IL, USA), anti-eNOS (Pierce), anti-Ki67 (Ventana Medical Systems, AZ, USA), anti-BrdU (YLEM, Avezzano, Italy), anti-human CD31 (Ventana), anti-CD4 (Ventana), and rabbit polyclonal anti-VCAM-1 (Abcam, CB, UK), anti-iNOS (Pierce), and anti-PlGF (Abcam). For rat tissues, mouse monoclonal anti-rat CD31 (BD Pharmingen, NJ, USA), was used. Morphometric evaluation of immunoreactivity was performed according to defined criteria (see Supplementary Materials and Methods ).
Anti plgf
Manufactured by Abcam
Sourced in United Kingdom
Anti-PlGF is a primary antibody that targets placental growth factor (PlGF), a protein involved in angiogenesis and vasculogenesis. This antibody can be used for various research applications, such as immunohistochemistry, Western blotting, and ELISA.
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Lab products found in correlation
Anti vcam 1, by Abcam (1 mentions)
Anti cd4, by Roche (1 mentions)
Anti human cd31, by Roche (1 mentions)
Anti ki 67, by Roche (1 mentions)
Rat anti cd31, by BD (1 mentions)
Anti vimentin, by Abcam (1 mentions)
Anti vegfa, by Abcam (1 mentions)
Anti cd68, by Abcam (1 mentions)
Anti cd31, by Abcam (1 mentions)
Trypsin inhibitor, by Merck Group (1 mentions)
Anti ki67, by Proteintech (1 mentions)
Mouse anti cd31, by Abcam (1 mentions)
Omni ecl femto light chemiluminescence kit, by Ipsen (1 mentions)
Horseradish peroxidase conjugated anti rabbit igg, by Abcam (1 mentions)
Anti vcam 1, by Cell Signaling Technology (1 mentions)
Anti fgf2, by Abcam (1 mentions)
Anti vegf, by Abcam (1 mentions)
4 protocols using anti plgf
1
Immunohistochemical Analysis of Vascular Markers
2
Histological and Immunohistochemical Analysis
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Histology and immunohistochemistry analysis was conducted as described previously [20 (link)]. Briefly, for histology stains, standard H&E was used to assess total cell infiltration. For immunohistochemistry analysis, sections were stained with primary antibodies including anti-VEGFA (Abcam, USA) for vascular endothelial growth factor expression, anti-PlGF (Abcam, USA) for placental growth factor expression, anti-NE (neutrophil elastase) (Abcam, USA) for neutrophils, anti-CD68 (Abcam, USA) for macrophages, anti-CD31 (Abcam, USA) for endothelial cells, anti-vimentin (Abcam, USA) for fibroblasts, and anti-SMC-α actin (Abcam, USA) for smooth muscle cells. Fluorescence microscopy was then used to visualize cell markers using Alexa Fluor 594-conjugated secondary antibodies. Sections were mounted with DAPI-containing mounting media (VECTASHIELD).
3
Immunohistochemical Analysis of ITGA6, Ki67, PLGF and CD31
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Immunohistochemistry was performed using the anti-ITGA6 (Sigma, 1:200), anti-Ki67 (Proteintech, 1:400), anti-PLGF (Abcam, 1:200) and anti-mouse CD31 (Abcam, 1:400) antibodies. Frozen or paraffin-embedded sections were used for immunohistochemical analysis. For immunohistochemical staining, tissue sections were first incubated with 0.1% trypsin at RT for 10 min followed by incubation with 0.1 μg/ml trypsin inhibitor (Sigma) for 5 min. The sections were then rinsed three times in PBS before blocking with 10% normal goat serum (15 min at RT). The sections were rinsed in PBS and incubated with a primary antibody overnight at 4 °C. Afterwards, the tissue sections were rinsed three times in PBS and incubated with the secondary antibody for 60 min at RT. The sections were then immersed in DAB for 5−10 min and counterstained with 10% Mayer’s haematoxylin. ITGA6 expression in tissues was evaluated according to the methods described by R Shao et al 37 (link).
4
Western Blot Analysis of Angiogenic Factors
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Protein extracts were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to polyvinylidene fluoride membranes. Blots were blocked with 5% milk in Tris-buffered saline containing 0.5% Tween-20 for 1 hr at room temperature. The membranes were incubated with primary antibodies at 4°C overnight, followed by incubation with the horseradish peroxidase-conjugated secondary antibodies at 37°C for 2 hr. The immunoreactive bands were visualized using Omni-ECL™ Femto Light Chemiluminescence Kit (Epizyme, China) and imaged by the ChemiDoc XRS Plus luminescent image analyzer (Bio-Rad, USA). The antibodies used in this study were as follows: anti-glyceraldehyde-3-phosphate dehydrogenase (1 : 5,000, Bioworld Technology, USA), anti-VCAM-1 (1 : 1,000, Cell Signaling Technology, USA), anti-VEGF (1 : 1,000, Abcam, UK), anti-PLGF (1 : 1,000, Abcam, UK), anti-FGF2 (1 : 1,000, Abcam, UK), anti-APLN (1 : 1,000, Abcam, UK), anti-cellular communication network factor 2 (CCN2) (1 : 1,000, Abcam, UK), horseradish peroxidase conjugated anti-rabbit IgG (1 : 5,000, Abcam, UK).
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