Tumors were fixed in formalin for 24 hours, paraffin embedded, sectioned and stained according to standard procedures. Briefly, slides were baked at 60°C for 1 hour then deparaffinized and rehydrated. After antigen retrieval in citrate buffer, endogenous peroxidases were inactivated by 3% hydrogen peroxide. Non-specific signals were blocked using Rodent Block M (Biocare RBM961). Then slides were stained using VIM antibody (Cell Signaling; 5741) overnight at 4°C. After overnight incubation, the slides were washed and incubated with secondary antibody (HRP-polymers, Biocare RMR622) for 30 min at room temperature. The slides were washed and stained with DAB substrate (ThermoFisher Scientific). The slides were then counterstained with hematoxylin and mounted with mounting medium (Richard-Allan Scientific). Cells were counted using imageJ based on a minimum of 9 20x images from each sample. P values were calculated using a wilcoxon test.
Vim antibody
Manufactured by Cell Signaling Technology
The VIM antibody is a tool used in research laboratories to detect and study the expression of the vimentin protein. Vimentin is an intermediate filament protein that plays a role in the structure and function of cells. The VIM antibody can be used in various immunoassay techniques, such as Western blotting, immunohistochemistry, and immunofluorescence, to investigate the presence and localization of vimentin in different cell types and tissues.
Automatically generated - may contain errors
Lab products found in correlation
Mounting medium, by Thermo Fisher Scientific (2 mentions)
Rodent block m, by Biocare Medical (2 mentions)
Rmr622, by Biocare Medical (2 mentions)
Dab substrate, by Thermo Fisher Scientific (2 mentions)
Peroxidase conjugated goat anti rabbit antibody or goatanti mouse antibody, by Merck Group (1 mentions)
Flag antibody, by Cell Signaling Technology (1 mentions)
Ecl immunoblotting kit, by Beyotime (1 mentions)
Gapdh antibody, by Santa Cruz Biotechnology (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
3 protocols using vim antibody
1
Immunohistochemical Analysis of Tumor Samples
2
Immunohistochemical Analysis of Tumor Samples
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Tumors were fixed in formalin for 24 hours, paraffin embedded, sectioned and stained according to standard procedures. Briefly, slides were baked at 60°C for 1 hour then deparaffinized and rehydrated. After antigen retrieval in citrate buffer, endogenous peroxidases were inactivated by 3% hydrogen peroxide. Non-specific signals were blocked using Rodent Block M (Biocare RBM961). Then slides were stained using VIM antibody (Cell Signaling; 5741) overnight at 4°C. After overnight incubation, the slides were washed and incubated with secondary antibody (HRP-polymers, Biocare RMR622) for 30 min at room temperature. The slides were washed and stained with DAB substrate (ThermoFisher Scientific). The slides were then counterstained with hematoxylin and mounted with mounting medium (Richard-Allan Scientific). Cells were counted using imageJ based on a minimum of 9 20x images from each sample. P values were calculated using a wilcoxon test.
3
Quantitative Protein Expression Analysis
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For the detection of protein, cytoplasm and nuclear protein extracts were prepared from cells. The protein concentration of each sample was determined using a NanodropTM spectrophotometer (Thermo Scienti c). Protein (100ug) from each sample was examined bySDS-PAGE(4% stacking and 10% separating gels) and then transferred overnight on to PVDF membranes(Millipore). The membranes were immunoblotted with the following: poly clonal anti-human TJP1 antibody (1:200, Abgent); CDH2 antibody (1:1000, Abcam); VIM antibody (1:1000, Cell Signaling Technology); EPS15 antibody (1:500, Abgent); GAPDH antibody (1:1000, Santa Cruz Biotechnology); Flag antibody (1:1000, Cell Signaling Technology) overnight. The blots were then incubated with peroxidase-conjugated goat anti-rabbit antibody or goatanti-mouse antibody (1:4000, Millipore) for 1 h. The PVDF membranes were subsequently subjected to immunoblotting analysis using an ECL immunoblotting kit according to the manufacturer's recommended protocol (Beyotime Institute of Biotechnology, China).
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