Paxillin antibody

Manufactured by Abcam

Sourced in United Kingdom

Paxillin is a cytoplasmic focal adhesion-associated protein involved in regulating cell spreading, migration, and adhesion. The Paxillin antibody is a tool used to detect and study this protein in biological samples.

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Lab products found in correlation

Goat anti rabbit igg af 647, by Thermo Fisher Scientific (3 mentions) Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (3 mentions) Click it edu cell proliferation assay, by Thermo Fisher Scientific (3 mentions) Yap antibody, by Cell Signaling Technology (3 mentions) β1 integrin antibody, by BD (2 mentions) Cm1950, by Leica (2 mentions) Rabbit anti mouse collagen 1 polyclonal antibody, by Abcam (2 mentions) Af 488 phalloidin to stain actin, by Thermo Fisher Scientific (2 mentions) Lsm 510 meta, by Zeiss (1 mentions) Lsm 710, by Zeiss (1 mentions) Rhodamine conjugated phalloidin, by Thermo Fisher Scientific (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions) Alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Goat anti rabbit igg h l alexa fluor 488 antibody, by Abcam (1 mentions) Hoechst 33342, by Beyotime (1 mentions) Confocal laser scanning microscope, by Zeiss (1 mentions) Goat anti mouse igg h l alexa fluor 647 antibody, by Abcam (1 mentions) Ptbp1 antibody, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

Paxillin (25 citations) Anti-paxillin antibody (7 citations)

6 protocols using paxillin antibody

Immunohistochemistry for Cell Mechanics

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For immunohistochemical staining, media was first removed from the gels. The gels were then fixed with 4% paraformaldehyde in serum free DMEM at 37°C for 30 – 45 minutes. Gels were then washed 3 times in PBS containing calcium (cPBS). Gels were stained following standard immunohistochemistry protocols directly following washing. The following antibodies/reagents were used for immunohistochemistry: YAP antibody (Cell signaling, Catalogue # 4912:, dilution of 1:100), Paxillin antibody (Abcam, Catalogue # ab32084, dilution of 1:200), Prolong Gold antifade reagent with DAPI (Invitrogen), AF-488 Phalloidin to stain actin (Life Technologies, dilution of 1:80), Goat anti-Rabbit IgG AF 647 (Invitrogen, Catalogue # a21245, dilution of 1:500). The Click-IT EdU cell proliferation assay (Invitrogen) was used to identify proliferating cells.

Chaudhuri O., Gu L., Darnell M., Klumpers D., Bencherif S.A., Weaver J.C., Huebsch N, & Mooney D.J. (2015). Substrate stress relaxation regulates cell spreading. Nature communications, 6, 6364.

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Visualization of Actin and Focal Adhesions in hMSCs

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In order to analyze subcellular features of the actin cytoskeleton and focal adhesion, hMSCs were cultured with the microrods still adhered to the silicon wafers with or without MGF for 48 h. Cells were fixed with 4 % paraformaldehyde in phosphate buffered saline (PBS) for 10 min at room temperature, rinsed three times with PBS and permeabilized by 0.1 % Triton X-100 in PBS for 10 min, and washed 3 times with PBS. Cells were pre-incubated in blocking solution (PBS, 1 % bovine serum albumin (BSA)) for 15 min and then incubated with rhodamine conjugated phalloidin (Molecular Probes) at a dilution of 1:400 to stain actin, or paxillin antibody (Abcam) at a dilution of 1:250 for 1.5 h followed by another incubation with secondary antibody Alexa Fluor 488 (Invitrogen) at a dilution of 1:1,000 for 45 min to stain the focal adhesions of the cells. 4′, 6-Diamidino-2-phenylindole (DAPI) (Sigma) was used for nuclear staining, which artificially stained the microrods as well. Confocal images of actin and focal adhesions were obtained with Zeiss LSM 510 META and LSM 710 microscopes.

Doroudian G., Pinney J., Ayala P., Los T., Desai T.A, & Russell B. (2014). Sustained delivery of MGF peptide from microrods attracts stem cells and reduces apoptosis of myocytes. Biomedical microdevices, 16(5), 705-715.

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Quantifying Focal Adhesion and Cytoskeleton

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After 2 and 24 h, HUVECs were fixed with 4% PFA. Adhesion was assessed by staining with a paxillin antibody (1:100; Abcam, Cambridge, UK) using the above-described immunocytochemistry protocol. Projection of the z-stack images were analyzed using ImageJ to calculate the area of paxillin, F-actin (phalloidin), and the co-localization of this focal adhesion protein with the cytoskeleton of the cells.

van Dijk C.G., Louzao-Martinez L., van Mulligen E., Boermans B., Demmers J.A., van den Bosch T.P., Goumans M.J., Duncker D.J., Verhaar M.C, & Cheng C. (2020). Extracellular Matrix Analysis of Human Renal Arteries in Both Quiescent and Active Vascular State. International Journal of Molecular Sciences, 21(11), 3905.

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Immunohistochemical Analysis of 3D Cell Cultures

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For immunohistochemical staining, media was first removed from the gels. The gels were then fixed with 4% paraformaldehyde in serum free DMEM at 37°C for 30 – 45 minutes. Gels were then washed 3 times in PBS containing calcium (cPBS), and incubated overnight in 30% sucrose in cPBS at 4°C. The gels were then placed in a mix of 50% of a 30% sucrose in cPBS solution, and 50% OCT (Tissue-Tek) for several hours. Then the media was removed, the gels were embedded in OCT and frozen. The frozen gels were sectioned with a cryostat (Leica CM1950) to a thickness of 30 – 100 μm, and stained following standard immunohistochemistry protocols. The following antibodies/reagents were used for immunohistochemistry: Rabbit-anti-mouse Collagen I polyclonal antibody (Abcam, cat. # 34710), YAP antibody (Cell signaling, cat. # 4912), Paxillin antibody (Abcam, cat. # 32084), β1 integrin antibody (BD Biosciences, cat. # 550531), Prolong Gold antifade reagent with DAPI (Invitrogen), AF-488 Phalloidin to stain actin (Invitrogen), Goat anti-Rabbit IgG AF 647 (Invitrogen). The Click-IT EdU cell proliferation assay (Invitrogen) was used to identify proliferating cells.

Chaudhuri O., Gu L., Klumpers D., Darnell M., Bencherif S.A., Weaver J.C., Huebsch N., Lee H.P., Lippens E., Duda G.N, & Mooney D.J. (2015). Hydrogels with tunable stress relaxation regulate stem cell fate and activity. Nature materials, 15(3), 326-334.

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Immunofluorescence Staining of Paxillin and PTBP1

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Grow cells in 6-well plates containing confocal slides (1 × 10⁴ cells per well). After washing with PBS, cells were fixed with 4% paraformaldehyde for 15 min, and 0.5%Triton X-100 was permeable for 20 min. After the slides were soaked with PBS, 1%bovine serum albumin was blocked for 30 min, each slides were dripped with sufficient amount of diluted Paxillin antibody (Abcam, UK, Cat. no. ab32084) and incubated overnight at 4℃ in a wet box. Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody (Abcam, UK, Cat. no. ab150077) was added in the dark and incubated in a wet box at 37℃. Then, cells were incubated overnight at 4℃ in a wet box with diluted PTBP1 antibody (Invitrogen, USA, Cat. no. 32-4800), followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 647) antibody (Abcam, UK, Cat. no. ab150115). Hoechst 33342 (Beyotime Biotechnology, China, Cat. no. C1025) was added to the drops and incubated for 5 min, and the specimens were nucleated. The tablets were sealed with a sealing solution containing an anti-fluorescence quench agent, and the acquired images were observed under a confocal laser scanning microscope (Carl Zeiss, Germany) at 200x magnification.

Chu Z., Zhu M., Luo Y., Hu Y., Feng X., Wang H., Sunagawa M, & Liu Y. (2023). PTBP1 plays an important role in the development of gastric cancer. Cancer Cell International, 23, 195.

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Immunohistochemical Analysis of 3D Cell Cultures

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For immunohistochemical staining, media was first removed from the gels. The gels were then fixed with 4% paraformaldehyde in serum free DMEM at 37°C for 30 – 45 minutes. Gels were then washed 3 times in PBS containing calcium (cPBS), and incubated overnight in 30% sucrose in cPBS at 4°C. The gels were then placed in a mix of 50% of a 30% sucrose in cPBS solution, and 50% OCT (Tissue-Tek) for several hours. Then the media was removed, the gels were embedded in OCT and frozen. The frozen gels were sectioned with a cryostat (Leica CM1950) to a thickness of 30 – 100 μm, and stained following standard immunohistochemistry protocols. The following antibodies/reagents were used for immunohistochemistry: Rabbit-anti-mouse Collagen I polyclonal antibody (Abcam, cat. # 34710), YAP antibody (Cell signaling, cat. # 4912), Paxillin antibody (Abcam, cat. # 32084), β1 integrin antibody (BD Biosciences, cat. # 550531), Prolong Gold antifade reagent with DAPI (Invitrogen), AF-488 Phalloidin to stain actin (Invitrogen), Goat anti-Rabbit IgG AF 647 (Invitrogen). The Click-IT EdU cell proliferation assay (Invitrogen) was used to identify proliferating cells.

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