Anti α sma ab5694

Manufactured by Abcam

Sourced in United Kingdom

Anti-α-SMA (ab5694) is a primary antibody that binds to alpha-smooth muscle actin (α-SMA), a cytoskeletal protein found in vascular smooth muscle cells and other cell types. This antibody can be used in various research applications to detect and identify α-SMA-expressing cells.

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α-SMA (863 citations) Ab5694 (709 citations) Anti-α-SMA (354 citations) Anti-SMA (22 citations) α-SMA (ab5694 (15 citations) Anti-α-SMA antibody (ab5694) (3 citations) -SMA (ab5694), (2 citations)

7 protocols using anti α sma ab5694

Immunofluorescence Analysis of Tumor Cells

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Immunofluorescence analysis on tumor cells and tissues was performed as described [23 (link),24 (link)]. Staining was carried out with the following primary antibodies: anti-αSMA (ab5694), anti-TNC (ab108930), and anti-Collagen-I (ab34710) all from Abcam (Cambridge, UK), and revealed by Alexa Fluor 488-conjugated secondary antibody. All images were captured with a Leica TCS SP5 AOBS confocal laser-scanning microscope (Leica Microsystems, Wetzlar, Germany). Immunofluorescence acquisition settings were kept constant within each cell line or tumor tissue. Mean fluorescence intensity (MFI) was evaluated with ImageJ software, measuring the mean pixel intensity in each channel, background subtracted. MFI was normalized on DAPI tumors.

Modica C., Olivero M., Zuppini F., Milan M., Basilico C, & Vigna E. (2021). HGF/MET Axis Induces Tumor Secretion of Tenascin-C and Promotes Stromal Rewiring in Pancreatic Cancer. Cancers, 13(14), 3519.

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Protein Extraction and Immunoblotting Protocol

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Preparation of whole cell lysates and kidney tissue homogenates, and immunoblotting were performed as previously described [32 (link)]. The primary antibodies were obtained as follows: anti-α-SMA (ab5694; Abcam), anti-FN (ab2413; Abcam), anti-col-I (ab138492; Abcam), and anti-E-cadherin (sc-7870; Santa Cruz Biotechnology or ab15148; Abcam). Anti-p-Smad2 (Ser465/467)/Smad3 (Ser423/425) (8828), anti-p-Erk1/2 (Thr202/Tyr204) (9101), and anti-p-p38MAPK (Thr180/Tyr182) (4511) were obtained from Cell Signaling Technology (Danvers, MA, USA).

Wu Y., Wang L., Deng D., Zhang Q, & Liu W. (2017). Renalase Protects against Renal Fibrosis by Inhibiting the Activation of the ERK Signaling Pathways. International Journal of Molecular Sciences, 18(5), 855.

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Immunohistochemical Analysis of PDAC Samples

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Four-µm sections of paraffin slides obtained from patients with PDAC were provided from the pathology department. Primary antibodies—anti-CD3 (ab5690), anti-CD8 (ab17147), anti-CD20 (ab78237), anti-CD66b (ab197678), and anti-α-sma (ab5694)—were purchased from Abcam PLC (Cambridge, UK). A Masson-Goldner staining kit (100485) for collagen staining was bought from Merck KGaA (Darmstadt, Germany). Secondary antibodies—horse anti-mouse IgG (BA-2000) and horse anti-rabbit IgG (BA-1100)—were purchased from VECTOR Laboratories (Burlingame, CA, USA), along with the Avadin/Biotin Blocking Kit (SP-2001), the alkaline phosphatase enzyme detection system (ABC-AP) (AK-5000), and the ImmPACT Vector Red Alkaline Phosphatase Substrate Kit (SK-5105).

Miksch R.C., Schoenberg M.B., Weniger M., Bösch F., Ormanns S., Mayer B., Werner J., Bazhin A.V, & D’Haese J.G. (2019). Prognostic Impact of Tumor-Infiltrating Lymphocytes and Neutrophils on Survival of Patients with Upfront Resection of Pancreatic Cancer. Cancers, 11(1), 39.

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Immunohistochemical Profiling of Macrophage Activation

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BLM was purchased from Tokyo Chemical Industry (Tokyo, Japan). Anti-α-SMA (ab5694) and anti-CD163 antibodies (ab182422) were purchased from Abcam PLC (Cambridge, UK). Anti-CD68 antibody (MCA1957GA) was purchased from AbD Serotec (Raleigh, NC, USA). Anti-NOS2 antibody (SC-650) was purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). Anti-phospho-SMAD3 antibody (44-246G), anti-Arginase-I antibody (PA5-29645), and anti-TLR2 antibody (PA5-20020) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Anti-vimentin antibody (MAB-2105SP) was purchased from R&D Systems (Minneapolis, MN, USA). Mouse enzyme-linked immunosorbent assay (ELISA) kits for TNF-α (430904) and INF-γ (430804) were purchased from BioLegend (San Diego, CA, USA). The QuickZyme Total Collagen Assay kit was purchased from QuickZyme Biosciences (Leiden, The Netherlands).

Park G.T., Yoon J.W., Yoo S.B., Song Y.C., Song P., Kim H.K., Han J., Bae S.J., Ha K.T., Mishchenko N.P., Fedoreyev S.A., Stonik V.A., Kim M.B, & Kim J.H. (2021). Echinochrome A Treatment Alleviates Fibrosis and Inflammation in Bleomycin-Induced Scleroderma. Marine Drugs, 19(5), 237.

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Comprehensive Kidney Tissue Immunostaining Protocol

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For immunofluorescence staining of kidney tissues, the antibodies used were guinea pig anti-Nephrin (clone GP-N2, Progen, Heidelberg, Germany), mouse anti-synaptopodin (clone G1D4, Progen), and rat anti-mouse CD44 (clone IM7, BD Biosciences). For immunohistochemistry, the antibodies used were rat anti-F4/80 (ab6640, Abcam, Cambridge, UK), rabbit anti-WT1 (clone C-19, Santa Cruz Biotechnology, CA, USA), rabbit anti-α-SMA (ab5694, Abcam), and mouse anti-PCNA (clone PC10, DAKO). All antibodies were diluted at 1:100 in DAKO antibody diluent (Agilent, CA, USA). The following polyclonal rabbit antibodies used for primary reaction of immunoblots were obtained from Cell Signaling Technology (MA, USA): anti-phosphorylated Smad1/5/8 (#95115), anti-phosphorylated AMPK (#2531), anti-AMPK (#2532), anti-phosphorylated p38 (#4511), anti-p38 (#9212), anti-phosphorylated ERK (#4370), anti-ERK (#4695), anti-phosphorylated STAT3 (#9145), anti-phosphorylated p53 (#9284), and anti-phosphorylated mTOR (#2971). Rabbit antibodies anti-α-SMA (ab5694) and anti-NRF2 (ab31163) were from Abcam. Rabbit antibodies anti-STAT3 (clone C-20), anti-p53 (clone FL393), and anti-vinculin (clone H-300) were obtained from Santa Cruz Biotechnology.

Omachi K., Kaseda S., Yokota T., Kamura M., Teramoto K., Kuwazuru J., Kojima H., Nohara H., Koyama K., Ohtsuki S., Misumi S., Takeo T., Nakagata N., Li J.D., Shuto T., Suico M.A., Miner J.H, & Kai H. (2021). Metformin ameliorates the severity of experimental Alport syndrome. Scientific Reports, 11, 7053.

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Quantitative Histological Analysis of Kidney Fibrosis

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Paraffin sections mounted on Superfrost Ultra Plus Adhesion Slides (Thermo Fisher Scientific Inc, Waltham, MA, USA) were deparaffinized and rehydrated in ethanol. Desmin, αSMA and fibronectin immunohistochemistry was performed with polyclonal antibodies (anti-desmin MS 376-S1, Thermo Fisher, 1:1000; anti- αSMA ab5694, Abcam, 1:1000; anti-FN HPA0027066, Atlas Antibodies, 1:1000), using the avidin–biotin method and developed by incubation with diaminobenzidine (Vector Laboratories, Burlingame, CA, USA). Pictures were taken with Zeiss AxioCam 512 of the stained sections for further analysis. Glomerular tuft was delineated manually and standard glomerulus size parameters (e.g., Feret diameter) as wells as the PAS, desmin, and αSMA positive area were determined using the CellProfiler cell image analysis software.
Lipid deposition was determined by oil-Red-O (O0625, Sigma-Aldrich, Budapest, Hungary) staining of 5-μm-thick cryosections.

Bukosza E.N., Kaucsár T., Godó M., Lajtár E., Tod P., Koncsos G., Varga Z.V., Baranyai T., Tu N.M., Schachner H., Sőti C., Ferdinandy P., Giricz Z., Szénási G, & Hamar P. (2019). Glomerular Collagen Deposition and Lipocalin-2 Expression Are Early Signs of Renal Injury in Prediabetic Obese Rats. International Journal of Molecular Sciences, 20(17), 4266.

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Immunohistochemical Analysis of Mouse Kidneys

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Mouse kidneys were promptly perfused with 4% paraformaldehyde and fixed in paraffin for sectioning after removal. Three-micron-thick kidney sections were mounted for immunohistochemical staining. Cells that had grown on glass coverslips were fixed with 4% paraformaldehyde for 10 min at room temperature. Then, the cells were treated with 5% Triton for 30 min. Primary antibodies were incubated at 4 °C overnight. The next day, the tissues and cells were incubated with secondary antibodies under the guidance of the immunohistochemical staining protocol. The antibodies used were as follows: anti-CXCR7 (SAB2700198, Sigma), anti-fibronectin (F3648; Sigma–Aldrich), anti-α-SMA (ab5694; Abcam), and anti-β-catenin (610154; BD Biosciences, ab15180; Abcam). The secondary antibodies used were as follows: AffiniPure donkey anti-rabbit IgG (H + L) (711-005-152; Jackson ImmunoResearch) and AffiniPure donkey anti-mouse IgG (H + L) (711-005-150; Jackson ImmunoResearch). DAB (SK-4100) and AEC (SK-4200) substrate kits were obtained from Vector Laboratories. An Olympus microscope was used to observe and photograph the sections.

Meng P., Liu C., Li J., Fang P., Yang B., Sun W, & Zhang Y. (2024). CXC chemokine receptor 7 ameliorates renal fibrosis by inhibiting β-catenin signaling and epithelial-to-mesenchymal transition in tubular epithelial cells. Renal Failure, 46(1), 2300727.

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