Total protein was extracted from cells using RIPA buffer (Beyotime, Shanghai, China) in the presence of a protease inhibitor mixture (Pierce Chemical, Dallas, Texas, USA) and quantified using a BCA protein assay kit (Thermo Scientific, Waltham, USA). Western blotting was performed according to standard procedures. The following primary antibodies were used: anti– periostin (Proteintech, #66,491– 1– lg, 1:3000), anti– ERK1/2 (CST, #9102, 1:1000), anti– phospho ERK1/2 (CST, #4370, 1:1000), anti– P38 (CST, #9212, 1:1000), anti– phospho P38 (CST, #9215, 1:1000), and anti– β– actin (Affinity Biosciences, #AF7018, 1:1000).
Anti periostin
Manufactured by Proteintech
Sourced in China
Anti–periostin is a laboratory equipment product that can be used to detect and measure the levels of periostin, a protein involved in various biological processes. The core function of this product is to provide researchers with a tool to analyze the expression and distribution of periostin in their experiments.
Automatically generated - may contain errors
Lab products found in correlation
Anti tgf β, by Abcam (2 mentions)
Anti il 1β, by Bioworld Technology (2 mentions)
Anti tnf α, by Bioworld Technology (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Ripa buffer, by Beyotime (1 mentions)
Anti β actin, by Affinity Biosciences (1 mentions)
Dm2500 fluorescence microscope, by Leica camera (1 mentions)
Alexa fluor 647 goat anti rabbit igg secondary antibody, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Triton x 100, by Merck Group (1 mentions)
Anti calnexin, by Cell Signaling Technology (1 mentions)
Anti gapdh, by CWBIO (1 mentions)
Anti β catenin, by Cell Signaling Technology (1 mentions)
Anti α sma, by Bioworld Technology (1 mentions)
Anti cd81, by Cell Signaling Technology (1 mentions)
Anti histone, by Cell Signaling Technology (1 mentions)
Anti cd63, by Cell Signaling Technology (1 mentions)
Anti tsg101, by Cell Signaling Technology (1 mentions)
Anti galectin 3, by Abcam (1 mentions)
4 protocols using anti periostin
1
Protein Extraction and Western Blotting
2
Immunofluorescence Staining of Heart Slices
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For immunofluorescence staining 5-μm-thick heart slices were blocked and permeabilized with a blocking and permeabilization solution made of 5% FBS and 0.1% Triton-X100 (Sigma-Aldrich) in PBS for 1 h at RT and stained with 1/200 polyclonal rabbit IgG to caveolin-1 (#3238S; Cell Signaling Technology, Danvers, MA), 1/100 monoclonal mouse IgG anti-4 hydroxynonenal antibody (ab48506; Abcam, Cambridge, UK) or 1/400 anti-periostin (Proteintech, Manchester, UK), in the same solution overnight at 4 °C. Heart slices were then washed and stained with an Alexa Fluor® 647 goat anti-rabbit IgG secondary antibody (Invitrogen, Carlsbad, CA, USA) or Alexa Fluor® 488 goat anti-mouse IgG (Invitrogen, Carlsbad, CA, USA), respectively for 2 h at RT in the dark. Heart sections were washed and counterstained with 200 ng/mL DAPI (ThermoFisher Scientific) for 10 min in the dark. The heart slices were finally washed and mounted with FluorSaveTM reagent (CalbioChem, San Diego, CA, USA). Slides were imaged with a Leica DM2500 fluorescence microscope and analyzed with ImageJ software.
3
Comprehensive Protein Expression Analysis
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Cells and exosomes were lysed with RIPA lysis buffer containing protease and phosphatase inhibitors. Nuclear proteins were extracted with a kit (CWBIO, Beijing, China). Protein samples were separated by SDS-PAGE, and then transferred to PVDF membranes (Millipore, Burlington, MA, USA). After blocking for 1 h, the PVDF membranes were incubated with primary antibodies overnight at 4℃. The primary antibodies used were anti-CD81 (1：1,000; Cell Signaling Technology, Danvers, MA, USA), anti-CD63 (1：1,000; Cell Signaling Technology), anti-TSG-101 (1：1,000; Cell Signaling Technology), anti-calnexin (1：1,000; Cell Signaling Technology)，anti-Galectin-3 (1：1,500; Abcam, Cambridge, UK), anti-α-SMA (1：1,000; Bioworld Technology, Bloomington, MN, USA), anti-Collagen Ⅰ (1：1,000; Bioworld Technology), anti-Pe-riostin (1：1,000; Proteintech, Wuhan, China), anti-IL-1β (1：1,000; Bioworld Technology), anti-TNF-α (1：1,000; Bioworld Technology), anti-TGF-β (1：1,000; Abcam), anti-β-catenin (1：1,000; Cell Signaling Technology), anti-GAPDH (1：3,000; CWBIO, Beijing, China), and anti-Histone (1：1,000; Cell Signaling Technology). The next day, membranes were incubated at 37℃ for 1 h with hor-seradish peroxidase-linked goat anti-rabbit or anti-mouse antibodies (1：3,000; ABM, Richmond, Canada).
4
Immunohistochemical Analysis of Myocardial Tissues
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Myocardial tissues were fixed with 4% paraformaldehyde, embedded in a paraffin block, and cut into 3-μm sections. After a series of deparaffinization and dehydration steps, endogenous peroxidase activity was blocked with 3% H2O2, and then freshly prepared 0.01 mol/l citrate buffer was used for antigen retrieval. After washing three times with PBS, the sections were blocked with 5% BSA for 30 min, and then incubated with the following primary antibodies: anti-α-SMA (1：200; MAXIM, Fujian, China), anti-Periostin (1：200; Proteintech), anti-Collagen I (1：200; Boster, Wuhan, China), anti-IL-1β (1：200; Bioworld Technology), anti-TNF-α (1：200; Bioworld Technology), and anti-TGF-β (1：300; Abcam) overnight at 4℃. Then the sections were incubated with biotin-la-beled goat anti-mouse/rabbit secondary antibodies at 37℃ for 30 min. DAB chromogenic solution was used for color development, and hematoxylin was used for counter staining. Immunohistochemical images were captured with a Panoramic Scanner MIDI (3DHISTECH, Budapest, Hungary).
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