Colon tissue sections (4 mm) were deparaffinized in xylene twice for 10 min, rehydrated in a graded ethanol series once for 5 min, incubated in sodium citrate buffer for 10 min for antigen retrieval, blocked with 10% bovine serum albumin for 1 h, and incubated with antibodies against Claudin-1 (1:100; Abcam, Cambridge, UK; ab242370), zonula occludens 1 (ZO-1) (1:100; Abcam; ab276131), Occludin (1:100; Abcam; ab216327), mucin 2 (MUC2) (1:500; Abcam; ab272692), myeloperoxidase (MPO) (1:500; Abcam; ab208670), CD11b (1:100; Abcam; ab133357), IL-17 (2 mg/mL; Abcam; ab79056), IL-10 (10 mg/mL; Abcam; ab189392) overnight, and IL-1β (1:500; Abcam: ab283818). The sections were then incubated with secondary antibodies (SP9000; Zsbio Biotechnology, Beijing, China) for 1 h. The images were acquired using a BX53F microscope (Olympus, Tokyo, Japan).
Ab242370
Manufactured by Abcam
Sourced in United Kingdom
Ab242370 is a primary antibody. It is a purified rabbit monoclonal antibody that recognizes a specific protein target. The product is intended for use in various research applications.
Automatically generated - may contain errors
Lab products found in correlation
Ab272692, by Abcam (2 mentions)
Ab216327, by Abcam (2 mentions)
Bx53f microscope, by Olympus (1 mentions)
Ab79056, by Abcam (1 mentions)
Ab133357, by Abcam (1 mentions)
Ab208670, by Abcam (1 mentions)
Ab276131, by Abcam (1 mentions)
Ab189392, by Abcam (1 mentions)
Ab283818, by Abcam (1 mentions)
Ab221547, by Abcam (1 mentions)
Ab16667, by Abcam (1 mentions)
3 3 diaminobenzidine kit, by ZSGB-BIO (1 mentions)
Phosphatase inhibitor, by GenDEPOT (1 mentions)
Ripa buffer, by Merck Group (1 mentions)
Sc 47778, by Santa Cruz Biotechnology (1 mentions)
Protease inhibitor cocktail solution, by GenDEPOT (1 mentions)
Ab53032, by Abcam (1 mentions)
Ab15102, by Abcam (1 mentions)
Nbp2 14977, by Novus Biologicals (1 mentions)
Dp2 bsw software, by Olympus (1 mentions)
5 protocols using ab242370
1
Immunohistochemical Analysis of Colon Tissue
2
Immunohistochemical and Histopathological Analysis of Mouse Intestine
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The small intestinal tissue of mice was fixed in 4% paraformaldehyde for 24 h, embedded in paraffin, and cut into sections at a thickness of 5 μm (The histopathology associated index of intestine was evaluated though the criterion in Supplementary Table 1 ). For immunohistochemical staining, the intestinal sections were subjected to deparaffinization, hydration, antigen retrieval, quenching of endogenous peroxidase, and blocking procedures. All slices were then incubated with the primary antibodies against pSTAT3 (CST #9145) and Ki-67 (Abcam, England, Ab16667) at 4°C overnight followed by incubation with biotinylated secondary antibodies for 30 min and visualization using a 3,3′-Diaminobenzidine Kit (ZSGB-BIO, Beijing, China). For immunofluorescence staining for the detection of MUC2 (Abcam, England, Ab272692), Lgr5 (NBP1-28904SS), ZO-1 (Abcam, England, Ab221547), and Claudin-1 (Abcam, England, Ab242370), the colonic sections were processed by the methods described above. Periodic Acid-Schiff stain (PAS) staining (Biossci, China) was performed following the manufacturer’s protocols.
3
Western Blot Analysis of Protein Expression
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Penis tissue, MCPs, and MCP-EVs were lysed in RIPA buffer (Sigma-Aldrich) supplemented with protease inhibitor cocktail Solution (Ca# P3100-010, GenDEPOT, Katy, TX, USA) and phosphatase inhibitors (Ca# P3200-005, GenDEPOT). An equal amount of protein (50 µg/lane) from these samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) on 8-15% gels and then transferred to polyvinylidene difluoride membranes. After blocking with 5% nonfat dry milk for 1 hour at room temperature, membranes were probed with antibodies against Hebp1 (Ca# NBP2-14977, 1:100; NOVUS Biologicals), claudin-1 (Ca# ab242370, 1:1000; Abcam), claudin-2 (Ca# ab53032, 1:1000; Abcam), claudin-3 (Ca# ab15102, 1:1000; Abcam), claudin-11 (Ca# 36-4500, 1:1000; Thermo Fisher Scientific), and/or β-actin (Ca# sc-47778, 1:5000; Santa Cruz Biotechnology Inc.) for 2 hours. After washing three times, signals were visualized using an ECL detection system (Ca# EBP-1073, Bionote Inc., Hwaseong-si, Gyeonggi-do, Korea). Results were quantified densitometrically using an image analysis system (Image J 1.34; NIH).
4
Quantifying Claudin-1 Expression by IHC
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Immunohistochemistry was performed as previously described by Tian et al (Tian et al., 2020 (link)). Specifically, the slides were dewaxed sequentially, endogenous peroxidase was removed, antigen‐binding sites were exposed, and sections were exposed and permeabilized according to a previously described technique. Then, the sections were treated overnight at 4 °C with claudin‐1 antibodies (1:200, Abcam, #ab242370). The antigen–antibody response sites were detected for 30 min using HRP‐labeled secondary antimouse IgG (1:1000, Abcam, #ab205719), followed by staining with DAB (GeneTex). The nuclei were counterstained with hematoxylin. A fluorescein microscope (Olympus) equipped with DP2‐BSW software was used to acquire digital images at 400× magnification. ImageProline Plus 5.1 (Media Cybernetics) was used for image processing and analysis. The relative abundance of Claudin‐1 was calculated by dividing (integrated optical density) by the area.
5
Western Blot Analysis of Tight Junction Proteins
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Mouse tissues were homogenized in RIPA buffer containing protease and phosphatase inhibitor cocktails (Beyotime) using a tissue homogenizer (Wonbio, Shanghai, China) according to the manufacturer's instructions. Cells were lysed in RIPA buffer (P0013B, Beyotime) containing protease and phosphatase inhibitor cocktails (P1049, Beyotime). After centrifugation, the supernatants were collected and quantified using a bicinchoninic acid (BCA) protein assay kit (P0011, Beyotime). Protein lysates were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (20325ES62, Yeasen), transferred to 0.45 μm polyvinylidene difluoride membranes (Millipore, Burlington, MA, USA), blocked in 5% nonfat milk in PBS plus Tween-20 (0.1% [v/v]) buffer, and incubated with primary antibodies at 4 °C overnight. The blot was incubated with secondary antibodies for 1 h, and detection used Clarity Western ECL Substrate (Biorad, Hercules, CA, USA) with imaging using a Tanon fluorescence imaging system. ImageJ software package was used for densitometry analysis, and band density was normalized to β-actin expression. Antibody details were: anti-ZO-1 (ab190085, Abcam, 1:5000), anti-occludin (ab216327, Abcam, 1:5000), anti-claudin-1 (ab242370, Abcam, 1:5000), and anti-β-actin (3700, Cell Signaling Technology, 1:5000).
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