We followed the methods of Wang et al. [21 (link)]. After blocking, the aorta sections were incubated overnight at 4°C with a rabbit anti-AGE antibody (1 : 200, ab23722, Abcam, Cambridge, UK), rabbit anti-RAGE antibody (1 : 200, ab3611, Abcam, Cambridge, UK), rabbit anti-Nox4 antibody (1 : 150, ab133303, Abcam, Cambridge, UK), mouse anti-3-nitrotyrosine antibody (1 : 200, ab61392, Abcam, Cambridge, UK), rabbit anti-NF-κB p65 antibody (1 : 200, D14E12, CST, Beverly, MA, USA), or mouse anti-Glo1 antibody (1 : 200, MA1-13029, Invitrogen, Waltham, MA, USA). The sections were washed and then incubated with a secondary antibody (1 : 200, peroxidase-conjugated anti-rabbit (ZB-2301) or anti-mouse (ZB-2305) antibody, ZSGB-BIO, Beijing, China) for 1 hour. Color was developed using DAB. Images were captured using an Olympus DP71 microscope. The mean IOD of staining (IOD/area) from 5 random fields on one section was assessed using Image-Pro Plus 6.0 software.
Ma1 13029
Manufactured by Thermo Fisher Scientific
Sourced in United States
The MA1-13029 is a laboratory product manufactured by Thermo Fisher Scientific. It is a core piece of equipment designed to perform a specific function within the laboratory environment. However, due to the need to maintain an unbiased and factual approach without extrapolation, a detailed description of the product's core function cannot be provided. The information available is limited, and providing any further details would involve interpretation or speculation, which is not within the scope of this request.
Automatically generated - may contain errors
Lab products found in correlation
Zb 2301, by ZSGB-BIO (2 mentions)
Zb 2305, by ZSGB-BIO (2 mentions)
Ab3611, by Abcam (2 mentions)
Ab133303, by Abcam (2 mentions)
Ecl western blotting substrate, by Thermo Fisher Scientific (2 mentions)
Ta 09, by ZSGB-BIO (2 mentions)
Dp71 microscope, by Olympus (1 mentions)
Ab23722, by Abcam (1 mentions)
Ab61392, by Abcam (1 mentions)
Image pro plus 6, by Media Cybernetics (1 mentions)
P0013, by Beyotime (1 mentions)
Ab89443, by Abcam (1 mentions)
Ab11079, by Abcam (1 mentions)
Kgp1100, by Keygen Biotech (1 mentions)
3 protocols using ma1 13029
1
Immunohistochemical Analysis of AGE-RAGE Pathway
2
Western Blot Analysis of Protein Targets
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The solubilized protein samples were boiled with loading buffer for 5 min. Equal amounts of protein (30 μg) were separated using 12% SDS-polyacrylamide gel electrophoresis and electrotransferred onto a nitrocellulose membrane. The membrane was blocked with 5% skim milk and incubated with a rabbit anti-RAGE antibody (1 : 1000, ab3611, Abcam, Cambridge, UK), rabbit anti-Nox4 antibody (1 : 1000, ab133303, Abcam, Cambridge, UK), rabbit anti-NF-κB p65 antibody (1 : 1000, D14E12, CST, Beverly, MA, USA), or mouse anti-Glo1 antibody (1 : 3000, MA1-13029, Invitrogen, Waltham, MA, USA) overnight at 4°C. After a thorough washing, the membrane was incubated with a secondary antibody (1 : 5000, peroxidase-conjugated anti-rabbit (ZB-2301) or anti-mouse (ZB-2305) antibody, ZSGB-BIO, Beijing, China) for an hour at room temperature. β-Actin was used as the loading control (1 : 3000 TA-09, ZSGB-BIO, Beijing, China). The bands were visualized using ECL Western Blotting Substrate (Thermo Fisher Scientific, Hudson, NH, USA) and quantified by Image-Pro Plus 6.0 software.
3
Quantification of Nrf2 and Glo1 Proteins
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To obtain total protein samples, frozen samples of kidney cortex were homogenized in ice‐cold radioimmunoprecipitation assay lysis buffer (P0013; Beyotime, Beijing, China) with 1% protease inhibitors cocktail and phenylmethane sulfonyl fluoride. The protein concentration of the supernatants was determined by the bicinchoninic acid assay kit and adjusted to the same level in all samples. The separation of nuclear and cytosol protein was carried out according to the manufacturer's instructions (KGP1100; KeyGEN BioTECH, Beijing China).
The method of western blot has been previously reported elsewhere24 . The membrane was incubated overnight with a primary mouse anti‐Nrf2 antibody (1:1,000 ab89443; Abcam, Cambridge, UK) or mouse anti‐Glo1 antibody (1:3,000, MA1‐13029; Invitrogen, Waltham, MA, USA) at 4°C. After washing, the membrane was incubated with a secondary antibody (1:5,000 HRP‐coupled anti‐mouse antibody, ZB‐2305; ZSGB‐BIO, Beijing, China) for 1 h at room temperature. The membrane was probed for β‐actin (1:3,000, TA‐09; ZSGB‐BIO) as a loading control for total protein or cytoplasmic protein. Histone was used as a loading control for nuclear protein (1:3,000, ab11079; Abcam). The bands were visualized using the ECL Western Blotting Substrate (Thermo Fisher, Hudson, NY, USA), and quantified with Image‐Pro Plus 6.0 software (Media Cybernetics, Rockville, MD, USA).
The method of western blot has been previously reported elsewhere
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