The treated lung tissues or BEAS‐2B cells were lysed in radio‐immunoprecipitation assay (RIPA) buffer for 20 min, and supernatants were collected. The BCA assay kit was applied to determine the concentrations of proteins. Further, 20 μg proteins were loaded into the lanes of 10% sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS‐PAGE) before electroblotting onto polyvinylidene fluoride (PVDF) membranes (Bio‐Rad). After blocked with 3% BSA (Sigma‐Aldrich) at room temperature for 1 h, the membranes were incubated with antibodies against CD9 (Abcam, ab223052), CD61 (R&D, AF2266), CD63 (Abcam, ab217345), CAV1 (Abcam, ab84811), NRF2 (Abcam, ab62352), HO1 (Protein Tech, 27282‐1‐AP), NQO1 (Proteintech, 67240‐1‐Ig), Histone H3 (Abcam, ab176842) and β‐Tubulin (Sigma, SAB4500088) overnight at 4°C. After rinsing, the membranes were then incubated with a HRP‐labelled secondary antibodies that binds primary antibody for 1 h. Finally, the membranes were scanned using electrochemiluminescence (ECL; Millipore) and quantified using ImageJ software.
Ab84811
Manufactured by Abcam
Sourced in United Kingdom
Ab84811 is a laboratory equipment product. It is a high-quality tool designed for use in scientific research and experiments.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Bio-Rad (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Enhanced chemi luminescence (ecl), by Merck Group (1 mentions)
Ab62352, by Proteintech (1 mentions)
Ab217345, by Abcam (1 mentions)
Loading buffer, by Beyotime (1 mentions)
Ab22683, by Abcam (1 mentions)
Odyssey instrument, by LI COR (1 mentions)
Ab6046, by Abcam (1 mentions)
Ab150435, by Abcam (1 mentions)
Ab177952, by Abcam (1 mentions)
3 protocols using ab84811
1
Protein Expression Analysis in Lung Cells
2
Protein Expression Analysis in Neonatal Rat Cardiomyocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Whole-cell proteins were isolated from neonatal rat ventricular myocytes. Samples
containing 100 μg of protein in 10 µL of loading buffer (Beyotime Biotechnology, Shanghai,
China) were loaded and separated by electrophoresis on 8% sodium-dodecyl sulfate
polyacrylamide gels. Then, proteins in the gels were transferred to polyvinylidene
fluoride membranes and blocked for 1 hour using 5% nonfat milk, followed by incubation
with primary antibodies to calreticulin (ab22683, Abcam, Cambridge, UK, observed band size
65 kDa) and CaV1.3 (ab84811, Abcam, observed band size 245 kDa) overnight at
4°C on a shaker. The membranes were washed three times with 0.05% phosphate buffered
saline plus 0.05% Tween and incubated with corresponding fluorescent secondary antibodies
(926-32211 and 926-32210, LI-COR Biosciences, Lincoln, NE, USA) for 1 hour in the dark at
room temperature. Finally, the bands on the membranes were detected with the Odyssey
instrument (LI-COR Biosciences), and Odyssey software v1.2 was used to analyze and
quantify the bands. The intensities of proteins were normalized to the respective β-actin
intensity of each gel.
containing 100 μg of protein in 10 µL of loading buffer (Beyotime Biotechnology, Shanghai,
China) were loaded and separated by electrophoresis on 8% sodium-dodecyl sulfate
polyacrylamide gels. Then, proteins in the gels were transferred to polyvinylidene
fluoride membranes and blocked for 1 hour using 5% nonfat milk, followed by incubation
with primary antibodies to calreticulin (ab22683, Abcam, Cambridge, UK, observed band size
65 kDa) and CaV1.3 (ab84811, Abcam, observed band size 245 kDa) overnight at
4°C on a shaker. The membranes were washed three times with 0.05% phosphate buffered
saline plus 0.05% Tween and incubated with corresponding fluorescent secondary antibodies
(926-32211 and 926-32210, LI-COR Biosciences, Lincoln, NE, USA) for 1 hour in the dark at
room temperature. Finally, the bands on the membranes were detected with the Odyssey
instrument (LI-COR Biosciences), and Odyssey software v1.2 was used to analyze and
quantify the bands. The intensities of proteins were normalized to the respective β-actin
intensity of each gel.
3
Cardiac Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Western Blot was used to study the protein expression of SERCA2a/RyR2/CaV1.3/NCX/β-tubulin (β-tubulin, Abcam, ab6046; SERCA2a, Abcam, ab150435; RyR2, Abcam, ab21796; Cav1.3, Abcam, ab84811; and NXC, Abcam, ab177952). The total protein was obtained from the SANCs. Protein samples are separated on 8% and 12% SDS-PAGE gel and transferred to nitrocellulose membranes. Transfer the protein to the PVDF membrane, block it with 5% skimmed milk powder at room temperature for 1 hour, and then incubate it with the primary antibody overnight at 4°C. The cells were incubated with the first antibody overnight. The ImageJ software was used to analyze the data by optical density analysis.
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