To extract protein from cells, Ripa buffer with 50x protease inhibitor was used to lyses cells. We then used 10% sodium dodecyl sulfate (SDS) gel wells to isolate proteins at 80 V in the start and 120 V when samples reached into separating gel, and then transferred protein into nitrocellulose membranes (NC, Millipore). At the end of transfer, NC membranes were washed in PBS and incubated in 5% BSA to block protein at room temperature for 1 h. After incubated with primary antibodies overnight at 4°C, the membranes were washed by PBST following the incubation with secondary antibody for 1 h. Protein levels were evaluated by using infrared imaging system LI‐COR Odyssey. (Odyssey, LI‐COR). The following antibodies were used: anti‐MCM4 (#12973, 1:1000 dilution), anti‐PCNA (#13110, 1:1000 dilution), anti‐Caspase‐3 (#9662, 1:1000 dilution), anti‐Cleaved Caspase‐3 (#9664, 1:1000 dilution), anti‐P21 (#2947, 1:1000 dilution) and anti‐GAPDH (#5174, 1:10000 dilution) purchased from Cell Signalling Technology.
Anti mcm4
Manufactured by Cell Signaling Technology
Anti-MCM4 is a primary antibody that binds to the MCM4 (Minichromosome Maintenance Complex Component 4) protein. MCM4 is a subunit of the MCM complex, which is essential for the initiation and regulation of DNA replication in eukaryotic cells.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Merck Group (2 mentions)
Anti actin, by Merck Group (2 mentions)
Goat anti mouse hrp, by Merck Group (2 mentions)
Goat anti rabbit hrp, by Cell Signaling Technology (2 mentions)
Crescendo ecl substrate, by Merck Group (2 mentions)
Anti smad2 3, by Cell Signaling Technology (2 mentions)
Anti mcm2, by Cell Signaling Technology (2 mentions)
Anti p21, by Santa Cruz Biotechnology (2 mentions)
Anti mcm2, by Abcam (2 mentions)
Anti androgen receptor, by Abcam (2 mentions)
Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Anti gapdh, by Cell Signaling Technology (1 mentions)
Anti pcna, by Cell Signaling Technology (1 mentions)
Anti p21, by Cell Signaling Technology (1 mentions)
Anti caspase 3, by Cell Signaling Technology (1 mentions)
3 protocols using anti mcm4
1
Protein Extraction and Western Blot
2
Placental and Embryonic Protein Isolation and Immunoblotting
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Protein was isolated from E13.5 placentas and embryos by acetone
precipitation from RNA-isolation buffer (Buffer RLT or TRK) and resuspending in
SUTEB loading buffer (8M Urea, 1% SDS, 10mM EDTA, 10mM Tris-HCl, pH 6.8).
Protein lysates were run on 4–20% SDS-PAGE acrylamide gels and
transferred to PVDF membrane (Millipore). Immunoblots were probed with anti-Mcm2
(Epitomics/Abcam), anti-Mcm2(Cell Signaling Technology), anti-androgen receptor
(Epitomics/Abcam), anti-SMAD2/3(Cell Signaling), anti-p21(Santa Cruz),
anti-MCM4(Cell Signaling Technology), anti-actin (Sigma). Secondary antibodies
used included goat anti-rabbit-HRP (Cell Signaling) and goat anti-mouse-HRP
(Sigma). Crescendo ECL substrate(Millipore) was used and immunoblots digitally
scanned using a cDigit scanner. Quantification of immunoblots was performed
using ImageStudio software.
precipitation from RNA-isolation buffer (Buffer RLT or TRK) and resuspending in
SUTEB loading buffer (8M Urea, 1% SDS, 10mM EDTA, 10mM Tris-HCl, pH 6.8).
Protein lysates were run on 4–20% SDS-PAGE acrylamide gels and
transferred to PVDF membrane (Millipore). Immunoblots were probed with anti-Mcm2
(Epitomics/Abcam), anti-Mcm2(Cell Signaling Technology), anti-androgen receptor
(Epitomics/Abcam), anti-SMAD2/3(Cell Signaling), anti-p21(Santa Cruz),
anti-MCM4(Cell Signaling Technology), anti-actin (Sigma). Secondary antibodies
used included goat anti-rabbit-HRP (Cell Signaling) and goat anti-mouse-HRP
(Sigma). Crescendo ECL substrate(Millipore) was used and immunoblots digitally
scanned using a cDigit scanner. Quantification of immunoblots was performed
using ImageStudio software.
3
Placental and Embryonic Protein Isolation and Immunoblotting
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