Recombinant Anti-PCNA antibody [EPR3821] (ab92552)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.	-Rabbit (1:5000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Upstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.
Protocol tips
-Rabbit (1:5000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Publication protocol

Cultured or transfected cells were harvested and washed twice with PBS. Then, the cells were lysed in ice-cold radio immunoprecipitation assay buffer (Beyotime, Shanghai, People’s Republic of China) containing 0.01% protease and phosphatase inhibitor (Sigma, Shanghai, People’s Republic of China) and incubated on ice for 30 min. Cell lysis was centrifuged at 12,000× g for 10 min at 4°C, and the proteins in supernatant were obtained (20−30 μg) and separated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel, then transferred electrophoretically to a polyvinylidene fluoride membrane (Millipore, Shanghai, People’s Republic of China). The membrane was blocked with 5% bovine serum albumin in tris-buffered saline (TBS) and Polysorbate 20 (also known as Tween 20) and incubated with primary antibodies against ARHGAP1 (Ab194425, 1:1,000 dilution; Abcam), matrix metallopeptidase 2 (MMP2; Ab92536, 1:1,000 dilution; Abcam), zinc finger E-box binding homeobox 1 (ZEB1; Ab124512, 1:1,000 dilution; Abcam), Cyclin B1 (CCNB1; #12231, 1:1,000 dilution; Cell Signaling Technology, Inc.), twist family bHLH transcription factor 1 (Twist; Ab175430, 1:500 dilution; Abcam), proliferating cell nuclear antigen (PCNA; Ab92552, 1:5,000 dilution; Abcam) and GAPDH (#5174, 1:1,500 dilution; CST). Blots were then incubated for 1 h at 37°C with goat anti-mouse or anti-rabbit secondary antibody (Beyotime), and the intensities were measured using enhanced chemiluminescence (Thermo Fisher Scientific Inc.)

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Manufacturer protocol

Download the product protocol from Abcam for Recombinant Anti-PCNA antibody [EPR3821] (ab92552) below.

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