Cells were washed twice in PBS, collected, lysed and boiled in SDS sample buffer at 99°Cfor 5 min. Equal amount of proteins of each cell extracts were resolved by 10–12% Bis-Tris SDS-PAGE and transferred to polyvinylidene difluoride membrane (PVDF, Millipore, Billerica, MA). Non-specific binding was blocked by incubation in 5% skim milk in TBST at room temperature for 1–2 h. Blots were then probed with various primary antibodies, AcH3 (06-599, Millipore, Billerica, MA), H3K9me3 (ab8898, Abcam, Cambridge, UK), Zscan4 (#5114, custom-made), Histone H3 (ab1791, Abcam, Cambridge, UK) and β-actin (P30002, Abmart, Shanghai, China) by overnight incubation at 4°C in 5% skim milk in TBST. Immunoreactive bands were then probed for 1–2 h at room temperature with the appropriate horseradish peroxidase-conjugated secondary anti-Rabbit IgG-HRP (GE Healthcare, NA934V, Piscataway, NJ). The protein bands were detected by Enhanced ECL Amersham™ prime Western blotting detection reagent (GE Healthcare, RPN2232).
Zscan4
Manufactured by Abcam
Sourced in United Kingdom
Zscan4 is a laboratory equipment product offered by Abcam. Zscan4 is a protein that plays a role in the regulation of gene expression and cellular development.
Automatically generated - may contain errors
Lab products found in correlation
Histone h3, by Abcam (1 mentions)
β actin p30002, by Abmart (1 mentions)
Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions)
H3k9me3, by Abcam (1 mentions)
Amersham ecl, by Cytiva (1 mentions)
Sds page gel, by Thermo Fisher Scientific (1 mentions)
Immobilon p membrane, by Merck Group (1 mentions)
Enhanced chemi luminescence (ecl), by Cytiva (1 mentions)
H3k4me3, by Diagenode (1 mentions)
Gapdh, by Santa Cruz Biotechnology (1 mentions)
2 protocols using zscan4
1
Protein Expression and Western Blot Analysis
2
Western Blot Analysis of Oocyte Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Oocytes were washed in PBS and frozen at −80°C. Thirty oocytes were lysed in Reducing SDS loading buffer and denatured for 5 min. Proteins were separated by gradient precast 4-12% SDS-PAGE gel (Thermo Fisher) and transferred to Immobilon P membrane (Millipore) by the semi-dry blotting system. Membranes were blocked by 5% skimmed milk for 1 h and incubated with H3K4me3 (Diagenode, 1541003; 1:500), H3 (Abcam, ab1791; 1:500), Zscan4 (Abcam, 97748; 1:250) and Gapdh (Santa Cruz Biotechnology, 97166; 1:500) primary antibodies diluted in 1% milk/TTBS overnight. The membranes were incubated in peroxidase donkey anti-rabbit secondary antibody (Jackson ImmunoResearch, 711-035-152, 1:10,000) for 1 h at room temperature. Proteins were visualised by chemiluminescence using ECL (Amersham), and imaged and quantified using an Azure 600 Imager.
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