The nuclear fractions and lysates from the untreated Skov-3 and GO/CDDP-treated Skov-3 cells were resuspended into 500 μL RIPA buffer and the protein concentration was quantified using the Bradford protein assay (Pierce). Same amount of proteins was subjected to immunoprecipitation using LC3B MAb (Cell Signaling, cat. no. 3868) and Dynabeads® Protein A (Thermo Fisher, cat. 10001D). The immunocomplex associated with LC3 was separated by 12% SDS-PAGE and the bands that were apparently upregulated after GO/CDDP treatment were cut for Liquid Chromatography/Mass Spectrometry (LC/MS/MS) analysis. The LC3-immunocomplex was also subjected to SDS-PAGE and subsequent Western blot using the following primary antibodies: rabbit MAb (1:500 dilution) specific for histone H4 (D2X4V, Cell Signaling) or acetyl-histone H4(Lys16) (Cell Signaling), mouse MAb specific for histone H1 (Merck) or rabbit polyclonal Ab specific for phosphorylated histone H1 (Merck). The secondary antibodies were HRP-conjugated goat anti-rabbit (Genetex) or anti-mouse (SeraCare) IgG. The images were developed using the chemiluminescence reagent (Western Lightning® ECL Pro, PerkinElmer) and captured using the GeneGnome system (Syngene). The band intensities were quantified using Image Pro Plus 6.0.
Hrp conjugated goat anti rabbit
Manufactured by GeneTex
HRP-conjugated goat anti-rabbit is a secondary antibody used in immunoassay techniques. It is produced by conjugating horseradish peroxidase (HRP) to goat-derived antibodies that specifically bind to rabbit primary antibodies. This antibody can be used to detect and quantify the presence of rabbit-derived target proteins or antigens in samples.
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Lab products found in correlation
Dynabeads protein a, by Thermo Fisher Scientific (1 mentions)
Western lightning ecl pro, by PerkinElmer (1 mentions)
Genegnome system, by Syngene (1 mentions)
Total β catenin, by Cell Signaling Technology (1 mentions)
Phospho β catenin, by Cell Signaling Technology (1 mentions)
Dulbecco s modified eagle medium dmem, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Matrigel coating, by BD (1 mentions)
2 protocols using hrp conjugated goat anti rabbit
1
Quantitative Histone Modification Analysis
2
Investigating Cell Migration and Invasion
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Dulbecco’s Modified Eagle Medium (DMEM) was purchased from Life Technologies (Carlsbad, CA). 8.0 micron Transwell dishes with Matrigel coating for invasion assays or without ECM coating for the migration assays were purchased from BD Biosciences (San Jose, CA). MRx102 was a kind gift from MyeloRx LLC (Vallejo, CA).
WIF1 antibody (MABN722) was purchased from EMD/Millipore (Temecula, CA). The GAPDH, phospho β-catenin, and total β-catenin, and p53 antibodies were purchased from Cell Signaling (Danvers, MA). The phosphor Akt and total Akt antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX). HRP-conjugated goat anti-rabbit and goat anti-mouse secondary antibody were purchased from Genetex (Irvine, CA).
WIF1 antibody (MABN722) was purchased from EMD/Millipore (Temecula, CA). The GAPDH, phospho β-catenin, and total β-catenin, and p53 antibodies were purchased from Cell Signaling (Danvers, MA). The phosphor Akt and total Akt antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX). HRP-conjugated goat anti-rabbit and goat anti-mouse secondary antibody were purchased from Genetex (Irvine, CA).
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