The primary antibodies used for immunofluorescence and/or western blot hybridisation were γH2AX (rabbit; Active Motif, #39117), γH2AX (rabbit; Abcam, #ab2893), γH2AX (mouse; clone JBW301; Upstate/Millipore, #05–636), BrdU (mouse; clone 131–14871; Chemicon/Millipore, #MAB4072), BrdU (rabbit; Rockland Immunochemicals, #600–401-C29), cyclin B1 (rabbit; Santa Cruz Biotechnology, #sc-752), 53BP1 (rabbit; Santa Cruz Biotechnology, #sc-22760), Rad51 (mouse; Abcam, #ab213), GM-130 (rabbit; Cell Signaling Technology, #12480P), DNA-topoisomerase 1 (rabbit; Abcam, #ab3825), p21 (rabbit; Cell Signaling Technology, #2947P) and histone H3 (rabbit; Abcam, #ab1791). The secondary antibodies conjugated to either Alexa Fluor 488 or Alexa Fluor 594 were purchased from Molecular Probes/Life Technologies; the horseradish peroxidase-conjugated anti-mouse and anti-rabbit IgG were purchased from Amersham/GE Healthcare.
Horseradish peroxidase conjugated anti mouse and anti rabbit igg
Manufactured by Cytiva
Sourced in United States
Horseradish peroxidase-conjugated anti-mouse and anti-rabbit IgG is a secondary antibody conjugate used for detecting primary antibodies in various immunoassays and immunochemical techniques. It provides a sensitive and reliable method for signal amplification and visualization of target proteins or molecules.
Automatically generated - may contain errors
Lab products found in correlation
Immobilion substrate, by Merck Group (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 594, by Thermo Fisher Scientific (1 mentions)
Vinculin, by Merck Group (1 mentions)
Mouse anti ezh2, by BD (1 mentions)
Tubb3, by Santa Cruz Biotechnology (1 mentions)
H3k27me3, by Merck Group (1 mentions)
Actin, by Merck Group (1 mentions)
Histone h3, by Abcam (1 mentions)
Ab9484, by Cell Signaling Technology (1 mentions)
Mouse monoclonal antibody against, by Cell Signaling Technology (1 mentions)
Phosstop, by Roche (1 mentions)
Imagequant las 4000 system, by GE Healthcare (1 mentions)
Nupage, by Thermo Fisher Scientific (1 mentions)
5 protocols using horseradish peroxidase conjugated anti mouse and anti rabbit igg
1
Immunofluorescence and Western Blot Antibodies
2
Western Blot Analysis of Molecular Markers
3
Western Blot Analysis of Phospho-p53
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Western blot analysis was performed as previously described [6 (link)]. For Western blot analysis of phospho-p53, we added phosphatase inhibitors in lysis buffer (PhosSTOP, Roche). The primary antibodies were: rabbit polyclonal antibody against SOX14 (Abcam, Cambridge, UK, ab149047, diluted 1:400), mouse monoclonal anti-α-Tubulin (Calbiochem, MA, USA, CP06, diluted 1:30000), mouse monoclonal antibody against GAPDH (Abcam, Cambridge, UK, ab9484, diluted 1:5000), mouse monoclonal antibody against cleaved PARP (Cell Signaling, Technology, Danvers, MA, USA, Asp214, diluted 1:1000), mouse monoclonal antibody against p53 (DO-1) (Santa Cruz Biotechnology, Texas, USA, sc-126X, diluted 1:1000), rabbit monoclonal antibody against p21Waf1/Cip1 (Cell Signaling, Technology, Danvers, MA, USA, 2947, diluted 1:1000) and rabbit antibody against phospho-p53 (Cell Signaling, Technology, Danvers, MA, USA, diluted 1:1000). Secondary antibodies used for Western blot were: horseradish peroxidase-conjugated anti-mouse and anti-rabbit IgG (Amersham Biosciences, NJ, USA, diluted 1:10000 or Active motif, La Hulpe, Belgium, 1:5000). Immunoreactive bands were detected by chemiluminescence (Immobilion substrate, Millipore, MA, USA). The density of protein bands on Western blots was quantified using ImageJ software.
4
Cell Fractionation and Western Blotting
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Cell fractionations were prepared as described [17 (link)]. Briefly, cells were resuspended in buffer CSK (10mM PIPES, pH = 6.8, 100mM NaCl, 1mM EGTA, 1mM EDTA, 300mM Sucrose, 1.5mM MgCl2, 0.1% Triton-X-100 and protease inhibitors) and incubated in ice for 5min. Samples were centrifuged at 1500g for 5min. Supernatant was collected and stored (soluble fraction). Pellets (insoluble fraction) were washed once in CSK buffer and then resuspended in sample buffer (0.05 M Tris-HCl (pH 6.8), 2% SDS, 6% β-mercaptoethanol) and boiled for 5min. Western blotting was performed as described [64 (link)]. Briefly, cells were lysed in 0.05 M Tris-HCl (pH 6.8), 2% SDS, 6% β-mercaptoethanol and boiled for 5min. SDS-PAGE electrophoresis was done using NuPAGE 3% to 8% Tris-acetate or NuPAGE 4% to 12% Tris-glycine gels (Invitrogen) and proteins were transferred to a nitrocellulose membrane. Primary antibodies were diluted in blocking buffer (5% milk in TBS-Tween 20) and incubated overnight. Horseradish peroxidase—conjugated anti-mouse and anti-rabbit IgG (Amersham) were used as secondary antibodies. Images were acquired using ImageQuant LAS4000 system (GE Healthcare).
5
Quantitative Western Blot Analysis
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Whole cell lysates were isolated as previously described (Popovic et al., 2014) . Proteins were quantified and separated by SDS-PAGE as previously described (Popovic et al., 2014) . After blocking with 5% non-fat milk at +4°C for 24 h, membranes were incubated for 1 h at RT with the following primary antibodies: mouse monoclonal antibody against p53 (DO-1) (Santa Cruz Biotechnology, Texas, USA, sc-126X, diluted 1:1000), rabbit monoclonal antibody against p21 Waf1/Cip1 (Cell Signaling, Technology, Danvers, MA, USA, 2947, diluted 1:1000), mouse monoclonal antibody against α -tubulin (Calbiochem, MA, USA, CP06, diluted 1:30000)). Afterwards, the membranes were incubated for 1 h at RT with the following secondary antibodies: horseradish peroxidase-conjugated anti-mouse and antirabbit IgG (Amersham Biosciences, NJ, USA, diluted 1∶10000). Immunoreactive bands were detected by chemiluminescence (Immobilion substrate, Millipore, MA, USA).
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