The following antibodies were used: GSK3β (9315, 1:1,000) (Cell Signaling), CD34 (340667, 1:100), ICAM-1 (559771, 1:100) (BD Biosciences, San Jose, CA, USA) and anti CD11a antibody (52895, 1:100) (Abcam, Cambridge, UK). For western blot analysis p65 (sc-372, 1:2,000), c-Rel (sc-71, 1:1,000), p50 (sc-7178, 1:1,000), p105 (sc-7178, 1:1,000), actin (sc-69879, 1:10,000), hnRNPA1 (sc-32301, 1:2,000), tubulin (sc-5546, 1:1,000) histone H3 (sc-10809, 1:1,000), ERK1 and ERK2 antibodies (sc- (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA).
C rel
Manufactured by Santa Cruz Biotechnology
Sourced in United States, United Kingdom
C-Rel is a laboratory equipment product manufactured by Santa Cruz Biotechnology. It is a protein that regulates the expression of genes involved in cellular processes. The core function of C-Rel is to serve as a transcription factor, binding to specific DNA sequences and modulating the transcription of target genes.
Automatically generated - may contain errors
Lab products found in correlation
β actin, by Merck Group (3 mentions)
Anti p65, by Santa Cruz Biotechnology (3 mentions)
β actin, by Santa Cruz Biotechnology (3 mentions)
β actin, by Cell Signaling Technology (3 mentions)
γ 32p atp, by PerkinElmer (2 mentions)
Mcl 1, by Santa Cruz Biotechnology (2 mentions)
Sirna transfection reagent, by Santa Cruz Biotechnology (2 mentions)
Traf2, by Santa Cruz Biotechnology (2 mentions)
Traf6, by Santa Cruz Biotechnology (2 mentions)
Polyclonal antibodies generated against dream, by Santa Cruz Biotechnology (2 mentions)
Nf κb2, by Santa Cruz Biotechnology (2 mentions)
Tak1 rabbit mab d94d7, by Cell Signaling Technology (2 mentions)
Phospho sapk jnk thr183 tyr185 rabbit mab 81e11, by Cell Signaling Technology (2 mentions)
Sapk jnk rabbit mab 56g8, by Cell Signaling Technology (2 mentions)
Ikkα pab, by Cell Signaling Technology (2 mentions)
Phospho ikkα β ser176 180 pab, by Cell Signaling Technology (2 mentions)
Ikkβ pab, by Cell Signaling Technology (2 mentions)
Rip2 pab, by Cell Signaling Technology (2 mentions)
P65 rela pab, by Merck Group (2 mentions)
Lipids dimethyldioctadecylammonium bromide and chlolesterol for liposome preparation, by Merck Group (2 mentions)
30 protocols using c rel
1
Immunoblotting Analysis of Cellular Proteins
2
Protein Fractionation and Immunoblotting
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Total protein was fractionated by 9% SDS–PAGE, transferred
onto nitrocellulose and then blocked blots with Tris-buffered saline and Tween
20 (0.1%) containing 5% BSA before incubation overnight
with primary antibodies (1:1,000) to S100A9 (ab73987 Abcam), PCNA (ab2426
Abcam), Cyclin D1 (ab16663 Abcam), p105/p50 (ab7971 Abcam), RelA (ab7970 Abcam),
c-Rel (sc71 Santa Cruz), CyP2E1 (ab28146 Abcam) or GAPDH (Abcam), total
p38α (9218 Cell Signaling), P-p38α (9216 Cell Signaling), HA
or Flag-HRP conjugate (Sigma). Next day, membranes were washed in T-TBS and then
incubated with horseradish peroxidase-conjugated mouse anti-rabbit IgG. Blots
were washed and antigen was detected using enhanced chemiluminescence (Amersham
Biosciences). Images have been cropped for presentation. Full-size images are
presented inSupplementary Fig.
10 .
onto nitrocellulose and then blocked blots with Tris-buffered saline and Tween
20 (0.1%) containing 5% BSA before incubation overnight
with primary antibodies (1:1,000) to S100A9 (ab73987 Abcam), PCNA (ab2426
Abcam), Cyclin D1 (ab16663 Abcam), p105/p50 (ab7971 Abcam), RelA (ab7970 Abcam),
c-Rel (sc71 Santa Cruz), CyP2E1 (ab28146 Abcam) or GAPDH (Abcam), total
p38α (9218 Cell Signaling), P-p38α (9216 Cell Signaling), HA
or Flag-HRP conjugate (Sigma). Next day, membranes were washed in T-TBS and then
incubated with horseradish peroxidase-conjugated mouse anti-rabbit IgG. Blots
were washed and antigen was detected using enhanced chemiluminescence (Amersham
Biosciences). Images have been cropped for presentation. Full-size images are
presented in
10
3
Comprehensive Antibody Panel for DNA Damage Response
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Antibodies used were c-Rel (sc-71 Santa Cruz), c-Myc (sc-42 Santa Cruz), ATR (sc1887 Santa Cruz), RelA (sc-372 Santa Cruz), RelB (4954 Cell Signaling), p105/p50 (ab7971 Abcam), p100/p52 (4882 Cell Signaling), β-Actin (A5441 Sigma), CHK1 (phospho S345) (2341 Cell Signaling), CHK1 (2360 Cell Signaling) RelA (phospho 536) (3031 Cell Signaling), active caspase 3 (9664 Cell Signaling), γH2AX (9718 Cell Signaling), RPA2/RPA32 Ser 33 (10148 Cell Signaling), RPA2/RPA32 (52488 Cell Signaling), ATRIP (11327-1-AP Proteintech), Rad17 (13358-1-AP Proteintech), ATR (13934 Cell Signaling), Karyopherin Beta (Santa Cruz sc-137016). Antibodies to the murine form of Claspin was generated by Moravian Biotechnologies. Anti-rabbit IgG (A6154 Sigma and 7074 Cell Signaling) and anti-mouse IgG (A9044 Sigma) HRP-linked secondary antibodies were used for western blot detection.
4
Measuring Sp1 and NF-κB DNA Binding
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Nuclear protein was extracted from lungs. Sp1 and NF-B DNA binding activities were measured as we previously described [23 (link), 33 (link)]. Sp1 (5'-ATTCGATCGGGGCGGGGCC AG-3’) and NF-κB (5’-AGTTGAGGGGACTTTCC CAGGC-3) consensus oligonucleotide probes were end-labeled with [γ-32P] ATP (PerkinElmer, Waltham, MA). Nuclear protein (10 μg) was incubated with 50,000 cpm 32P-labeled Sp1 or NF-κB probe for 30 minutes in binding buffer consisted of 10 mM Tris-Cl, pH 7.5, 1 mM MgCl2, 50 mM NaCl, 0.5 mM DTT, 0.5 mM EDTA, 4% glycerol and 1 μg of poly-(dI•dC). The specificity of Sp1 or NF-κB DNA binding was determined in competition reactions, in which a 50-fold molar excess unlabelled Sp1 or NF-κB, or an unrelated oligonucleotide was added to the binding reaction 10 minutes prior to the addition of radiolabeled probe. In the supershift assay, antibody against RelA/p65, RelB, c-Rel or p50, or a combination of p50+p65 (all from Santa Cruz Biotech, Dallas, TX) was added to the reaction mixture immediately after the addition of radiolabeled NF-κB probe. Reaction was subjected to non-denaturing 4% polyacrylamide gel electrophoresis. Gel was vacuum-dried and exposed to X-ray film.
5
Phosphorylation-Specific Antibody Immunoblotting
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Phosphospecific JNK, AKT, p38, p65, IκBα and ERK antibodies and TNF-α-neutralising antibody were all purchased from Cell Signaling Technology. NORE1A monoclonal antibody, CD40L, p21, p53, c-Rel, RNA polymerase II, rabbit isotype and histone H3 antibodies were all from Santa Cruz Biotechnology. CD40L–APC and isotype–APC conjugates were from eBioscience. For immunoblotting, 10–50 mg protein was separated by SDS-PAGE, transferred onto polyvinylidene difluoride membrane (for phosphoproteins) or Biotrace nitrocellulose membranes (Pall, Port Washington, NY, USA) (for nonphosphoproteins), and blocked with 10% of BSA (for phosphoproteins) or nonfat milk (for nonphosphoproteins) dissolved in PBS supplemented with 0.1% Tween 20 for 1 h. After three washes with PBS supplemented with 0.1% Tween 20, membranes were incubated overnight at 4 °C with primary antibody and for 1 h at room temperature with the appropriate secondary antibody followed by ECL (Amersham Biosciences, Piscataway, NJ, USA). Cytoplasmic and nuclear extracts from EJ cell were prepared by using the NE-PER nuclear and cytoplasmic extraction kit (Thermo Scientific, MA, USA, cat. 78835) according to the manufacturer's instructions.
6
Nuclear Protein Extraction and Western Blotting
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Cultured cells were harvested and whole cell lysates were prepared according to the method previously described [30 (link)]. Nuclear extracts were prepared using a Nuclear Extract kit (Cat. no. 40010, Active Motif, Carlsbad, CA) following the manufacturer’s instructions. Protein concentration was determined using the BCA Assay Reagent (Cat. no. 23228, Pierce, Rockford, IL). Western blotting was performed as previously described [30 (link)]. The following antibodies were used for immunodetection with appropriate dilutions: Mcl-1 (sc-819, 1:1000), p50 (sc-114, 1:1000), p52 (sc-298, 1:1000), p65 (sc-8008, 1:1000), c-Rel (sc-272, 1:1000), RelB (sc-226, 1:1000) and GAPDH (sc-47724, 1:2000) (all from Santa Cruz, CA); Histone H3 (#9715, 1:1000) were purchased from Cell Signaling Technology (Beverly, MA); β-actin (A5316, 1:5000) was purchased from Sigma (St. Louis, MO).
7
Antibodies and Reagents for NF-κB Signaling
8
Protein Extraction and Western Blot
9
Quantifying NF-κB and SP-1 DNA Binding
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MDA-MB-231 and MEF cells were harvested in their exponential growth phase with or without TNFα (5 ng/ml, R&D Systems, Minneapolis, MN, USA) treatment for 15 min and assayed for NF-κB and SP-1 (as a control) DNA-binding activity as described previously.39 (link) Antibodies for supershift assays were purchased from Santa Cruz (c-Rel, cat. no. sc-070) and Millipore (p65, cat. no. 06-418; p50, cat. no. 06-886).
10
Antigen-Specific Immune Modulation Assay
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Antibodies targeting IκBα (C-21, 1:1,000), p65 (C-20, 1:1,000), Lamin B (C-20, 1:1,000), ERK (K-23, 1:2,000), phospho-ERK (E-4, 1:1,000), JNK2 (C-17, 1:1,000), p38 (H-147, 1:1,000), p105/p50 (C-19, 1:1,000), TBK1 (108A429, 1:1,000), IRF3 (SC-9082, 1:1,000), RelB (C-19; 1:1,000), NIK (H248; 1:1,000), and c-Rel (sc-71, 1:1,000), as well as a control rabbit IgG (sc-2027), were obtained from Santa Cruz Biotechnology. Antibodies targeting phospho-IκBα (Ser32, 14D4, 1:1,000), phospho-JNK (Thr180/Tyr185, #9251, 1:1,000), phospho-p38 (Thr180/Tyr182, 3D7, 1:1,000), phospho-p105 (Ser933, 18E6, 1:1,000), phospho-TBK1 (Ser172, D52C2, 1:1,000), phospho-IRF3 (Ser396,4D4G, 1:1,000), Uhrf1 (D6G8E, 1:1,000), and p100/p52 (4882; 1:1,000) were purchased from Cell Signaling Technology. Anti-actin (C-4, 1:10,000) was from Sigma-Aldrich. Other fluorescence-labeled antibodies are listed in the flow cytometry and cell sorting sections.
dcas9-Tet1 and gRNA plasmids were provided by Li Shen (Zhejiang University, Hangzhou, China). pMD2. G and psPAX2 plasmids were provided by Yichuan Xiao (Chinese Academy of Sciences, Shanghai, China).
CpG (2216) was purchased from Sigma-Aldrich. R848 and pI:C were purchased from Amersham Biosciences. The inactivated virus vaccine VaxigripTetra (IIV4) was purchased from Sanofi Pasteur.
dcas9-Tet1 and gRNA plasmids were provided by Li Shen (Zhejiang University, Hangzhou, China). pMD2. G and psPAX2 plasmids were provided by Yichuan Xiao (Chinese Academy of Sciences, Shanghai, China).
CpG (2216) was purchased from Sigma-Aldrich. R848 and pI:C were purchased from Amersham Biosciences. The inactivated virus vaccine VaxigripTetra (IIV4) was purchased from Sanofi Pasteur.
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