Revision 1

#55764Store at -20C

1 Kit

(6 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
NF-κB p65 (D14E12) XP® Rabbit mAb 8242 20 µl 65 kDa Rabbit IgG
Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb 3033 20 µl 65 kDa Rabbit IgG
RelB (D7D7W) Rabbit mAb 10544 20 µl 70 kDa Rabbit IgG
c-Rel (D4Y6M) Rabbit mAb 12707 20 µl 68-78 kDa Rabbit IgG
NF-κB1 p105/p50 (D4P4D) Rabbit mAb 13586 20 µl 50 Active form. 120 Precursor kDa Rabbit IgG
NF-κB2 p100/p52 (D7A9K) Rabbit mAb 37359 20 µl 120, 52 kDa Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The NF-κB Family Antibody Sampler Kit II provides an economical means of detecting members of the NF-κB family. The kit includes enough antibody to perform two western blots with each primary antibody.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammatory and immune responses (1,2). There are five family members in mammals: RelA, c-Rel, RelB, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Both p105 and p100 are proteolytically processed by the proteasome to produce p50 and p52, respectively. Rel proteins bind p50 and p52 to form dimeric complexes that bind DNA and regulate transcription. In unstimulated cells, NF-κB is sequestered in the cytoplasm by IκB inhibitory proteins (3-5). NF-κB-activating agents can induce the phosphorylation of IκB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-κB to enter the nucleus where it regulates gene expression (6-8). NIK and IKKα (IKK1) regulate the phosphorylation and processing of NF-κB2 (p100) to produce p52, which translocates to the nucleus (9-11).

  1. Baeuerle, P.A. and Henkel, T. (1994) Annu Rev Immunol 12, 141-79.
  2. Baeuerle, P.A. and Baltimore, D. (1996) Cell 87, 13-20.
  3. Haskill, S. et al. (1991) Cell 65, 1281-9.
  4. Thompson, J.E. et al. (1995) Cell 80, 573-82.
  5. Whiteside, S.T. et al. (1997) EMBO J 16, 1413-26.
  6. Traenckner, E.B. et al. (1995) EMBO J 14, 2876-83.
  7. Scherer, D.C. et al. (1995) Proc Natl Acad Sci USA 92, 11259-63.
  8. Chen, Z.J. et al. (1996) Cell 84, 853-62.
  9. Senftleben, U. et al. (2001) Science 293, 1495-9.
  10. Coope, H.J. et al. (2002) EMBO J 21, 5375-85.
  11. Xiao, G. et al. (2001) Mol Cell 7, 401-9.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    XP is a registered trademark of Cell Signaling Technology, Inc.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    使用に関する制限

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    Orders: 877-616-CELL (2355)•[email protected]• Support: 877-678-TECH (8324)•[email protected]• Web: cellsignal.com
    For Research Use Only. Not for Use in Diagnostic Procedures.
    Revision 1
    #55764

    NF-κB Family Antibody Sampler Kit II

    NF-κB Family Antibody Sampler Kit II: Image 1 Expand Image
    Simple Western™ analysis of lysates (1.0 mg/mL) from HeLa cells treated with hTNF-α (20 ng/mL, 5 minutes) using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb #3033. The virtual lane view (left) shows a single target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    NF-κB Family Antibody Sampler Kit II: Image 2 Expand Image
    Immunoprecipitation of Phospho-NF-κB p65 (Ser536) from HeLa extracts treated with hTNF-α #8902 (20 ng/ml, 5 min). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb. Western blot analysis was performed using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.
    NF-κB Family Antibody Sampler Kit II: Image 3 Expand Image
    Simple Western™ analysis of lysates (1 mg/mL) from HeLa cells using NF-κB p65 (D14E12) XP® Rabbit mAb #8242. The virtual lane view (left) shows a single target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody. This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
    NF-κB Family Antibody Sampler Kit II: Image 4 Expand Image
    Western blot analysis of extracts from various cell lines using RelB (D7D7W) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). KARPAS cell line source: Dr. Abraham Karpas at the University of Cambridge.
    NF-κB Family Antibody Sampler Kit II: Image 5 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HDLM-2 cells and RelB (D7D7W) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Library was prepared using DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795. The figure shows binding across BIRC3/IAP2, a known target gene of RelB (see additional figure containing ChIP-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 6 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either RelB (C1E4) Rabbit mAb #4922 or RelB (D7D7W) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across BIRC3 gene.
    NF-κB Family Antibody Sampler Kit II: Image 7 Expand Image
    Western blot analysis of extracts from various cell lines and tissues using c-Rel (D4Y6M) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 8 Expand Image
    Western blot analysis of extracts from various cell lines and rat spleen using NF-κB1 p105/p50 (D4P4D) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 9 Expand Image
    CUT&RUN was performed with HeLa cells treated with human TNF-α #8902 (30 ng/ml, 1 hr) and NF-κB1 p105/p50 (D4P4D) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across NFKBIA, a known target gene of NFkB (see additional figure containing CUT&RUN-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 10 Expand Image
    Western blot analysis of extracts from HeLa and NIH/3T3 cells, untreated or TNF-α treated (#2169, 20 ng/ml for 5 minutes), using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb (upper) or NF-κB p65 Antibody #3034 (lower).
    NF-κB Family Antibody Sampler Kit II: Image 11 Expand Image
    Western blot anlaysis of extracts from various cell lines using NF-κB2 p100/p52 (D7A9K) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). KARPAS cell line source: Dr. Abraham Karpas at the University of Cambridge.
    NF-κB Family Antibody Sampler Kit II: Image 12 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either NF-κB2 p100/p52 (18D10) Rabbit mAb #3017, NF-κB2 p100/p52 (D7A9K) Rabbit mAb, or NF-κB2 p100/p52 (D9S3M) Rabbit mAb #52583, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across BIRC3 gene.
    NF-κB Family Antibody Sampler Kit II: Image 13 Expand Image
    After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
    NF-κB Family Antibody Sampler Kit II: Image 14 Expand Image
    Western blot analysis of extracts from various cell lines using NF-κB p65 (D14E12) XP® Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 15 Expand Image
    Immunoprecipitation of RelB from Raji cell extracts. Lane 1 represents 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 and lane 3 is RelB (D7D7W) Rabbit mAb. Western blot was performed using RelB (D7D7W) Rabbit mAb. A conformation specific secondary antibody was used to avoid cross reativity with IgG.
    NF-κB Family Antibody Sampler Kit II: Image 16 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HDLM-2 cells and RelB (D7D7W) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Library was prepared using DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 11 (upper), including BIRC3/IAP2 (lower), a known target gene of RelB (see additional figure containing ChIP-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 17 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either RelB (C1E4) Rabbit mAb #4922 or RelB (D7D7W) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 11 (upper), including BIRC3 gene (lower).
    NF-κB Family Antibody Sampler Kit II: Image 18 Expand Image
    Western blot analysis of extracts from Neuro-2a cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® c-Rel siRNA I (Mouse Specific) #13058 (+) or SignalSilence® c-Rel siRNA II (Mouse Specific) #13170 (+), using c-Rel (D4Y6M) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). The c-Rel (D4Y6M) Rabbit mAb confirms silencing of c-Rel expression, while the β-Actin (D6A8) Rabbit mAb is used as a loading control.
    NF-κB Family Antibody Sampler Kit II: Image 19 Expand Image
    Immunoprecipitation of NF-κB1 p105/p50 from Raji cell extracts using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or NF-κB1 (D4P4D) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot was performed using NF-κB1 p105/p50 (D4P4D) Rabbit mAb. Mouse Anti-rabbit IgG (Light-Chain Specific) (L57A3) mAb #3677 was used as a secondary antibody.
    NF-κB Family Antibody Sampler Kit II: Image 20 Expand Image
    CUT&RUN was performed with HeLa cells treated with human TNF-α #8902 (30 ng/ml, 1 hr) and NF-κB1 p105/p50 (D4P4D) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 14 (upper), including NFKBIA (lower), a known target gene of NFkB (see additional figure containing CUT&RUN-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 21 Expand Image
    Western blot analysis of extracts from THP-1 cells, differentiated with TPA (#9905, 80 nM for 24h) and treated with 1 μg/ml LPS for the indicated times, using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb (upper) and NF-κB p65 (C22B4) Rabbit mAb #4764 (lower).
    NF-κB Family Antibody Sampler Kit II: Image 22 Expand Image
    Immunoprecipitation of NF-κB2 p100/p52 from HDLM-2 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is NF-κB2 p100/p52 (D7A9K) Rabbit mAb. Western blot analysis was performed using NF-κB2 p100/p52 (D7A9K) Rabbit mAb. A conformation-specific secondary antibody was used to avoid cross-reactivity with IgG.
    NF-κB Family Antibody Sampler Kit II: Image 23 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either NF-κB2 p100/p52 (18D10) Rabbit mAb #3017, NF-κB2 p100/p52 (D7A9K) Rabbit mAb, or NF-κB2 p100/p52 (D9S3M) Rabbit mAb #52583, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 11 (upper), including BIRC3 gene (lower).
    NF-κB Family Antibody Sampler Kit II: Image 24 Expand Image
    Immunohistochemical analysis of paraffin-embedded human colon carcinoma using RelB (D7D7W) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 25 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HDLM-2 cells and either RelB (D7D7W) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human IAP2 promoter primers, SimpleChIP® Human IκBα Promoter Primers #5552, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 26 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either RelB (D7D7W) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552 and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 27 Expand Image
    Confocal immunofluorescent analysis of C2C12 cells, untreated (left) or treated with Mouse Tumor Necrosis Factor-α (mTNF-α) #5178 (20 ng/ml, 30 min; right), using NF-κB1 p105/p50 (D4P4D) Rabbit mAb (green). Actin filaments were labeled with DyLight™ 554 Phalloidin #13054 (red). Blue pseudocolor= DRAQ5® #4084 (fluorescent DNA dye).
    NF-κB Family Antibody Sampler Kit II: Image 28 Expand Image
    CUT&RUN was performed with HeLa cells treated with human TNF-α #8902 (30 ng/ml, 1 hr) and either NF-κB1 p105/p50 (D4P4D) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552 and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 29 Expand Image
    Confocal immunofluorescent analysis of HeLa cells, serum starved (left) or TNF-α treated (#8902 at 20 ng/ml for 20 min, right), using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor® phalloidin 555 (red).
    NF-κB Family Antibody Sampler Kit II: Image 30 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HDLM-2 cells and either NF-κB2 p100/p52 (D7A9K) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552, Human IAP2 Promoter Primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 31 Expand Image
    CUT&RUN was performed with HDLM-2 cells and either NF-κB2 p100/p52 (D7A9K) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552 and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 32 Expand Image
    Immunohistochemical analysis using NF-κB p65 (D14E12) XP® Rabbit mAb on SignalSlide® NF-κB p65 IHC Controls #12873 (paraffin-embedded HCT116 cells, untreated (left) or treated with hTNF-α #8902 (right)).
    NF-κB Family Antibody Sampler Kit II: Image 33 Expand Image
    Immunohistochemical analysis of paraffin-embedded human lung carcinoma using RelB (D7D7W) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 34 Expand Image
    Flow cytometric analysis of NIH/3T3 cells using NF-κB1 p105/p50 (D4P4D) Rabbit mAb (solid line) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed line). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
    NF-κB Family Antibody Sampler Kit II: Image 35 Expand Image
    Flow cytometric analysis of HeLa cells, untreated (blue) or treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 and Calyculin A #9902 (20 ng/ml and 100 nM, 15 min; green), using Phospho-NF-κB p65 (Ser536) (93H1) Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
    NF-κB Family Antibody Sampler Kit II: Image 36 Expand Image
    Immunohistochemical analysis of paraffin-embedded human chronic cholecystitis using NF-κB p65 (D14E12) XP® Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 37 Expand Image
    Immunohistochemical analysis of paraffin-embedded HDLM-2 (left) and LNCaP (right) cell pellets using RelB (D7D7W) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 38 Expand Image
    Confocal immunofluorescent analysis of HT-1080 cells, untreated (left) or treated with hTNF-α #8902 (20 ng/ml, 20 min) (right), using NF-κB p65 (D14E12) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
    NF-κB Family Antibody Sampler Kit II: Image 39 Expand Image
    Immunohistochemical analysis of paraffin-embedded human tonsil using RelB (D7D7W) Rabbit mAb.
    NF-κB Family Antibody Sampler Kit II: Image 40 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells treated with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (30 ng/ml, 1 hr) and either NF-κB1 p105/p50 (D4P4D) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human IκBα Promoter Primers #5552, human IL-8 promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 41 Expand Image
    Flow cytometric analysis of HeLa cells using NF-κB p65 (D14E12) XP® Rabbit mAb (blue) compared to concentration matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red).
    NF-κB Family Antibody Sampler Kit II: Image 42 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and NF-κB p65 (D14E12) XP® Rabbit mAb, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across IL-8, a known target gene of NFκB (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
    NF-κB Family Antibody Sampler Kit II: Image 43 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and NF-κB p65 (D14E12) XP® Rabbit mAb, using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across chromosome 4 (upper), including IL-8 (lower), a known target gene of NFκB (see additional figure containing ChIP-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 44 Expand Image
    Confocal immunofluorescent analysis of PANC-1 cells (left, positive) and Kelly cells (right, negative) using RelB (D7D7W) Rabbit mAb (green). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).
    NF-κB Family Antibody Sampler Kit II: Image 45 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and either NF-κB p65 (D14E12) XP® Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by Real-Time PCR using SimpleChIP® Human IκBα Promoter Primers #5552, human IL-8 promoter primers, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 46 Expand Image
    Flow cytometric analysis of LNCaP cells (blue) and HDLM-2 cells (green) using RelB (D7D7W) Rabbit mAb (solid lines) or a concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
    NF-κB Family Antibody Sampler Kit II: Image 47 Expand Image
    CUT&RUN was performed with HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and NF-κB p65 (D14E12) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across LAMC2, a known target gene of NF-κB p65 (see additional figure containing CUT&RUN-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 48 Expand Image
    CUT&RUN was performed with HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and NF-κB p65 (D14E12) XP® Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 1 (upper), including LAMC2 (lower), a known target gene of NF-κB p65 (see additional figure containing CUT&RUN-qPCR data).
    NF-κB Family Antibody Sampler Kit II: Image 49 Expand Image
    CUT&RUN was performed with HeLa cells treated with hTNF-α #8902 (30 ng/ml, 1 hr) and either NF-κB p65 (D14E12) XP® Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human LAMC2 upstream primers, and human ITM2A upstream primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
    NF-κB Family Antibody Sampler Kit II: Image 50 Expand Image
    Immunoprecipitation of NF-kB p65 from CHO cell extracts. Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is NF-κB p65 (D14E12) XP® Rabbit mAb, #8242. Western blot was performed using NF-κB p65 (L8F6) Mouse mAb, #6956.
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