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89985
Huntingtin Interaction Antibody Sampler Kit
Primary Antibodies

Huntingtin Interaction Antibody Sampler Kit #89985

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Western blot analysis of extracts from various cell lines and rat spleen using NF-κB1 p105/p50 (D4P4D) Rabbit mAb #13586.

Western blot analysis of extracts from NIH/3T3 and 3T3-L1 cells (differentiated 6 days) using PPARγ (C26H12) Rabbit mAb #2435.

Western blot analysis of extracts from various cell lines using ACF1 Antibody #6255.

Western blot analysis of recombinant GST-SUMO-1 protein (38 kDa) and extracts from Jurkat cells using SUMO-1 (C9H1) Rabbit mAb #4940.

Western blot analysis of extracts from 293 and COS cells, using p53 (7F5) Rabbit mAb #2527.

Western blot analysis of extracts from various cell lines using CtBP1 (D2D6) Rabbit mAb #8684.

Chromatin immunoprecipitations were performed with cross-linked chromatin from HDLM-2 cells and either PPARγ (C26H12) 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 FOXN3 Intron 3 Primers #95568, human STON2 intron 4 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.

Confocal immunofluorescent analysis of 293T cells using CtBP1 (D2D6) Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red).

Chromatin immunoprecipitations were performed with cross-linked chromatin from HCT116 cells treated with UV (100 J/m2 followed by a 3 hour recovery) and either p53 (7F5) 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 CDKN1A Promoter Primers #6449, human MDM2 intron 2 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.

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using SUMO-1 (C9H1) Rabbit mAb in the presence of control peptide (left) or antigen specific peptide (right).

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.

Western blot analysis of extracts from various cell lines using ACF1 Antibody.

Confocal immunofluorescent analysis of 3T3-L1 cells using PPARγ (C26H12A8) Rabbit mAb (red) showing nuclear localization in differentiated cells. Lipid droplets have been labeled with BODIPY 493/503 (green). Blue pseudocolor = DRAQ5™ (fluorescent DNA dye).

Immunohistochemical analysis of paraffin-embedded human colon carcinoma using CtBP1 (D2D6) Rabbit mAb in the presence of control peptide (left) or antigen-specific peptide (right).

Flow cytometric analysis of HT-29 cells using p53 (7F5) Rabbit mAb (blue) compared to a nonspecific negative control antibody (red).

Western blot analysis of recombinant GST-SUMO-1 protein (38 kDa) and extracts from Jurkat cells using SUMO-1 (C9H1) Rabbit mAb.

Flow cytometric analysis of C2C12 cells using NF-κB1 p105/p50 (D4P4D) Rabbit mAb (blue) compared to Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody.

Immunohistochemical analysis of 3T3-L1 cells, undifferentiated (left) or differentiated (right) , using PPARγ (C26H12) Rabbit mAb.

Immunohistochemical analysis of paraffin-embedded human lung squamous cell carcinoma using CtBP1 (D2D6) Rabbit mAb.

Confocal Immunofluorescent analysis of HT-29 cells using p53 (7F5) Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red).

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).

Immunohistochemical analysis of paraffin-embedded mouse brown fat using PPARγ (C26H12) Rabbit mAb.

Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using CtBP1 (D2D6) Rabbit mAb.

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, using p53 (7F5) Rabbit mAb.

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.

Western blot analysis of extracts from NIH/3T3 and 3T3-L1 cells (differentiated 6 days) using PPARγ (C26H12) Rabbit mAb.

Western blot analysis of extracts from various cell lines using CtBP1 (D2D6) Rabbit mAb.

Immunohistochemical analysis of paraffin-embedded human colon carcinoma, using p53 (7F5) Rabbit mAb.

Western blot analysis of extracts from various cell lines and rat spleen using NF-κB1 p105/p50 (D4P4D) Rabbit mAb.

Immunohistochemical analysis of paraffin-embedded HT-29 (left) and SaOs-2 (right) cells, using p53 (7F5) Rabbit mAb. Note the lack of staining in p53-negative SaOs-2 cells.

Western blot analysis of extracts from 293 and COS cells, using p53 (7F5) Rabbit mAb.

To Purchase # 89985T
Product # Size Price
89985T
1 Kit  (6 x 20 µl) $ 432

Product Includes Quantity Applications Reactivity MW(kDa) Isotype
PPARγ (C26H12) Rabbit mAb 2435 20 µl
  • WB
  • IHC
  • IF
  • ChIP
H M 53, 57 Rabbit IgG
CtBP1 (D2D6) Rabbit mAb 8684 20 µl
  • WB
  • IP
  • IHC
  • IF
H M Mk 47 Rabbit IgG
p53 (7F5) Rabbit mAb 2527 20 µl
  • WB
  • IHC
  • IF
  • F
  • ChIP
H Mk 53 Rabbit IgG
SUMO-1 (C9H1) Rabbit mAb 4940 20 µl
  • WB
  • IP
  • IHC
H M R Mk Rabbit IgG
NF-κB1 p105/p50 (D4P4D) Rabbit mAb 13586 20 µl
  • WB
  • IP
  • IF
  • F
  • ChIP
H M R 50 Active form. 120 Precursor Rabbit IgG
ACF1 Antibody 6255 20 µl
  • WB
H Mk 203 Rabbit 
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

Product Description

The Huntingtin Interaction Antibody Sampler kit provides an economical means of detecting transcription-related proteins that interact with Huntingtin (Htt). This kit contains enough antibody to perform two western blot experiments per primary antibody.

Specificity / Sensitivity

Unless otherwise indicated, each antibody will recognize endogenous levels of total target protein. SUMO-1 (C9H1) Rabbit mAb detects recombinant SUMO-1 and endogenous levels of sumoylated proteins (e.g. SUMO-1-RanGAP at 90 kD). SUMO-1 (C9H1) Rabbit mAb does not detect recombinant SUMO-2 or SUMO-3. ACF1 Antibody recognizes endogenous levels of total ACF1 protein (isoforms 1 and 2).

Source / Purification

Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the residues surrounding Asp69 of human PPARγ, amino terminus of the human CtBP1 protein, full-length human p53 fusion protein, amino terminus of human SUMO-1, or residues surrounding Ile415 of mouse NF-kB1 P105/p50 protein. Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponds to the residues surrounding Met864 of human ACF1 protein. Antibodies are purified by protein A and peptide affinity chromatography

Background

Peroxisome proliferator-activated receptor gamma (PPARG) is a member of the ligand-activated nuclear receptor superfamily and functions as a transcriptional activator (1). Besides its role in mediating adipogenesis and lipid metabolism (2), PPAR gamma also modulates insulin sensitivity, cell proliferation and inflammation (3). CtBP1 is able to regulate gene activity through its intrinsic dehydrogenase activity (4,5) and by interacting with Polycomb Group (PcG) proteins during development (6). Along with its homologue, CtBP2, it acts as a transcriptional corepressor of zinc-finger homeodomain factor deltaEF1 to regulate a wide range of cellular processes through transrepression mechanisms (7). The p53 tumor suppressor protein plays a major role in cellular response to DNA damage and other genomic aberrations. Activation of p53 can lead to either cell cycle arrest and DNA repair or apoptosis (8). DNA damage induces phosphorylation of p53 at Ser15 and Ser20 and leads to a reduced interaction between p53 and its negative regulator, the oncoprotein MDM2 (9). MDM2 inhibits p53 accumulation by targeting it for ubiquitination and proteasomal degradation (10,11). Phosphorylation impairs the ability of MDM2 to bind p53, promoting both the accumulation and activation of p53 in response to DNA damage (9,12). Acetylation appears to play a positive role in the accumulation of p53 protein in stress response (13). Deacetylation of p53 occurs through interaction with the SIRT1 protein, a deacetylase that may be involved in cellular aging and the DNA damage response (14). Small ubiquitin-related modifier 1, 2 and 3 (SUMO-1, -2 and -3) are members of the ubiquitin-like protein family (15). The covalent attachment of the SUMO-1, -2 or -3 (SUMOylation) to target proteins is analogous to ubiquitination. Ubiquitin and the individual SUMO family members are all targeted to different proteins with diverse biological functions. Ubiquitin predominantly regulates degradation of its target (1). In contrast, SUMO-1 is conjugated to RanGAP, PML, p53 and IkB-alpha to regulate nuclear trafficking, formation of subnuclear structures, regulation of transcriptional activity and protein stability (16-20). Transcription factors of the nuclear factor kappaB (NF-kB)/Rel family play a pivotal role in inflammatory and immune responses (21, 22). In unstimulated cells, NF-kB is sequestered in the cytoplasm by IkB inhibitory proteins (23-25). NF-kB-activating agents can induce the phosphorylation of IkB proteins, targeting them for rapid degradation through the ubiquitin-proteasome pathway and releasing NF-kB to enter the nucleus where it regulates gene expression (26-28). ACF1 (BAZ1A) has distinct roles in development (29), regulation of chromatin structure (30), and DNA damage response (31, 32). Different developmental stages dictate the expression of ACF1 in Drosophila, and alterations in ACF1 expression during Drosophila development leads to deviation from normal chromatin organization (29).

  1. Chehab NH et al. (1999) Proc Natl Acad Sci U S A 96, 13777–82
  2. Levine, A.J. (1997) Cell 88, 323-31.
  3. Baeuerle, P.A. and Henkel, T. (1994) Annu Rev Immunol 12, 141-79.
  4. Ho, L. and Crabtree, G.R. (2010) Nature 463, 474-84.
  5. Balasubramanian, P. et al. (2003) FEBS Lett 537, 157-60.
  6. Schwartz, D.C. and Hochstrasser, M. (2003) Trends Biochem Sci 28, 321-8.
  7. Baeuerle, P.A. and Baltimore, D. (1996) Cell 87, 13-20.
  8. Chioda, M. et al. (2010) Development 137, 3513-22.
  9. Haskill, S. et al. (1991) Cell 65, 1281-9.
  10. Shieh, S.Y. et al. (1997) Cell 91, 325-34.
  11. Matunis, M.J. et al. (1996) J Cell Biol 135, 1457-70.
  12. Thompson, J.E. et al. (1995) Cell 80, 573-82.
  13. Sánchez-Molina, S. et al. (2011) Nucleic Acids Res 39, 8445-56.
  14. Furusawa, T. et al. (1999) Mol Cell Biol 19, 8581-90.
  15. Tibbetts, R.S. et al. (1999) Genes Dev 13, 152-7.
  16. Whiteside, S.T. et al. (1997) EMBO J 16, 1413-26.
  17. Lan, L. et al. (2010) Mol Cell 40, 976-87.
  18. Honda, R. et al. (1997) FEBS Lett 420, 25-7.
  19. Traenckner, E.B. et al. (1995) EMBO J 14, 2876-83.
  20. Tontonoz, P. et al. (1995) Curr Opin Genet Dev 5, 571-6.
  21. Sewalt, R.G. et al. (1999) Mol Cell Biol 19, 777-87.
  22. Scherer, D.C. et al. (1995) Proc Natl Acad Sci USA 92, 11259-63.
  23. Chen, Z.J. et al. (1996) Cell 84, 853-62.
  24. Rosen, E.D. et al. (1999) Mol Cell 4, 611-7.
  25. Murphy, G.J. and Holder, J.C. (2000) Trends Pharmacol Sci 21, 469-74.
  26. Kumar, V. et al. (2002) Mol Cell 10, 857-69.
  27. Duprez, E. et al. (1999) J Cell Sci 112 ( Pt 3), 381-93.
  28. Gostissa, M. et al. (1999) EMBO J 18, 6462-71.
  29. Rodriguez, M.S. et al. (1999) EMBO J 18, 6455-61.
  30. Desterro, J.M. et al. (1998) Mol Cell 2, 233-9.
  31. Ito, A. et al. (2001) EMBO J 20, 1331-40.
  32. Solomon, J.M. et al. (2006) Mol Cell Biol 26, 28-38.

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