Revision 3
Cell Signaling Technology

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Support: 877-678-TECH (8324)

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3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
RARα (E6Z6K) Rabbit mAb 62294 20 µl 60 kDa Rabbit IgG
RXRα (D6H10) Rabbit mAb 3085 20 µl 53 kDa Rabbit IgG
RXRβ Antibody 8715 20 µl 70-72 kDa Rabbit 
RARγ1 (D3A4) XP® Rabbit mAb 8965 20 µl 58 kDa Rabbit IgG
RXRγ Antibody 5629 20 µl 55 kDa Rabbit 
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 Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit provides an economical means to investigate the expression of various subtypes of retinoic acid and retinoid X receptors. The kit contains enough primary antibody to perform two western blot experiments per primary.

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

Nuclear retinoic acid (RA) receptors (RARs) consist of three subtypes encoded by separate genes: α (NR1B1), β (NR1B2), and γ (NR1B3). For each subtype, there are at least two isoforms, which are generated by differential promoter usage and alternative splicing and differ only in their N-terminal regions. Retinoids, which are metabolites of vitamin A, serve as ligands for RARs (1). RARs function as ligand-dependent transcriptional regulators and are found to be heterodimerized with retinoid X receptors (RXRs). These transcriptionally active dimers regulate the expression of genes involved in cellular differentiation, proliferation, and apoptosis (2,3). Consequently, RARs play critical roles in a variety of biological processes, including development, reproduction, immunity, and organogenesis (4-6). RAR mutations, fusion proteins, altered expression levels, or aberrant post-translational modifications result in multiple diseases due to altered RAR function and disruption of homeostasis.

In contrast to the ubiquitously expressed RARα subtype, RARγ displays a complex tissue-specific expression pattern (7). The hematopoietic system expresses significant levels of RARγ, and a recent study identified a role for RARγ in hematopoietic stem cell maintenance (8). RARγ is the predominant subtype in human and mouse epidermis, representing 90% of the RARs in this tissue (9-11). Given the high level of RARγ expression in the skin, it has been suggested that this nuclear receptor participates in a transcriptional program that governs maintenance and differentiation of normal epidermis and skin appendages. The transcriptional activity of RARγ is under stringent control, in part, through retinoic acid-induced phosphorylation and proteasomal degradation (12).

The human retinoid X receptors (RXRs) are encoded by three distinct genes (RXRα, RXRβ, and RXRγ) and bind selectively and with high affinity to the vitamin A derivative, 9-cis-retinoic acid. RXRs are type-II nuclear hormone receptors that are largely localized to the nuclear compartment independent of ligand binding. Nuclear RXRs form heterodimers with nuclear hormone receptor subfamily 1 proteins, including thyroid hormone receptor, retinoic acid receptors, vitamin D receptor, peroxisome proliferator-activated receptors, liver X receptors, and farnesoid X receptor (13). Since RXRs heterodimerize with multiple nuclear hormone receptors, they play a central role in transcriptional control of numerous hormonal signaling pathways by binding to cis-acting response elements in the promoter/enhancer region of target genes (14).

  1. Rochette-Egly, C. and Germain, P. (2009) Nucl Recept Signal 7, e005.
  2. Delacroix, L. et al. (2010) Mol Cell Biol 30, 231-44.
  3. Eifert, C. et al. (2006) Mol Reprod Dev 73, 796-824.
  4. Mark, M. et al. (2006) Annu Rev Pharmacol Toxicol 46, 451-80.
  5. Niederreither, K. and Dollé, P. (2008) Nat Rev Genet 9, 541-53.
  6. Mark, M. et al. (2009) Nucl Recept Signal 7, e002.
  7. Dollé, P. (2009) Nucl Recept Signal 7, e006.
  8. Purton, L.E. et al. (2006) J Exp Med 203, 1283-93.
  9. Fisher, G.J. et al. (1994) J Biol Chem 269, 20629-35.
  10. Zelent, A. et al. (1989) Nature 339, 714-7.
  11. Elder, J.T. et al. (1991) J Invest Dermatol 96, 425-33.
  12. Giannì, M. et al. (2002) EMBO J 21, 3760-9.
  13. Gronemeyer, H. et al. (2004) Nat Rev Drug Discov 3, 950-64.
  14. Mangelsdorf, D.J. et al. (1992) Genes Dev 6, 329-44.

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.

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    Revision 3
    #8589

    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit

    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 1 Expand Image
    Western blot analysis of extracts from various cell lines using RXRα (D6H10) Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 2 Expand Image
    Western blot analysis of extracts from 293T cells, either mock transfected or transfected with human RXRα, RXRβ, or RXRγ DYKDDDK-tagged constructs, using RXRγ Antibody (upper) and DYKDDDK Tag Antibody (Binds to same epitope as Sigma's Anti-FLAG® M2 Antibody) #2368 (lower).
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 3 Expand Image
    Western blot analysis of extracts from various cell lines using RARα (E6Z6K) Rabbit mAb. The NB-4 cell line contains the PML-RARα fusion protein.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 4 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.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 5 Expand Image
    Western blot analysis of extracts from various cell lines using RXRβ Antibody.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 6 Expand Image
    Western blot analysis of extracts from various cell lines using RARγ1 (D3A4) XP® Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 7 Expand Image
    Western blot analysis of extracts from 293T cells, either mock transfected (-) or transfected with Myc/DDK-tagged cDNA expression constructs encoding full-length human RXRα (hRXRα; +), RXRβ (hRXRβ; +), or RXRγ (hRXRγ; +), using RXRα (D6H10) Rabbit mAb (upper) and DYKDDDDK Tag Antibody (Binds to same epitope as Sigma's Anti-FLAG® M2 Antibody) #2368 (lower).
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 8 Expand Image
    Western blot analysis of extracts from various tissues using RXRγ Antibody.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 9 Expand Image
    Immunoprecipitation of RARα and PML-RARα from NB-4 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is RARα (E6Z6K) Rabbit mAb. Western blot analysis was performed with RARα (E6Z6K) Rabbit mAb. The NB-4 cell line contains the PML-RARα fusion. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127 was used as a secondary antibody.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 10 Expand Image
    Western blot analysis of extracts from 293T cells, either mock-transfected or transfected with a Myc/DDK-tagged cDNA expression construct encoding full-length human RXRα, RXRβ, and RXRγ, using RXRβ Antibody (upper) or DYKDDDDK Tag Antibody (Binds to same epitope as Sigma's Anti-FLAG® M2 Antibody) #2368 (lower).
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 11 Expand Image
    Western blot analysis of extracts from 293T cells, either mock transfected (-) or transfected with a Myc/DDK-tagged cDNA expression construct encoding full-length human RARγ1 (hRARγ1-Myc/DDK, +), using RARγ1 (D3A4) XP® Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 12 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from LS-180 cells treated with Vitamin D (10nM) for 3 hours and either RXRα (D6H10) 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 c-Fos Upstream Primers #25661, human UCA1 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.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 13 Expand Image
    Immunohistochemical analysis of paraffin-embedded cell pellets, HaCaT (positive, left) and Hep3B (negative, right), using RARγ1 (D3A4) XP® Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 14 Expand Image
    Immunohistochemical analysis of paraffin-embedded human lung carcinoma using RARγ1 (D3A4) XP® Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 15 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from NB-4 cells and either RARα (E6Z6K) Rabbit mAb #62294 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 RUNX1 Exon 2 Primers #67254, human GFI promoter primers, human GFI exon 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.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 16 Expand Image
    Immunohistochemical analysis of paraffin-embedded human skin using RARγ1 (D3A4) XP® Rabbit mAb.
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 17 Expand Image
    Confocal immunofluorescent analysis of HaCaT cells (positive, left) and Hep3B cells (negative, right) using RARγ1 (D3A4) XP® Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red).
    Retinoic Acid and Retinoid X Receptors Antibody Sampler Kit: Image 18 Expand Image
    Flow cytometric analysis of T-47D cells using RARγ1 (D3A4) XP® Rabbit mAb (blue) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.