Revision 2
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.
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UniProt ID:

#P10275

Entrez-Gene Id:

367

Product Includes Quantity Reactivity MW(kDa) Isotype
Androgen Receptor (D6F11) XP® Rabbit mAb 5153 100 µl H 110 Rabbit IgG
Phospho-Androgen Receptor (Ser258) (E4L1I) Rabbit mAb 16969 100 µl H 110 Rabbit IgG

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

Description

PhosphoPlus® Duets from Cell Signaling Technology (CST) provide a means to assess protein activation status. Each Duet contains an activation-state and total protein antibody to your target of interest. These antibodies have been selected from CST's product offering based upon superior performance in specified applications.

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 antibodies.

Background

Androgen receptor (AR), a zinc finger transcription factor belonging to the nuclear receptor superfamily, is activated by phosphorylation and dimerization upon ligand binding (1). This promotes nuclear localization and binding of AR to androgen response elements in androgen target genes. Research studies have shown that AR plays a crucial role in several stages of male development and the progression of prostate cancer (2,3).

Upon ligand binding, AR becomes phosphorylated on many residues, including Ser258 (4).

  1. Li, J. and Al-Azzawi, F. (2009) Maturitas 63, 142-8.
  2. Avila, D.M. et al. (2001) J. Steroid. Biochem. Mol. Biol. 76, 135-142.
  3. Montgomery, J.S. et al. (2001) J. Pathol. 195, 138-146.
  4. Gioeli, D. et al. (2002) J Biol Chem 277, 29304-14.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    PhosphoPlus is a registered 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

    法的な権限を与えられたCSTの担当者が署名した書面によって別途明示的に合意された場合を除き、 CST、その関連会社または代理店が提供する製品には以下の条件が適用されます。お客様が定める条件でここに定められた条件に含まれるものを超えるもの、 または、ここに定められた条件と異なるものは、法的な権限を与えられたCSTの担当者が別途書面にて受諾した場合を除き、拒絶され、 いかなる効力も効果も有しません。

    研究専用 (For Research Use Only) またはこれに類似する表示がされた製品は、 いかなる目的についても FDA または外国もしくは国内のその他の規制機関により承認、認可または許可を受けていません。 お客様は製品を診断もしくは治療目的で使用してはならず、また、製品に表示された内容に違反する方法で使用してはなりません。 CST が販売または使用許諾する製品は、エンドユーザーであるお客様に対し、使途を研究および開発のみに限定して提供されるものです。 診断、予防もしくは治療目的で製品を使用することまたは製品を再販売 (単独であるか他の製品等の一部であるかを問いません) もしくはその他の商業的利用の目的で購入することについては、CST から別途許諾を得る必要があります。 お客様は以下の事項を遵守しなければなりません。(a) CST の製品 (単独であるか他の資材と一緒であるかを問いません) を販売、使用許諾、貸与、寄付もしくはその他の態様で第三者に譲渡したり使用させたりしてはなりません。また、商用の製品を製造するために CST の製品を使用してはなりません。(b) 複製、改変、リバースエンジニアリング、逆コンパイル、 分解または他の方法により製品の構造または技術を解明しようとしてはなりません。また、 CST の製品またはサービスと競合する製品またはサービスを開発する目的で CST の製品を使用してはなりません。(c) CST の製品の商標、商号、ロゴ、特許または著作権に関する通知または表示を除去したり改変したりしてはなりません。(d) CST の製品をCST 製品販売条件(CST’s Product Terms of Sale) および該当する書面のみに従って使用しなければなりません。(e) CST の製品に関連してお客様が使用する第三者の製品またはサービスに関する使用許諾条件、 サービス提供条件またはこれに類する合意事項を遵守しなければなりません。

    Revision 2
    #25015

    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet

    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 1 Expand Image
    Western blot analysis of extracts from LNCaP (AR+), VCaP (AR & AR-V7+), 22Rv1 (AR & AR-V7+), and PC-3 (AR-) cells using Androgen Receptor (D6F11) XP® Rabbit mAb (upper), and β-Actin (13E5) Rabbit mAb (lower).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 2 Expand Image
    Simple Western™ analysis of lysates (0.1 mg/mL) from LNCaP cells using Androgen Receptor (D6F11) XP® Rabbit mAb #5153. The virtual lane view (left) shows a single target band (as indicated) at 1:50 and 1:250 dilutions of primary antibody. The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:50 (blue line) and 1:250 (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.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 3 Expand Image
    Western blot analysis of extracts from LNCaP cells grown in phenol red free media and 5% charcoal stripped FBS for 3 d then either untreated (-), dihydrotestosterone (DHT)-treated (+), or DHT-treated and subsequently treated with λ phosphatase and calf intestinal phosphatase (CIP) (+), using Phospho-Androgen Receptor (Ser258) (E4L1I) Rabbit mAb (upper) or Androgen Receptor (D6F11) XP® Rabbit mAb #5153 (lower).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 4 Expand Image
    Western blot analysis of extracts from LNCaP (AR+), MCF7 (AR+), PC-3 (AR-), and DU 145 (AR-) cells using Androgen Receptor (D6F11) XP® Rabbit mAb (upper) and β-Actin Antibody #4967 (lower).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 5 Expand Image
    CUT&RUN was performed with LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and Androgen Receptor (D6F11) 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 KLK2, a known target gene of Androgen Receptor (see additional figure containing CUT&RUN-qPCR data).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 6 Expand Image
    Immunoprecipitation of phospho-androgen receptor (Ser258) protein from LNCaP cell extracts. Cells were grown in phenol red free media and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT) (5 nM, 6 hr). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-Androgen Receptor (Ser258) (E4L1I) Rabbit mAb. Western blot analysis was performed using Phospho-Androgen Receptor (Ser258) (E4L1I) Rabbit mAb. Anti-rabbit IgG, HRP-linked Antibody #7074 was used as a secondary antibody.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 7 Expand Image
    Immunohistochemical analysis of paraffin-embedded LNCaP (AR+, left) and DU145 (AR-, right) using Androgen Receptor (D6F11) XP® Rabbit mAb.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 8 Expand Image
    CUT&RUN was performed with LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and Androgen Receptor (D6F11) 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 19 (upper), including KLK2 (lower), a known target gene of Androgen Receptor (see additional figure containing CUT&RUN-qPCR data).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 9 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT) (5 nM, 6 hr) and either Phospho-Androgen Receptor (Ser258) (E4L1I) 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 KLK3 Promoter Primers #32784 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.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 10 Expand Image
    Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using Androgen Receptor (D6F11) XP® Rabbit mAb.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 11 Expand Image
    CUT&RUN was performed with LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and either Androgen Receptor (D6F11) 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 SimpleChIP® Human KLK2 Intron 1 Primers #62086 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.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 12 Expand Image
    Confocal immunofluorescent analysis of LNCaP (positive, left) and DU145 (negative, right) cells using Androgen Receptor (D6F11) XP® Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 13 Expand Image
    Flow cytometric analysis of DU-145 cells (blue) and LNCaP cells (green) using Androgen Receptor (D6F11) XP® 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.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 14 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and Androgen Receptor (D6F11) XP® Rabbit mAb, using SimpleChIP® Plus 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 KLK2, a known target gene of Androgen Receptor (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 15 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and Androgen Receptor (D6F11) XP® Rabbit mAb, using SimpleChIP® Plus 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 19 (upper), including KLK2 (lower), a known target gene of Androgen Receptor (see additional figure containing ChIP-qPCR data).
    PhosphoPlus® Androgen Receptor (Ser258) Antibody Duet: Image 16 Expand Image
    Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells grown in phenol red free medium and 5% charcoal stripped FBS for 3 d then treated with dihydrotestosterone (DHT, 10 nM) for 4 hours and either Androgen Receptor (D6F11) XP® 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 KLK2 Intron 1 Primers #62086, SimpleChIP® Human KLK3 Promoter Primers #32784, 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.