Revision 1

#83700Store at -20C

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.
Applications:

WB, IF-IC, ChIP, ChIP-seq

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

30

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q99801

Entrez-Gene Id:

4824

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 5 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:100
Chromatin IP 1:100
Chromatin IP-seq 1:100

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.

Specificity / Sensitivity

NKX3.1 (D2Y1A) XP® Rabbit mAb recognizes endogenous levels of total NKX3.1 protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein specific to human NKX3.1 protein.

Background

NKX3.1 is a homeobox transcription factor that in mammals plays a defining role in embryonic prostate morphogenesis. The expression of mammalian NKX3.1 is androgen-dependent, restricted primarily to developing and mature prostate epithelium, and is frequently reduced or lost in prostate cancer (1-3). The human NKX3.1 gene is located on chromsome 8p21.2, within a region that shows loss of heterozygosity (LOH) in >50% of prostate cancer cases (2). Allelic loss at the NKX3.1 locus is also common in high grade Prostate Intraepithelial Neoplasia (PIN), thought to be a putative precursor lesion to invasive prostate adenocarcinomas, suggesting that LOH at the NKX3.1 locus is a critical early step in prostate cancer development (4). Notably, the remaining NKX3.1 allele is intact in the majority of LOH cases, leading to the suggestion that NKX3.1 functions as a haploinsufficient tumor suppressor (4-6). Due to its highly restricted expression in prostate epithelial cells, NKX3.1 has been suggested as a diagnostic marker of prostate carcinoma (7), and may have additional utility as a biomarker of metastatic lesions originating in the prostate (8).

  1. Bhatia-Gaur, R. et al. (1999) Genes Dev 13, 966-77.
  2. He, W.W. et al. (1997) Genomics 43, 69-77.
  3. Bowen, C. et al. (2000) Cancer Res 60, 6111-5.
  4. Magee, J.A. et al. (2003) Cancer Cell 3, 273-83.
  5. Voeller, H.J. et al. (1997) Cancer Res 57, 4455-9.
  6. Bethel, C.R. et al. (2006) Cancer Res 66, 10683-90.
  7. Epstein, J.I. et al. (2014) Am J Surg Pathol 38, e6-e19.
  8. Conner, J.R. and Hornick, J.L. (2015) Adv Anat Pathol 22, 149-67.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IF-IC: Immunofluorescence (Immunocytochemistry) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
SimpleChIP 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.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 1
#83700

NKX3.1 (D2Y1A) XP® Rabbit mAb

Western Blotting Image 1: NKX3.1 (D2Y1A) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from human prostate and various cell lines using NKX3.1 (D2Y1A) XP® Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower). DND-41 cell are negative for NKX3.1 expression, confirming specificity of the antibody.
Immunofluorescence Image 1: NKX3.1 (D2Y1A) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of LNCaP (left; positive) and HeLa (right; negative) cells using NKX3.1 (D2Y1A) XP® Rabbit mAb (green). Actin filaments were labeled with DyLight 554 Phalloidin #13054 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Chromatin Immunoprecipitation Image 1: NKX3.1 (D2Y1A) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells and NKX3.1 (D2Y1A) XP® Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across CYP2B7P, a known target gene of NKX3.1 (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
Chromatin Immunoprecipitation Image 2: NKX3.1 (D2Y1A) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells and NKX3.1 (D2Y1A) XP® Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795. The figure shows binding across chromosome 19 (upper), including CYP2B7P (lower), a known target gene of NKX3.1 (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 3: NKX3.1 (D2Y1A) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells and either NKX3.1 (D2Y1A) 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 CYP2B7P Pseudogene Promoter Primers #84846, human MALSU1 downstream 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.