Order 3, Get Your 4th Free!* | Learn More >>
72209
Brachyury (D2Z3J) Rabbit mAb (PE Conjugate)
Antibody Conjugates
Monoclonal Antibody

Brachyury (D2Z3J) Rabbit mAb (PE Conjugate) #72209

Reviews ()
Citations (0)
Filter:
  1. F
Flow Cytometry Image 1: Brachyury (D2Z3J) Rabbit mAb (PE Conjugate)

Flow cytometric analysis of HeLa cells (blue) and MUG-Chor1 cells (green) using Brachyury (D2Z3J) Rabbit mAb (PE Conjugate) (solid line) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control (PE Conjugate) #5742 (dashed line).

To Purchase # 72209S
Product # Size Price
72209S
100 µl  (50 tests) N/A

Supporting Data

REACTIVITY H
SENSITIVITY Endogenous
MW (kDa)
Source/Isotype Rabbit IgG

Application Key:

  • W-Western
  • IP-Immunoprecipitation
  • IHC-Immunohistochemistry
  • ChIP-Chromatin Immunoprecipitation
  • IF-Immunofluorescence
  • F-Flow Cytometry
  • E-P-ELISA-Peptide

Species Cross-Reactivity Key:

  • H-Human
  • M-Mouse
  • R-Rat
  • Hm-Hamster
  • Mk-Monkey
  • 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
  • All-All Species Expected

Product Description

This Cell Signaling Technology antibody is conjugated to phycoerythrin (PE) and tested in-house for direct flow cytometric analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated Brachyury (D2Z3J) Rabbit mAb #81694.

Product Usage Information

Application Dilution
Flow Cytometry 1:50

Storage

Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.

Protocol

PRINT

View >Collapse >

Flow Cytometry Protocol for the Detection of Transcription Factors

A. Solutions and Reagents

NOTE: Prepare solutions with RODI (reverse osmosis deionized) or equivalent grade water.

  1. 20X Phosphate Buffered Saline (PBS): (#9808) To prepare 1 L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix.
  2. Formaldehyde (methanol free).
  3. 2% Formaldehyde (stock formaldehyde diluted in PBS; make fresh on day of experiment).
  4. Triton™ X-100.
  5. Incubation Buffer: Dissolve 0.5 g bovine serum albumin (BSA) (#9998) in 100 ml 1X PBS. Store at 4°C.

B. Fixation

NOTE: Immunostaining of surface antigens should be performed on live cells prior to fixation/permeabilization, as per manufacturer’s requirements.

  1. Adjust cell suspension of freshly isolated cells to 1–2 x 106 cells in 100 μl Incubation Buffer per assay tube.
  2. Add antibodies against surface antigens to the assay tubes as per manufacturers’ recommended volume or concentration and incubate for 30 min on ice.
  3. Add 2 ml of Incubation Buffer and wash by centrifugation.
  4. Aspirate supernatant and resuspend in 500 μl of 2% formaldehyde.
  5. Fix for 15 min at room temperature.
  6. Wash 2X by centrifugation in Incubation Buffer.

C. Permeabilization

  1. Add 1ml of 0.1% Triton™ X-100 (v/v in PBS) to the cell pellet.
  2. Resuspend and let stand for 30 min at room temperature.
  3. Wash 2X by centrifugation in Incubation Buffer.

D. Immunostaining

  1. Resuspend cell pellets in 100 μl of antibody working solution, diluted according to individual antibody datasheet or product webpage in Incubation Buffer.
  2. Incubate for 1 hr at room temperature.
  3. Wash 2X by centrifugation in Incubation Buffer.
  4. If using a fluorochrome-conjugated primary antibody, resuspend cells in 350 μl of Incubation Buffer and analyze on flow cytometer; for unconjugated or biotinylated primary antibodies, proceed to Step 5.
  5. Resuspend cells in fluorochrome-conjugated secondary antibody diluted in Incubation Buffer at the recommended dilution.
  6. Incubate for 30 min at room temperature.
  7. Wash 2X by centrifugation in Incubation Buffer.
  8. Resuspend cells in 350 μl of Incubation Buffer and analyze on flow cytometer.

posted December 2013

Protocol Id: 626

Specificity / Sensitivity

Brachyury (D2Z3J) Rabbit mAb (PE Conjugate) recognizes endogenous levels of total Brachyury protein.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro256 of human Brachyury protein.

Background

Brachyury protein, encoded by the T gene, is a transcription factor that is vital for the formation of posterior mesoderm and axial development during vertebrate embryogenesis (1). In the mouse, brachyury is necessary for mesodermal morphogenetic cell movements during gastrulation. Brachyury mutant mice die in utero and display deficient mesoderm formation including an abnormal notochord, missing posterior somites, and a reduced allantois (2). Human brachyury is expressed in the notochord, as well as in chordoma tumors that occur along the spine, making it a good marker for notochord and notochord-derived tumors (3,4). A common polymorphism in the human T gene has also been shown to be associated with development of the multifactorial neural tube defect, spina bifida (5,6).

  1. Edwards, Y.H. et al. (1996) Genome Res 6, 226-33.
  2. Wilson, V. et al. (1995) Development 121, 877-86.
  3. Vujovic, S. et al. (2006) J Pathol 209, 157-65.
  4. Yang, X.R. et al. (2009) Nat Genet 41, 1176-8.
  5. Morrison, K. et al. (1996) Hum Mol Genet 5, 669-74.
  6. Jensen, L.E. et al. (2004) Hum Genet 115, 475-82.

Pathways & Proteins

Explore pathways + proteins related to this product.

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's 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's 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.

For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
Powered by Translations.com GlobalLink OneLink SoftwarePowered By OneLink