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82389
NUT (C52B1) Rabbit mAb (Alexa Fluor® 555 Conjugate)
Antibody Conjugates
Monoclonal Antibody

NUT (C52B1) Rabbit mAb (Alexa Fluor® 555 Conjugate) #82389

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  1. IF
Immunofluorescence Image 1: NUT (C52B1) Rabbit mAb (Alexa Fluor® 555 Conjugate)

Confocal immunofluorescent analysis of rat testis (left, positive) or small intestine (right, negative) using NUT (C52B1) Rabbit mAb (Alexa Fluor® 555 Conjugate) (yellow). Actin filaments were labeled with DyLight™ 650 Phalloidin #12956 (red). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).

To Purchase # 82389S
製品番号 サイズ 価格 在庫
82389S
100 µl  (50 tests)

Supporting Data

REACTIVITY H R
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 Alexa Fluor® 555 fluorescent dye and tested in-house for direct immunofluorescent analysis in rat tissue. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated NUT (C52B1) Rabbit mAb #3625.

Product Usage Information

Application Dilution
Immunofluorescence (Frozen) 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

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Immunofluorescence (Frozen)

A. Solutions and Reagents

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

  1. 20X Phosphate Buffered Saline (PBS): (9808) To prepare 1L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix. Adjust pH to 8.0.
  2. Formaldehyde: 16%, methanol free, Polysciences, Inc. (cat# 18814), use fresh and store opened vials at 4°C in dark, dilute in 1X PBS for use.
  3. Blocking Buffer (1X PBS / 5% normal goat serum (#5425) / 0.3% Triton™ X-100): To prepare 10 ml: add 0.5 ml normal goat serum and 0.5 ml 20X PBS to 9.0 ml dH2O, mix. While stirring, add 30 µl Triton™ X-100.
  4. Antibody Dilution Buffer: (1X PBS / 1% BSA / 0.3% Triton™ X-100): To prepare 10 ml, add 30 µl Triton™ X-100 to 10 ml 1X PBS. Mix well then add 0.1 g BSA (#9998), mix.
  5. Prolong® Gold AntiFade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).

B. Specimen Preparation - Frozen/Cryostat Sections (IF-F)

  1. For fixed frozen tissue proceed with Immunostaining (Section C).
  2. For fresh, unfixed frozen tissue, please fix immediately, as follows:
    1. Cover sections with 4% formaldehyde dilute in 1X PBS.

      NOTE: Formaldehyde is toxic, use only in fume hood.

    2. Allow sections to fix for 15 minutes at room temperature.
    3. Aspirate liquid, rinse three times in 1X PBS for 5 minutes each.
    4. Proceed with Immunostaining (Section C).

C. Immunostaining

NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.

  1. Block specimen in Blocking Buffer for 60 min.
  2. While blocking, prepare primary antibody by diluting as indicated on protocol on product webpage in Antibody Dilution Buffer.
  3. Aspirate blocking solution, apply diluted primary antibody.
  4. Incubate overnight at 4°C.
  5. Rinse three times in 1X PBS for 5 min each.
  6. Coverslip slides with Prolong® Gold Antifade Reagent (#9071) or Prolong® Gold Antifade Reagent with DAPI (#8961).
  7. For best results, allow mountant to cure overnight at room temperature. For long-term storage, store slides flat at 4°C protected from light.

posted November 2006

revised November 2013

Protocol Id: 222

Specificity / Sensitivity

NUT (C52B1) Rabbit mAb (Alexa Fluor® 555 Conjugate) detects endogenous levels of total NUT protein. The antibody also detects endogenous levels of the BRD4-NUT fusion protein found in NUT midline carcinoma (NMC).

Species Reactivity:

Human, Rat

Species predicted to react based on 100% sequence homology:

Monkey

Source / Purification

Monoclonal antibody is produced by immunizing animals with a recombinant protein corresponding to the human NUT protein.

Background

Nuclear protein in testis (NUT) is normally confined to the germ cells of the testis and ovary (1,2). NUT midline carcinoma (NMC) is a recently recognized cancer that is defined by the presence of chromosomal rearrangements involving the NUT gene on chromosome 15q14 (3). In most cases the chromosomal translocation occurs between NUT and BRD4 on chromosome 19, resulting in the formation of a BRD4-NUT fusion protein. In the remaining tumors, variant NUT rearrangements are present involving BRD3, a very close homolog of BRD4. BRD4-NUT and BRD3-NUT encode fusion proteins that appear to contribute to carcinogenesis by blocking epithelial cell differentiation. NMCs, which are aggressive and highly lethal carcinomas, are morphologically indistinguishable from other poorly differentiated carcinomas. Given the limited expression of endogenous NUT protein, this antibody can be used to detect NUT fusion proteins in tissues by immunohistochemistry and immunofluorescence (2).

  1. French, C.A. et al. (2003) Cancer Res 63, 304-7.
  2. Haack, H. et al. (2009) Am J Surg Pathol 33, 984-91.
  3. French, C.A. et al. (2008) Oncogene 27, 2237-42.

Pathways & Proteins

Explore pathways + proteins related to this product.

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For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
DyLight is a trademark of Thermo Fisher Scientific, Inc. and its subsidiaries.
ProLong is a registered trademark of Life Technologies Corporation.
This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer, excluding contract research or any fee for service research, and the buyer must not (1) use this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; or (c) manufacturing or quality assurance or quality control, and/or (2) sell or transfer this product or its components for resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or [email protected]
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.
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