Nanog Homeobox (NANOG) antibody

Details for Product No. ABIN967666
Request Want additional data for this product?

The Independent Validation Initiative strives to provide you with high quality data. Find out more

Antigen
Synonyms 2410002E02Rik, ENK, ecat4
Reactivity
Mouse (Murine)
(162), (42), (20), (15), (3), (1), (1)
Host
Mouse (BALB/c)
(122), (51), (12), (3)
Clonality (Clone)
Monoclonal ()
Conjugate
Un-conjugated
(7), (6), (6), (5), (5), (5), (2), (2), (1), (1), (1), (1), (1), (1)
Application
Western Blotting (WB), BioImaging (BI)
(147), (100), (40), (34), (31), (23), (18), (16), (10), (5), (1), (1)
Pubmed 6 references available
Quantity 0.1 mg
Options
Shipping to United States (Change)
Request Want additional data for this product?

The Independent Validation Initiative strives to provide you with high quality data. Find out more

Catalog No. ABIN967666
Contact our Customer Service for availability and price in your country.
Add to Basket

Order hotline:

  • +1 404 474 4654
  • +1 888 205 9894 (TF)
Immunogen Mouse Nanog Recombinant Protein
Clone M55-312
Isotype IgG1, kappa
Characteristics 1. Please refer to us for technical protocols.
2. For fluorochrome spectra and suitable instrument settings, please refer to us.
3. Triton is a trademark of the Dow Chemical Company.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
Purification Purified from tissue culture supernatant or ascites by affinity chromatography.
Alternative Name Nanog
Background The M55-312 monoclonal antibody reacts with mouse Nanog (named for Tir Na Nog, the land of the ever-young of Celtic mythology), which is a homeobox transcription factor required for the maintenance of the undifferentiated state of pluripotent stem cells. Nanog expression counteracts the differentiation-promoting signals induced by the extrinsic factors LIF (Leukemia Inhibitory Factor) and BMP (Bone Morphogenic Protein). When Nanog expression is down-regulated, cell differentiation can proceed. Proteins that regulate Nanog expression include transcription factors Oct4, SOX2, FoxD3, and Tcf3 and tumor suppressor p53.
Molecular Weight 42 kDa
Research Area Embryogenesis, Embryonic stem cells, Cell Cycle
Application Notes Recommended Protocol for Bioimaging:
1. Seed the cells in appropriate culture medium at an appropriate cell density in an 96-well Imaging Plate, and culture overnight to 48 hours.
2. Remove the culture medium from the wells, and wash (one to two times) with 100 myl of 1× PBS.
3. Fix the cells by adding 100 µl of fresh 3.7% Formaldehyde in PBS or fixation buffer to each well and incubating for 10 minutes at room temperature (RT).
4. Remove the fixative from the wells, and wash the wells (one to two times) with 100 myl of 1× PBS.
5. Permeabilize the cells using either cold methanol (a), Triton™ X-100 (b), or Saponin (c): a. Add 100 µl of -20°C 90% methanol to each well and incubate for 5 minutes at RT. b. Add 100 µl of 0.1% Triton™ X-100 to each well and incubate for 5 minutes at RT. c. Add 100 µl of 1× Perm/Wash buffer to each well and incubate for 15 to 30 minutes at RT. Continue to use 1× Perm/Wash buffer for all subsequent wash and dilutions steps.
6. Remove the permeabilization buffer from the wells, and wash one to two times with 100 myl of appropriate buffer (either 1× PBS or 1× Perm/Wash buffer, see step 5.c.).
7. Optional blocking step: Remove the wash buffers, and block the cells by adding 100 µl of blocking buffer or 3% FBS in appropriate dilution buffer to each well and incubating for 15 to 30 minutes at RT.
8. Dilute the antibody to its optimal working concentration in appropriate dilution buffer. Titrate purified (unconjugated) antibodies and second-step reagents to determine the optimal concentration. If using a Bioimaging Certified antibody conjugate, dilute it 1:10.
9. Add 50 µl of diluted antibody per well and incubate for 60 minutes at RT. Incubate in the dark if using fluorescently labeled antibodies.
10. Remove the antibody, and wash the wells three times with 100 myl of wash buffer. An optional detergent wash (100 myl of 0.05% Tween in 1× PBS) can be included prior to the regular wash steps.
11. If the antibody being used is fluorescently labeled, then move to step 12. Otherwise, if using a purified unlabeled antibody, repeat steps 8 to 10 with a fluorescently labeled second-step reagent to detect the purified antibody.
12. After the final wash, counter-stain the nuclei by adding 100 ml of a 2 mg/ml solution of Hoechst 33342 in 1× PBS to each well at least 15 minutes before imaging.
13. View and analyze the cells on an appropriate imaging instrument.
Restrictions For Research Use only
Format Liquid
Concentration 0.5 mg/ml
Buffer Aqueous buffered solution.
Preservative Sodium azide
Precaution of Use This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Storage 4 °C
Supplier Images
anti-Nanog Homeobox (NANOG) antibody Western blot analysis of anti-mouse Nanog. Mouse embryonic stem cell lysate E14 12.5 µg/lane was probed with the Nanog monoclonal antibody at the following concentrations: 2.0 (lane 1), 1.0 (lane 2), and 0.5 µg/ml (lane 3). Nanog is identified as a 42 KDa band.
anti-Nanog Homeobox (NANOG) antibody (2) Immunofluorescent staining of mouse embryonic stem (ES) cell line. ES-E14TG2a cells (ATCC CRL-1821) were seeded in a 96-well imaging plate at ~10,000 cells per well. After overnight incubation, the cells were fixed, permeabilized with Triton™ X-100, and stained with Mouse anti-Mouse Nanog antibody according to the Recommended Assay Procedure. The second step reagent was Alexa Fluor® 555 goat anti-mouse Ig (Invitrogen) (pseudo colored green). Cell nuclei were counterstained with Hoechst 33342 (pseudo colored blue). The images were captured on a BD Pathway™ 435 High-Content Bioimager System using a 20X objective and merged using BD AttoVision™ software. This antibody also stains F9 cells (mouse embryonal carcinoma, ATCC CRL-1720). It also worked with the Saponin and cold methanol fix/perm protocols, however Saponin permeabilization resulted in higher background staining.
anti-Nanog Homeobox (NANOG) antibody (3) Immunofluorescent staining of mouse embryonic stem (ES) cell line
Product cited in: Mitsui, Tokuzawa, Itoh et al.: "The homeoprotein Nanog is required for maintenance of pluripotency in mouse epiblast and ES cells." in: Cell, Vol. 113, Issue 5, pp. 631-42, 2003 (PubMed).

Chambers, Colby, Robertson et al.: "Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells." in: Cell, Vol. 113, Issue 5, pp. 643-55, 2003 (PubMed).

Chambers: "The molecular basis of pluripotency in mouse embryonic stem cells." in: Cloning and stem cells, Vol. 6, Issue 4, pp. 386-91, 2005 (PubMed).

Suzuki, Raya, Kawakami et al.: "Nanog binds to Smad1 and blocks bone morphogenetic protein-induced differentiation of embryonic stem cells." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 103, Issue 27, pp. 10294-9, 2006 (PubMed).

Sun, Li, Yang et al.: "Mechanisms controlling embryonic stem cell self-renewal and differentiation." in: Critical reviews in eukaryotic gene expression, Vol. 16, Issue 3, pp. 211-31, 2006 (PubMed).

Pan, Thomson: "Nanog and transcriptional networks in embryonic stem cell pluripotency." in: Cell research, Vol. 17, Issue 1, pp. 42-9, 2007 (PubMed).

Validation Images
Did you look for something else?
back to top