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Purified Mouse anti-CDX-2
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Purified Mouse anti-CDX-2
Western Blot analysis of CDX-2 in human colorectal adenocarcinoma. Lysate from DLD-1 cells (ATCC CCL-221) was probed with Purified Mouse anti-CDX-2 monoclonal antibody at titrations of 0.5 (lane 1), 0.25 (lane 2), and 0.125 µg/ml (lane 3).  CDX-2 is identified as a band of 38-40 kDa.
Purified Mouse anti-CDX-2

Immunofluorescent staining of human colorectal adenocarcinoma.  DLD-1 cells (ATCC CCL-221) were seeded in a 96-well imaging plate (Cat. No. 353219) at ~10,000 cells per well.  After overnight incubation, the cells were fixed, permeabilized with Triton™ X-100, and stained with Purified Mouse anti-CDX-2 (pseudocolored green) according to the Recommended Assay Procedure.  The second-step reagent was Alexa Fluor® 555 goat anti-mouse Ig (Invitrogen), and counterstaining was with Hoechst 33342 (pseudocolored blue).  The left image shows CDX-2 alone, and the right image shows CDX-2 merged with Hoechst staining.  Images were captured on a BD Pathway™ 435 confocal bioimager using a 20x objective and merged using BD AttoVision™ software.  This antibody also worked with both the alcohol and saponin fix/perm protocols (see Recommended Assay Procedure).

Western Blot analysis of CDX-2 in human colorectal adenocarcinoma. Lysate from DLD-1 cells (ATCC CCL-221) was probed with Purified Mouse anti-CDX-2 monoclonal antibody at titrations of 0.5 (lane 1), 0.25 (lane 2), and 0.125 µg/ml (lane 3).  CDX-2 is identified as a band of 38-40 kDa.

Immunofluorescent staining of human colorectal adenocarcinoma.  DLD-1 cells (ATCC CCL-221) were seeded in a 96-well imaging plate (Cat. No. 353219) at ~10,000 cells per well.  After overnight incubation, the cells were fixed, permeabilized with Triton™ X-100, and stained with Purified Mouse anti-CDX-2 (pseudocolored green) according to the Recommended Assay Procedure.  The second-step reagent was Alexa Fluor® 555 goat anti-mouse Ig (Invitrogen), and counterstaining was with Hoechst 33342 (pseudocolored blue).  The left image shows CDX-2 alone, and the right image shows CDX-2 merged with Hoechst staining.  Images were captured on a BD Pathway™ 435 confocal bioimager using a 20x objective and merged using BD AttoVision™ software.  This antibody also worked with both the alcohol and saponin fix/perm protocols (see Recommended Assay Procedure).

商品详情
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BD Pharmingen™
CDX2, CDX3, CDX-3
Human (QC Testing)
Mouse BALB/c IgG1, κ
Human CDX-2 Recombinant Protein
Western blot (Routinely Tested), Bioimaging (Tested During Development)
38-40 kDa
0.5 mg/ml
AB_1645587
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


推荐的实验流程

Bioimaging

1.        Seed the cells in appropriate culture medium at an appropriate cell density in a BD Falcon™ 96-well Imaging Plate (Cat. No. 353219), and

        culture overnight to 48 hours.

2.        Remove the culture medium from the wells, wash (one to two times) with 100 μl of 1× PBS.

3.        Fix the cells by adding 100 µl of fresh 3.7% Formaldehyde in PBS or BD Cytofix™ fixation buffer (Cat. No. 554655) 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 μl 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 or -20°C BD™ Phosflow Perm Buffer III (Cat. No. 558050) 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 (Cat. No. 554723) 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 μl 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 BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656) 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 μl of wash buffer.  An optional detergent wash (100 μl 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 μl of a 2 μg/ml solution of Hoechst 33342 (eg, Sigma-Aldrich Cat. No. B2261) in 1× PBS to each well at least 15 minutes before imaging.

13.        View and analyze the cells on an appropriate imaging instrument.  

Bioimaging:  For more detailed information please refer to http://www.bdbiosciences.com/support/resources/protocols/ceritifed_reagents.jsp

Western blot:  For more detailed information please refer to http://www.bdbiosciences.com/pharmingen/protocols/Western_Blotting.shtml

商品通知

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. 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.
  4. Triton is a trademark of the Dow Chemical Company.
  5. This antibody has been developed and certified for the bioimaging application. However, a routine bioimaging test is not performed on every lot. Researchers are encouraged to titrate the reagent for optimal performance.
560171 Rev. 1
抗体详情
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M39-711

The M39-711 monoclonal antibody reacts with caudal-type homeobox transcription factor 2 (CDX-2).  In embryonic development, CDX-2 is first expressed in the outer layer of the morula and later in the trophoblast, but not in embryonic stem cells.  Along with Oct3/4, it is involved in the segregation of the trophoectoderm lineage from the inner cell mass.  In the adult, CDX-2 expression is limited to the mucosa of the small and large intestines, where it regulates the synthesis of several intestine-specific proteins.  Loss of CDX-2 expression is observed in some colorectal carcinomas, while gain of CDX-2 expression has been observed in gastric tumors and acute myeloid leukemias.

560171 Rev. 1
格式详情
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
560171 Rev.1
报价单和参考
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研发参考 (6)

  1. Hinoi T, Loda M, Fearon ER. Silencing of CDX2 expression in colon cancer via a dominant repression pathway. J Biol Chem. 2003; 278(45):44608-44616. (Biology). 查看参考
  2. Qualtrough D, Hinoi T, Fearon E, Paraskeva C. Expression of CDX2 in normal and neoplastic human colon tissue and during differentiation of an in vitro model system. Gut. 2002; 51:184-190. (Biology). 查看参考
  3. Ralston A, Rossant J. Genetic regulation of stem cell origins in the mouse embryo. Clin Genet. 2005; 68(2):106-112. (Biology). 查看参考
  4. Rawat VP, Cusan M, Deshpande A, et al. Ectopic expression of the homeobox gene Cdx2 is the transforming event in a mouse model of (12;13)(p13;q12) acute myeloid leukemia. Proc Natl Acad Sci U S A. 2004; 101(3):817-822. (Biology). 查看参考
  5. Scholl C, Bansal D, Döhner K, et al. The homeobox gene CDX2 is aberrantly expressed in most cases of acute myeloid leukemia and promotes leukemogenesis. J Clin Invest. 2007; 117(4):865-868. (Biology). 查看参考
  6. Tsukamoto T, Mizoshita T, Tatematsu M. Gastric-and-intestinal mixed-type intestinal metaplasia: aberrant expression of transcription factors and stem cell intestinalization. Gastric Cancer. 2006; 9(3):156-166. (Biology). 查看参考
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560171 Rev. 1

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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.