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RB744 Mouse Anti-Human CD326 (EpCAM)
RB744 Mouse Anti-Human CD326 (EpCAM)
Flow cytometric analysis of CD326 (EpCAM) expression on Human SK-BR-3 cells. Cells from the Human SK-BR-3 (Breast adenocarcinoma, ATCC® HTB-30™) cell line were stained with either BD Horizon™ RB744 Mouse IgG1, κ Isotype Control (Cat. No. 570519; dashed line histogram) or BD Horizon™ RB744 Mouse Anti-Human CD326 (EpCAM) antibody (Cat. No. 570597/570685; solid line histogram). DAPI Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histogram showing the expression of CD326 (EpCAM) [or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) cells. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 Cell Analyzer System and FlowJo™ Software.
Flow cytometric analysis of CD326 (EpCAM) expression on Human SK-BR-3 cells. Cells from the Human SK-BR-3 (Breast adenocarcinoma, ATCC® HTB-30™) cell line were stained with either BD Horizon™ RB744 Mouse IgG1, κ Isotype Control (Cat. No. 570519; dashed line histogram) or BD Horizon™ RB744 Mouse Anti-Human CD326 (EpCAM) antibody (Cat. No. 570597/570685; solid line histogram). DAPI Solution (Cat. No. 564907) was added to cells right before analysis. The fluorescence histogram showing the expression of CD326 (EpCAM) [or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) cells. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 Cell Analyzer System and FlowJo™ Software.
Product Details
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BD Horizon™
EPCAM; EGP; ESA; GA733-2; hEGP-2; KSA; M4S1; MIC18; MK-1; TACSTD1; TROP1
Human (QC Testing)
Mouse BALB/c IgG1, λ
Breast carcinoma–associated mucin BCA-225
Flow cytometry (Routinely Tested)
5 µl/test
4072
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions and unreacted dye was removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.

Recommended Assay Procedures

BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation). When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

Product Notices

  1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  2. 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.
  3. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. An isotype control should be used at the same concentration as the antibody of interest.
  6. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  7. Human donor specific background has been observed in relation to the presence of anti-polyethylene glycol (PEG) antibodies, developed as a result of certain vaccines containing PEG, including some COVID-19 vaccines. We recommend use of BD Horizon Brilliant™ Stain Buffer in your experiments to help mitigate potential background. For more information visit https://www.bdbiosciences.com/en-us/support/product-notices.
  8. When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
  9. Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
  10. Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
  11. For U.S. patents that may apply, see bd.com/patents.
570597 Rev. 1
Antibody Details
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EBA-1

The EBA-1 monoclonal antibody specifically binds to human CD326. CD326 is an approximately 40 kDa type 1 transmembrane glycoprotein and adhesion molecule that mediates intercellular adhesive interactions. CD326 is also known as epithelial adhesion molecule (EpCAM), epithelial glycoprotein 2 (EGP-2), and epithelial surface antigen (ESA). The epithelial cells present in non-squamous epithelia and tumors derived from such cells show EpCAM expression. The normal epithelial cells reactive with anti-EpCAM antibodies are those present in the (lower) respiratory tract; the (lower) gastrointestinal tract; tubules in the kidney; the surface epithelium of the ovary; the exocrine and endocrine pancreas; secondary germ cells of telogenic hair follicles; and secretory tubules of sweat glands in the skin, whereas the epidermis is negative. In addition, all epithelial cells in the thyroid and epithelial cells in the thymus show EpCAM expression, while the outer cortex and Hassall's corpuscles have low expression. In the liver, only the bile ducts appear to be positive with anti-EpCAM antibodies. Non-squamous- carcinoma cells have high EpCAM expression; some squamous carcinoma cells. Tumors arising from non-epithelial cells, such as lymphoma, mesothelioma, neuroblastoma, and melanoma, do not express EpCAM.

570597 Rev. 1
Format Details
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RB744
The BD Horizon RealBlue™ 744 (RB744) Dye is part of the BD® family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 498-nm and an emission maximum (Em Max) at 746-nm as measured using an antibody-dye conjugate. Driven by BD® innovation, RB744 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with minimal excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), we recommend using an optical filter centered near 750-nm (e.g., a 750/60-nm bandpass filter).
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RB744
Blue 488 nm
498 nm
746 nm
570597 Rev.1
Citations & References
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View product citations for antibody "570597" on CiteAb

Development References (13)

  1. Braun S, Pantel K, Müller P, et al. Cytokeratin-positive cells in the bone marrow and survival of patients with stage I, II, or III breast cancer. N Engl J Med. 2000; 342:525-533. (Biology). View Reference
  2. Carlsten M, Bjorkstrom NK, Norell H, et al. DNAX accessory molecule-1 mediated recognition of freshly isolated ovarian carcinoma by resting natural killer cells. Cancer Res. 2007; 67(3):1317-1325. (Clone-specific: Flow cytometry). View Reference
  3. De Leij L, Helrich W, Stein R, Mattes MJ. SCLC-cluster-2 antibodies detect the pancarcinoma/epithelial glycoprotein EGP-2. Int J Cancer. 1994; 8:60-63. (Biology). View Reference
  4. Diel IJ, Kaufmann M, Goerner R, Costa SD, Kaul S, Bastert G. Detection of tumor cells in bone marrow of patients with primary breast cancer: a prognostic factor for distant metastasis. J Clin Oncol. 1992; 10:1534-1539. (Biology). View Reference
  5. Hardingham JE, Kotasek D, Farmer B, et al. Immunobead-PCR: a technique for the detection of circulating tumor cells using immunomagnetic beads and the polymerase chain reaction. Cancer Res. 1993; 53(15):3455-3458. (Biology). View Reference
  6. Latza U, Niedobitek G, Schwarting R, Nekarda H, Stein H. Ber-EP4: new monoclonal antibody which distinguishes epithelia from mesothelial. J Clin Pathol. 1990; 43(3):213-219. (Biology). View Reference
  7. Momburg F, Moldenhauer G, Hämmerling GJ, Möller P. Immunohistochemical study of the expression of a Mr 34,000 human epithelium-specific surface glycoprotein in normal and malignant tissues. Cancer Res. 1987; 47:2883-2891. (Biology). View Reference
  8. Naume B, Borgen E, Beiske K, et al.. Immunomagnetic techniques for the enrichment and detection of isolated breast carcinoma cells in bone marrow and peripheral blood. J Hematother Stem Cell Res. 1997; 6:103-113. (Biology). View Reference
  9. Patriarca C, Macchi RM, Marschner AK, Mellstedt H. Epithelial cell adhesion molecule expression (CD326) in cancer: a short review. Cancer Treat Rev. 2012; 38(1):68-75. (Biology). View Reference
  10. Stahel RA, Gilks WR, Lehmann HP, Schenker T. Third International Workshop on Lung Tumor and Differentiation Antigens: overview of the results of the central data analysis. Int J Cancer. 1994; 8:6-26. (Biology). View Reference
  11. Takao M, Takeda K. Enumeration, characterization, and collection of intact circulating tumor cells by cross contamination-free flow cytometry. Cytometry A. 2011; 79(2):107-117. (Clone-specific: Flow cytometry). View Reference
  12. Trzpis M, McLaughlin PM, de Leij LM, Harmsen MC. Epithelial cell adhesion molecule: more than a carcinoma marker and adhesion molecule. Am J Pathol. 2007; 171(2):386-395. (Biology). View Reference
  13. Yemul S, Leon Ja, Pozniakoff T, Esser PD, Estabrook A. Radioimmunoimaging of human breast carcinoma xenografts in nude mouse model with 111In-labeled new monoclonal antibody EBA-1 and F(ab')2 fragments. Nucl Med Biol. 1993; 20:325-335. (Immunogen: ELISA, Radioimmunoassay, Western blot). View Reference
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570597 Rev. 1

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