Alexa Fluor® 594 anti-human CD326 (EpCAM) Antibody

Pricing & Availability
Clone
9C4 (See other available formats)
Regulatory Status
RUO
Other Names
Ep-CAM, tumor associated calcium signal transducer 1, epithelial cell surface antigen, epithelial glycoprotein 2, EGP2, adenocarcinoma associated antigen, TROP1
Isotype
Mouse IgG2b, κ
Ave. Rating
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Product Citations
publications
1_9C4_A594_CD326_Antibody_1_102820
Human paraffin-embedded placenta tissue slices were prepared with a standard protocol of deparaffination and rehydration. Antigen retrieval was done with Tris-Buffered Saline 1X (1.0M, pH7.4) at 95°C for 40 minutes. Tissue was washed with PBS/ 0.05% Tween20 twice for five minutes and blocked with 5% FBS and 0.2% Gelatin for 30 minutes. Then, the tissue was stained with 5µg/ml of anti-CD326 (clone 9C4) Alexa Fluor® 594 (red) at 4°C overnight. The nuclei were conterstained with DAPI(blue). The image was captured with a 10X objective.
  • 1_9C4_A594_CD326_Antibody_1_102820
    Human paraffin-embedded placenta tissue slices were prepared with a standard protocol of deparaffination and rehydration. Antigen retrieval was done with Tris-Buffered Saline 1X (1.0M, pH7.4) at 95°C for 40 minutes. Tissue was washed with PBS/ 0.05% Tween20 twice for five minutes and blocked with 5% FBS and 0.2% Gelatin for 30 minutes. Then, the tissue was stained with 5µg/ml of anti-CD326 (clone 9C4) Alexa Fluor® 594 (red) at 4°C overnight. The nuclei were conterstained with DAPI(blue). The image was captured with a 10X objective.
  • 2_9C4_A594_CD326_Antibody_2_102820
    Human colorectal adenocarcinoma cell line HT-29 was fixed with 1% paraformaldahyde (PFA), then stained with 10 µg/ml anti-human CD326 (clone 9C4) Alexa Fluor® 594 (red). Nuclei were counterstained with DAPI (blue). The image was acquired with a 40X objective.
  • 3_67_Human_Metastatic_Lymph_Node_CD4_EpCAM
    Confocal image of human metastatic lymph node sample acquired using the IBEX method of highly multiplexed antibody-based imaging: CD4 (blue) in Cycle 1 and EpCAM (red) in Cycle 3. Tissues were prepared using ~1% (vol/vol) formaldehyde and a detergent. Following fixation, samples are immersed in 30% (wt/vol) sucrose for cryoprotection. Images are courtesy of Drs. Andrea J. Radtke and Ronald N. Germain of the Center for Advanced Tissue Imaging (CAT-I) in the National Institute of Allergy and Infectious Diseases (NIAID, NIH).
Compare all formats See Alexa Fluor® 594 spectral data
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324228 100 µg 270 CHF
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Description

CD326 is also known as Ep-CAM, tumor associated calcium signal transducer 1, epithelial cell surface antigen, epithelial glycoprotein 2, EGP2, adenocarcinoma associated antigen, and TROP1. CD326 is a type I transmembrane protein containing six disulfide bridges and one THYRO domain. This cell surface glycosylated 40 kD protein is highly expressed in bone marrow, colon, lung, and most normal epithelial cells and is expressed on carcinomas of gastrointestinal origin. Recently, it has been reported that CD326 expression occurs during the early steps of erythrogenesis. CD326 functions as a homotypic calcium-independent cell adhesion molecule and is believed to be involved in carcinogenesis by its ability to induce genes involved in cellular metabolism and proliferation. CD326 antigen is an immunotherapeutic target for the treatment of human carcinomas.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human
Antibody Type
Monoclonal
Host Species
Mouse
Immunogen
DU.4475 breast carcinoma
Formulation
Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide.
Preparation
The antibody was purified by affinity chromatography and conjugated with Alexa Fluor® 594 under optimal conditions.
Concentration
0.5 mg/mL
Storage & Handling
The antibody solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application

IHC-P - Quality tested
ICC - Verified
SB - Reported in the literature, not verified in house

Recommended Usage

Each lot of this antibody is quality control tested by formalin-fixed paraffin-embedded immunohistochemical staining. For immunohistochemistry, the suggested use of this reagent is 5.0 - 10 µg per mL. For immunocytochemistry, a concentration range of 2.5 - 10 μg per mL is recommended. It is recommended that the reagent be titrated for optimal performance for each application.

* Alexa Fluor® 594 has an excitation maximum of 590 nm, and a maximum emission of 617 nm.


Alexa Fluor® and Pacific Blue™ are trademarks of Life Technologies Corporation.

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Excitation Laser
Green Laser (532 nm)/Yellow-Green Laser (561 nm)
Application Notes

Additional reported applications (for the revelant formats) include: immunofluorescence, immunohistochemistry3, and spatial biology (IBEX)4,5.

Additional Product Notes

Iterative Bleaching Extended multi-pleXity (IBEX) is a fluorescent imaging technique capable of highly-multiplexed spatial analysis. The method relies on cyclical bleaching of panels of fluorescent antibodies in order to image and analyze many markers over multiple cycles of staining, imaging, and, bleaching. It is a community-developed open-access method developed by the Center for Advanced Tissue Imaging (CAT-I) in the National Institute of Allergy and Infectious Diseases (NIAID, NIH).

Application References

(PubMed link indicates BioLegend citation)
  1. Lammers R, et al. 2002. Exp. Hematol. 30:537.
  2. Schultz LD, et al. 2010. P. Natl. Acad. Sci. USA 107:13022. PubMed
  3. Human Protein Atlas http://www.proteinatlas.org/ENSG00000119888/antibody (IHC)
  4. Radtke AJ, et al. 2020. Proc Natl Acad Sci USA. 117:33455-33465. (SB) PubMed
  5. Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
Product Citations
  1. Kure K, et al. 2020. Oncol Lett. 19:2286. PubMed
  2. Roy J, et al. 2020. Theranostics. 10:5778. PubMed
  3. Kenerson HL, et al. 2021. STAR Protocols. 2(2):100574. PubMed
RRID
AB_2563209 (BioLegend Cat. No. 324228)

Antigen Details

Structure
Type I transmembrane protein, contains six disulfide bridges, one THYRO domain, approximate molecular weight 40 kD.
Distribution

Highly expressed in bone marrow, colon, lung, and most normal epithelial cells. Also highly expressed on carcinomas of gastrointestinal origin. Expressed during early erythrogenesis.

Function
Homotypic calcium-independent cell adhesion. CD326 is believed to be involved in carcinogenesis by its ability to induce genes involved in cellular metabolism and proliferation.
Modification
Glycosylated.
Cell Type
Embryonic Stem Cells, Epithelial cells
Biology Area
Cell Biology, Immunology, Stem Cells
Molecular Family
Adhesion Molecules, CD Molecules
Antigen References

1. Strnad J, et al. 1989. Cancer Res. 49:314.
2. Munz M, et al. 2004. Oncogene 23:5748.
3. Rao CG, et al. 2005. Int. J. Oncol. 27:49.

Gene ID
4072 View all products for this Gene ID
UniProt
View information about CD326 on UniProt.org

Related FAQs

If an antibody clone has been previously successfully used in IBEX in one fluorescent format, will other antibody formats work as well?

It’s likely that other fluorophore conjugates to the same antibody clone will also be compatible with IBEX using the same sample fixation procedure. Ultimately a directly conjugated antibody’s utility in fluorescent imaging and IBEX may be specific to the sample and microscope being used in the experiment. Some antibody clone conjugates may perform better than others due to performance differences in non-specific binding, fluorophore brightness, and other biochemical properties unique to that conjugate.

Will antibodies my lab is already using for fluorescent or chromogenic IHC work in IBEX?

Fundamentally, IBEX as a technique that works much in the same way as single antibody panels or single marker IF/IHC. If you’re already successfully using an antibody clone on a sample of interest, it is likely that clone will have utility in IBEX. It is expected some optimization and testing of different antibody fluorophore conjugates will be required to find a suitable format; however, legacy microscopy techniques like chromogenic IHC on fixed or frozen tissue is an excellent place to start looking for useful antibodies.

Are other fluorophores compatible with IBEX?

Over 18 fluorescent formats have been screened for use in IBEX, however, it is likely that other fluorophores are able to be rapidly bleached in IBEX. If a fluorophore format is already suitable for your imaging platform it can be tested for compatibility in IBEX.

The same antibody works in one tissue type but not another. What is happening?

Differences in tissue properties may impact both the ability of an antibody to bind its target specifically and impact the ability of a specific fluorophore conjugate to overcome the background fluorescent signal in a given tissue. Secondary stains, as well as testing multiple fluorescent conjugates of the same clone, may help to troubleshoot challenging targets or tissues. Using a reference control tissue may also give confidence in the specificity of your staining.

How can I be sure the staining I’m seeing in my tissue is real?

In general, best practices for validating an antibody in traditional chromogenic or fluorescent IHC are applicable to IBEX. Please reference the Nature Methods review on antibody based multiplexed imaging for resources on validating antibodies for IBEX.

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*These products may be covered by one or more Limited Use Label Licenses (see the BioLegend Catalog or our website, www.biolegend.com/ordering#license). BioLegend products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products, reverse engineer functionally similar materials, or to provide a service to third parties without written approval of BioLegend. By use of these products you accept the terms and conditions of all applicable Limited Use Label Licenses. Unless otherwise indicated, these products are for research use only and are not intended for human or animal diagnostic, therapeutic or commercial use.

 

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This data display is provided for general comparisons between formats.
Your actual data may vary due to variations in samples, target cells, instruments and their settings, staining conditions, and other factors.
If you need assistance with selecting the best format contact our expert technical support team.

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