Alexa Fluor® 594 anti-mouse CD31 Antibody

Pricing & Availability
Clone
MEC13.3 (See other available formats)
Regulatory Status
RUO
Other Names
PECAM-1, EndoCAM
Isotype
Rat IgG2a, κ
Ave. Rating
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Product Citations
publications
1-MEC13.3_AF594_CD31_Antibody_FC_072414.jpg
C57BL/6 mouse splenocytes were stained with CD31 (clone MEC13.3) Alexa Fluor® 594 (filled histogram) or rat IgG2a, κ Alexa Fluor® 594 isotype control (open histogram). The data was acquired by BD LSRFortessa™ cell analyzer equipped with Yellow-Green Laser (561 nm).
  • 1-MEC13.3_AF594_CD31_Antibody_FC_072414.jpg
    C57BL/6 mouse splenocytes were stained with CD31 (clone MEC13.3) Alexa Fluor® 594 (filled histogram) or rat IgG2a, κ Alexa Fluor® 594 isotype control (open histogram). The data was acquired by BD LSRFortessa™ cell analyzer equipped with Yellow-Green Laser (561 nm).
  • 2-MEC13.3_AF594_CD31_Antibody_IHC_033114
    C57BL/6 mouse frozen liver section was fixed with 4% paraformaldehyde (PFA) for 10 minutes at room temperature and blocked with 5% FBS plus 5% rat serum for 1 hour at room temperature. Then the section was stained with 2.5 µg/ml of CD31 (clone MEC13.3) Alexa Fluor® 594 (red) overnight at 4°C. Nuclei were counterstained with DAPI (blue). The image was captured by 10X objective.
  • 3-MEC133_A594_CD31_Antibody_2_120718
    Dissected C57/B6 mouse stomach was immersed in 4% paraformaldehyde (PFA) overnight followed by 30% sucrose immersion overnight and frozen in OCT. Frozen section was blocked with 5% FBS and 5% mouse serum for 30 minutes at room temperature. Then the tissue section was stained with 5 µg/mL of anti-mouse CD31 (clone MEC13.3) Alexa Fluor® 594 (red) and 2.5 µg/mL anti-mouse CD326 (clone G8.8) Alexa Fluor® 488 (blue) and 2.5 µg/mL anti-mouse Tubulin Beta 3 (clone AA10) Alexa Fluor® 647 (green) overnight at 4°C. The image was captured by 10X objective.
  • 4_MEC133_A594_CD31_Antibody_3D-IHC_092121.png
    Paraformaldehyde-fixed (4%), 500 μm-thick mouse kidney section was processed according to the Ce3DTM Tissue Clearing Kit protocol (cat. no. 427701). The section was costained with anti-mouse Podoplanin Antibody (clone PMab-1) Alexa Fluor® 488 at 5 µg/mL (green), anti-mouse CD31 Antibody (clone MEC13.3) Alexa Fluor® 594 at 5 µg/mL (orange), and anti-Tubulin β 3 (TUBB3) Antibody (Clone TUJ1) Alexa Fluor® 647 at 5 µg/mL (magenta). The section was then optically cleared and mounted in a sample chamber. The image was captured with a 20X objective using Zeiss 780 confocal microscope and processed by Imaris image analysis software.
    Watch the video.
  • 43_Mouse_Lung_CD11c_CD206_CD31
    Confocal image of C57BL/6 mouse lung sample acquired using the IBEX method of highly multiplexed antibody-based imaging: CD11c (magenta) in Cycle 1, CD206 (blue) in Cycle 1, and CD31 (green) in Cycle 4. 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).
  • 47_Mouse_Lung_Ly6G_CD8_CD31
    Confocal image of C57BL/6 mouse lung sample acquired using the IBEX method of highly multiplexed antibody-based imaging: Ly-6G (yellow) in Cycle 2, CD31 (blue) in Cycle 3, and CD8 (red) in Cycle 4. 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).
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Description

CD31 is a 130-140 kD glycoprotein, also known as platelet endothelial cell adhesion molecule (PECAM-1), EndoCAM, and gpIIa. It is a member of the Ig superfamily, expressed on endothelial cells, platelets, granulocytes, monocytes/macrophages, dendritic cells, and T and B cell subsets, and is critical for cell-to-cell interactions. The primary ligands for CD31 have been reported to be CD38 and the vitronectin receptor (αv β3 integrin, CD51/CD61). Other reported functions of CD31 are neutrophil emigration to sites of inflammation, and angiogenesis.

Product Details
Technical data sheet

Product Details

Verified Reactivity
Mouse
Antibody Type
Monoclonal
Host Species
Rat
Immunogen
Polyoma middle T transformed EC line tEnd.1
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

FC, IHC-F - Quality tested
3D IHC - Verified

SB - Reported in the literature, not verified in house

Recommended Usage

Each lot of this antibody is quality control tested by immunohistochemical staining on frozen tissue sections. For immunohistochemistry, a concentration range of 2.0 - 5.0 μg/ml is suggested. For flow cytometric staining, the suggested use of this reagent is ≤ 0.25 µg per million cells in 100 µl volume. For 3D immunohistochemistry on formalin-fixed tissues, a concentration of 5.0 µg/mL is suggested. 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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Application Notes

Anti-mouse CD31 clones 390 and MEC13.3 bind to their respective non-overlapping epitopes in IgD2 of CD31.8 Additional reported applications (in the relevant formats) include: immunoprecipitation1, in vitro and in vivo blocking of CD31-mediated cell-cell interactions1-4, immunohistochemical staining1,5,6 of acetone-fixed frozen sections and zinc-fixed paraffin-embedded sections, and spatial biology (IBEX)12,13.
Special Note: The antibody works well on acetone-fixed frozen sections as well as Zinc-fixed paraffin-embedded sections. It sometime works on formalin-fixed and paraformaldehyde-fixed paraffin-embedded tissue sections but inconsistent results have been reported. This antibody is not recommended for formalin-fixed paraffin-embedded sections or for Western blot analysis. The Ultra-LEAF™ Purified antibody (Endotoxin < 0.01 EU/µg, Azide-Free, 0.2 µm filtered) is recommended for functional assays (Cat. No. 102529 and 102530).

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. Vecchi A, et al. 1994. Eur. J. Cell Biol. 63:247. (IP, IHC, Block)
  2. Christofidou-Solomidou M, et al. 1997. J. Immunol. 158:4872. (Block)
  3. DeLisser HM, et al. 1997. Am. J. Pathol. 151:671. (Block)
  4. Rosenblum WI, et al. 1994. Am. J. Pathol. 145:33. (Block)
  5. Baldwin HS, et al. 1994. Development 120:2539. (IHC)
  6. Voswinckel R, et al. 2003. Circ. Res. 93:372. (IHC)
  7. Leung VW, et al. 2009. Am J. Pathol. 175:1757. PubMed
  8. Chacko AM, et al. 2012. PLoS One 7:e34958.
  9. Giacomini C, et al. 2014. Exp Eye Res. 18:1. PubMed
  10. Morita R, et al. 2015. PNAS. 112:160. PubMed
  11. Ito A, et al. 2015. Brain Res. 1594:310. PubMed
  12. Radtke AJ, et al. 2020. Proc Natl Acad Sci U S A. 117:33455-65. (SB) PubMed
  13. Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
Product Citations
  1. Zhang Y, et al. 2018. Development. 145. PubMed
  2. Baptista AP et al. 2019. Immunity. 50(5):1188-1201 . PubMed
  3. Luan X, et al. 2021. Biomaterials. 275:120910. PubMed
  4. Fan H, et al. 2021. Int J Radiat Oncol Biol Phys. S0360-3016:02845. PubMed
  5. Liu Z, et al. 2016. Sci Rep. 6:34416. PubMed
  6. Nguyen TH, et al. 2022. Nat Microbiol. 7:62. PubMed
  7. Hofmann J, et al. 2021. Front Immunol. 11:599495. PubMed
  8. Tuong ZK, et al. 2021. Cell Rep. 37:110132. PubMed
  9. Kumar D, et al. 2015. PLoS Pathog. 11: 1005333. PubMed
  10. Di Pilato M, et al. 2021. Cell. 184(17):4512-4530.e22. PubMed
RRID
AB_2563319 (BioLegend Cat. No. 102520)

Antigen Details

Structure
Ig superfamily, 130-140 kD
Distribution

Endothelial cells, platelets, granulocytes, monocytes/macrophages, dendritic cells, T and B cell subsets

Function
Adhesion
Ligand/Receptor
CD38, αV3 integrin
Cell Type
B cells, Dendritic cells, Endothelial cells, Granulocytes, Macrophages, Monocytes, Neutrophils, Platelets, T cells
Biology Area
Angiogenesis, Cell Adhesion, Cell Biology, Immunology, Neuroinflammation, Neuroscience
Molecular Family
Adhesion Molecules, CD Molecules
Antigen References

1. Barclay AN, et al. 1997. The Leukocyte Antigen FactsBook Academic Press.
2. DeLisser HM, et al. 1994. Immunol. Today 15:490.
3. Newman PJ, et al. 1990. Science 247:1219.

Gene ID
18613 View all products for this Gene ID
UniProt
View information about CD31 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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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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