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Alexa Fluor® 647 anti-mouse CD23 Antibody

Pricing & Availability
Clone
B3B4 (See other available formats)
Regulatory Status
RUO
Other Names
FcεRII, IgE Fc Receptor, BLAST-2, Ly-42, B6
Isotype
Rat IgG2a, κ
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Product Citations
publications
1-B3B4_Alx647_080207
CD57BL/6 mouse splenocytes were stained with CD23 (clone B3B4) Alexa Fluor® 647 (filled histogram), or rat IgG2a, κ Alexa Fluor® 647 (open histogram).
  • 1-B3B4_Alx647_080207
    CD57BL/6 mouse splenocytes were stained with CD23 (clone B3B4) Alexa Fluor® 647 (filled histogram), or rat IgG2a, κ Alexa Fluor® 647 (open histogram).
  • 2-B3B4_A647_CD23_Antibody_IHCF_051519
    C57BL/6 mouse frozen spleen section was fixed with 4% paraformaldehyde (PFA) for ten minutes at room temperature and blocked with 5% FBS+ rat/mouse/hamster serum for 30 minutes at room temperature. Then the section was stained with 5 µg/ml of Alexa Fluor® 647 anti-mouse CD23 (red) (clone B3B4), 5 µg/ml Alexa Fluor® 488 anti-mouse CD3ε (green) (clone 145-2C11), and 5 µg/ml of Alexa Fluor® 594 anti-mouse F4/80 Antibody (blue) (Clone BM8) overnight at 4°C. The image was captured by 10X objective.
  • 60_Mouse_Lymph_Node_PD1_CD23_IgD
    Mice were injected subcutaneously with sheep red blood cells in a volume of 25 µl per site on days 0 and 4 and harvested on day 11. Confocal image of C57BL/6 mouse lymph node acquired using the IBEX method of highly multiplexed antibody-based imaging: PD-1 (green) in Cycle 1, CD23 (magenta) in Cycle 7, and IgD (blue) in Cycle 10. 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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101611 25 µg £70
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Description

CD23 is a 45 kD protein also known as low affinity IgE Fc receptor, FcεRII, BLAST-2, Ly-42, or B6. It is a member of the Ig family, expressed on conventional B (but not B-1) cells and follicular dendritic cells. CD23 responds to high levels of IgE by downregulating IgE secretion.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Mouse
Antibody Type
Monoclonal
Host Species
Rat
Immunogen
Complex of IgE with Fcε receptor isolated from the mouse B hybridoma cell line O1.2B2
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® 647 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 - Quality tested
IHC-F - Verified

SB - Reported in the literature, not verified in house
Recommended Usage

Each lot of this antibody is quality control tested by immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is ≤ 0.25 µg per million cells in 100 µl volume. For immunohistochemistry on frozen tissue sections, a concentration range of 2.5 - 10 µg/ml is suggested. It is recommended that the reagent be titrated for optimal performance for each application.

* Alexa Fluor® 647 has a maximum emission of 668 nm when it is excited at 633 nm / 635 nm.


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

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Excitation Laser
Red Laser (633 nm)
Application Notes

The B3B4 antibody is useful for blocking IgE activity in vivo. Additional reported applications (for the relevant formats) include: immunoprecipitation1, immunofluorescence microscopy, in vitro and in vivo blocking of ligand binding2-4, immunohistochemical staining of acetone-fixed frozen sections5, and spatial biology (IBEX)7,8.

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. Waldschmidt TJ, et al. 1988. J. Immunol. 140:2148. (IP)
  2. Rao M, et al. 1987. J. Immunol. 138:1845. (Block)
  3. Oshiba A, et al. 1997. J. Immunol. 159:4056. (Block)
  4. Dasic G, et al. 1999. Eur. J. Immunol. 29:2957. (Block)
  5. Maeda K, et al. 1992. J. Immunol. 148:2340. (IHC)
  6. Craig VJ, et al. 2011. Cancer Res. 71:3616. PubMed
  7. Radtke AJ, et al. 2020. Proc Natl Acad Sci U S A. 117:33455-65. (SB) PubMed
  8. Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
Product Citations
  1. Lino AC et al. 2018. Immunity. 49(1):120-133 . PubMed
  2. Duan L, et al. 2021. Immunity. 54:2256. PubMed
  3. Noviski M, et al. 2018. Elife. 7:e35074. PubMed
  4. Shiozawa S, et al. 2022. iScience. 25:103537. PubMed
RRID
AB_2103038 (BioLegend Cat. No. 101611)
AB_2103038 (BioLegend Cat. No. 101612)

Antigen Details

Structure
C-lectin family, type II transmembrane glycoprotein, 45 kD
Distribution

B cells, follicular dendritic cells

Function
Regulates B cell activation
Ligand/Receptor
IgE, CD19/CD21/CD81
Cell Type
B cells, Dendritic cells
Biology Area
Immunology
Molecular Family
CD Molecules, Fc Receptors
Antigen References

1. Barclay A, et al. 1997. The Leukocyte Antigen FactsBook Academic Press.
2. Delespesse G, et al. 1992. Immunol. Rev. 125:77.
3. Flores-Romo L, et al. 1993. Science 261:1038.

Gene ID
14128 View all products for this Gene ID
UniProt
View information about CD23 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.

Go To Top Version: 3    Revision Date: 04/20/2022

For Research Use Only. Not for diagnostic or therapeutic 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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