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Alexa Fluor® 647 anti-human CD4 Antibody

Pricing & Availability
Clone
RPA-T4 (See other available formats)
Regulatory Status
RUO
Workshop
IV T114
Other Names
T4
Isotype
Mouse IgG1, κ
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Product Citations
publications
1_RPA-T4_Alexa647_071805
Human peripheral blood lymphocytes stained with RPA-T4 Alexa Fluor® 647
  • 1_RPA-T4_Alexa647_071805
    Human peripheral blood lymphocytes stained with RPA-T4 Alexa Fluor® 647
  • 2_16_Human_Spleen_CD4_Vimentin
    Confocal image of human spleen sample acquired using the IBEX method of highly multiplexed antibody-based imaging: CD4 (cyan) in Cycle 3 and vimentin (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).
  • 3_23_Human_LN_CD4_CD31
    Confocal image of human lymph node sample acquired using the IBEX method of highly multiplexed antibody-based imaging: CD4 (cyan) in Cycle 4 and CD31 (magenta) in Cycle 5. 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® 647 spectral data
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300523 25 tests 90€
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300520 100 tests 215€
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Description

CD4, also known as T4, is a 55 kD single-chain type I transmembrane glycoprotein expressed on most thymocytes, a subset of T cells, and monocytes/macrophages. CD4, a member of the Ig superfamily, recognizes antigens associated with MHC class II molecules, and participates in cell-cell interactions, thymic differentiation, and signal transduction. CD4 acts as a primary receptor for HIV, binding to HIV gp120. CD4 has also been shown to interact with IL-16.

Product Details
Technical Data Sheet (pdf)

Product Details

Verified Reactivity
Human
Reported Reactivity
Chimpanzee
Antibody Type
Monoclonal
Host Species
Mouse
Formulation
Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide and BSA (origin USA)
Preparation
The antibody was purified by affinity chromatography and conjugated with Alexa Fluor® 647 under optimal conditions.
Concentration
Lot-specific (to obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.)
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
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 5 µl per million cells in 100 µl staining volume or 5 µl per 100 µl of whole blood.

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


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

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Excitation Laser
Red Laser (633 nm)
Application Notes

The RPA-T4 antibody binds to the D1 domain of CD4 (CDR1 and CDR3 epitopes) and can block HIV gp120 binding and inhibit syncytia formation. Additional reported applications (for the relevant formats) include: immunohistochemistry of acetone-fixed frozen sections3,4,5, blocking of T cell activation1,2, and spatial biology (IBEX)10,11.  This clone was tested in-house and does not work on formalin fixed paraffin-embedded (FFPE) tissue. The Ultra-LEAF™ purified antibody (Endotoxin < 0.01 EU/µg, Azide-Free, 0.2 µm filtered) is recommended for functional assays (Cat. No. 300569 - 300574).

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. Knapp W, et al. 1989. Leucocyte Typing IV. Oxford University Press. New York. (Activ)
  2. Moir S, et al. 1999. J. Virol. 73:7972. (Activ)
  3. Deng MC, et al. 1995. Circulation 91:1647. (IHC)
  4. Friedman T, et al. 1999. J. Immunol. 162:5256. (IHC)
  5. Mack CL, et al. 2004. Pediatr. Res. 56:79. (IHC)
  6. Lan RY, et al. 2006. Hepatology 43:729.
  7. Zenaro E, et al. 2009. J. Leukoc. Biol. 86:1393. (FC) PubMed
  8. Yoshino N, et al. 2000. Exp. Anim. (Tokyo) 49:97. (FC)
  9. Stoeckius M, et al. 2017. Nat. Methods. 14:865. (PG)
  10. Radtke AJ, et al. 2020. Proc Natl Acad Sci USA. 117:33455-33465. (SB) PubMed
  11. Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
Product Citations
  1. Junker F, et al. 2021. Cytometry A. 99:832. PubMed
  2. Lu T, et al. 2016. PLoS One. 11: 0148044. PubMed
  3. NULL, et al. 2020. J Clin Immunol. 40:1. PubMed
  4. Riese P, et al. 2022. Nat Commun. 13:6894. PubMed
  5. O'Connor MH, et al. 2021. Commun Biol. 4:563. PubMed
  6. Parent AV, et al. 2021. Cell Reports. 36(7):109538. PubMed
RRID
AB_389333 (BioLegend Cat. No. 300523)
AB_389333 (BioLegend Cat. No. 300520)

Antigen Details

Structure
Ig superfamily, type I transmembrane glycoprotein, 55 kD
Distribution

T cell subset, majority of thymocytes, monocytes/macrophages

Function
MHC class II co-receptor, lymphocyte adhesion, thymic differentiation, HIV receptor
Ligand/Receptor
MHC class II molecules, HIV gp120, IL-16
Cell Type
Dendritic cells, Macrophages, Monocytes, T cells, Thymocytes, Tregs
Biology Area
Immunology
Molecular Family
CD Molecules
Antigen References

1. Center D, et al. 1996. Immunol. Today 17:476.
2. Gaubin M, et al. 1996. Eur. J. Clin. Chem. Clin. Biochem. 34:723.

Gene ID
920 View all products for this Gene ID
UniProt
View information about CD4 on UniProt.org

Related FAQs

I am unable to see expression of T cell markers such as CD3 and CD4 post activation.
TCR-CD3 complexes on the T-lymphocyte surface are rapidly downregulated upon activation with peptide-MHC complex, superantigen or cross-linking with anti-TCR or anti-CD3 antibodies. PMA/Ionomycin treatment has been shown to downregulate surface CD4 expression. Receptor downregulation is a common biological phenomenon and so make sure that your stimulation treatment is not causing it in your sample type.
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.

Other Formats

View All CD4 Reagents Request Custom Conjugation
Description Clone Applications
APC anti-human CD4 RPA-T4 FC
Biotin anti-human CD4 RPA-T4 FC
FITC anti-human CD4 RPA-T4 FC,SB
PE anti-human CD4 RPA-T4 FC
PE/Cyanine5 anti-human CD4 RPA-T4 FC
PE/Cyanine7 anti-human CD4 RPA-T4 FC
Purified anti-human CD4 RPA-T4 FC,CyTOF®,Activ,Block,IHC
APC/Cyanine7 anti-human CD4 RPA-T4 FC
Alexa Fluor® 488 anti-human CD4 RPA-T4 FC,ICC
Alexa Fluor® 647 anti-human CD4 RPA-T4 FC,SB
Pacific Blue™ anti-human CD4 RPA-T4 FC
Brilliant Violet 421™ anti-human CD4 RPA-T4 FC,ICC
Alexa Fluor® 700 anti-human CD4 RPA-T4 FC,SB
PerCP anti-human CD4 RPA-T4 FC
PerCP/Cyanine5.5 anti-human CD4 RPA-T4 FC
Brilliant Violet 570™ anti-human CD4 RPA-T4 FC
Brilliant Violet 650™ anti-human CD4 RPA-T4 FC
Purified anti-human CD4 (Maxpar® Ready) RPA-T4 FC,CyTOF®
Alexa Fluor® 594 anti-human CD4 RPA-T4 FC,ICC
Brilliant Violet 510™ anti-human CD4 RPA-T4 FC
PE/Dazzle™ 594 anti-human CD4 RPA-T4 FC
Brilliant Violet 785™ anti-human CD4 RPA-T4 FC
Brilliant Violet 605™ anti-human CD4 RPA-T4 FC
Brilliant Violet 711™ anti-human CD4 RPA-T4 FC
APC/Fire™ 750 anti-human CD4 RPA-T4 FC
CD4 Fluorophore Sampler Kit RPA-T4 FC
CD4 Fluorophore Sampler Kit with Veri-Cells™ PBMC RPA-T4 FC
TotalSeq™-A0072 anti-human CD4 RPA-T4 PG
TotalSeq™-B0072 anti-human CD4 RPA-T4 PG
TotalSeq™-C0072 anti-human CD4 RPA-T4 PG
Ultra-LEAF™ Purified anti-human CD4 RPA-T4 FC,CyTOF®,Activ,Block,IHC
TotalSeq™-D0072 anti-human CD4 RPA-T4 PG
Go To Top Version: 4    Revision Date: 04/25/2022

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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