Block/Neutralize, Dot Blot, Electron Microscopy, ELISA, Flow (Intracellular), Flow Cytometry, Functional, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry Whole-Mount, Immunoprecipitation, In vitro assay, Western Blot

Cited:

Block/Neutralize, Cytometric Bead Assay Standard, Dot Blot, Electron Microscopy, ELISA, Flow Cytometry, Functional Assay, Immunoassay, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, In vitro assay, Western Blot

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG₁ kappa Clone # H5C6

Format

BSA Free

Concentration

1.0 mg/ml

View all CD63 Antibodies »

Product Specifications

Immunogen

Human splenic adherent cells.

Marker

Exosome Marker

Clonality

Monoclonal

Host

Mouse

Isotype

IgG₁ kappa

Scientific Data Images for CD63 Antibody (H5C6) - BSA Free

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Western Blot: CD63 Antibody (H5C6) [NBP2-42225] - THP1 whole cell protein was separated by SDS-PAGE on a 12% gel and transferred to PVDF membrane. The membrane was probed with anti-CD63 antibody at 2 ug/mL and detected with an anti-mouse HRP secondary antibody using chemiluminescence.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - The CD63 (H5C6) antibody was tested in HeLa cells at a 1:50 dilution against DyLight 488 (Green). Actin and nuclei were counterstained against Phalloidin 568 (Red) and DAPI (Blue), respectively.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - HeLa cells were fixed in 4% paraformaldehyde for 10 minutes and permeabilized in 0.05% Triton X-100 in PBS for 5 minutes. The cells were incubated with anti-CD63 Antibody [H5C6] conjugated to DyLight 550 (NBP2-42225R) at 5 ug/ml for 1 hour at room temperature. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 100X objective and digitally deconvolved.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on SK-MEL-28 cells with CD63 (H5C6) Antibody NBP2-42225R (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 5 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to DyLight 550.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - MDCK cells stained with CD63 antibody at a dilution of 1:50 followed by Donkey anti-mouse secondary antibody conjugated with Alexa Fluor 488 (1:500). Nuclei were stained with Hoechst 33342. ICC/IF image submitted by a verified customer review.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - U2OS cells were fixed in 4% paraformaldehyde for 10 minutes and permeabilized in 0.05% Triton X-100 in PBS for 5 minutes. The cells were incubated with anti-CD63 Antibody [H5C6] conjugated to Alexa Fluor 488 (NBP2-42225AF488) at 5 ug/ml for 1 hour at room temperature. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 100X objective and digitally deconvolved.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - A431 cells were fixed in 4% paraformaldehyde for 10 minutes and permeabilized in 0.05% Triton X-100 in PBS for 5 minutes. The cells were incubated with anti-CD63 Antibody [H5C6] conjugated to Biotin (NBP2-42225B) at 5 ug/ml for 60 minutes at room temperature and detected with Streptavidin Protein conjugated to DyLight 488 at a 1:500 dilution for 60 minutes. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 100X objective and digitally deconvolved.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - HeLa cells were fixed in 4% paraformaldehyde for 10 minutes and permeabilized in 0.05% Triton X-100 in PBS for 5 minutes. The cells were incubated with anti-CD63 Antibody [H5C6] conjugated to Biotin (NBP2-42225B) at 5 ug/ml for 1 hour at room temperature and detected with Streptavidin conjugated to DyLight 488. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 100X objective and digitally deconvolved.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - HeLa cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X PBS + 0.05% Triton X-100. The cells were incubated with anti-CD63 [H5C6] conjugated to Alexa Fluor 488 [NBP2-42225AF488] at 10ug/mL for 1 hour at room temperature. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Western Blot: CD63 Antibody (H5C6) - BSA Free [NBP2-42225] - Analysis of common EV markers, where 10 ug of total protein was loaded in each lane. cASCs-EVs expressed CD63 and CD9, while beta actin and lamin A showed lower expression. The displayed data represent at least three repeated experiments with consistent results. Image collected and cropped by CiteAb from the following publication (//pubmed.ncbi.nlm.nih.gov/32040478/) licensed under a CC-BY license.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on HeLa cells with CD63 [H5C6] Antibody NBP2-42225AF594 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 2.5 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to Alexa Fluor 594.

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Western Blot: CD63 Antibody (H5C6) [NBP2-42225] - The same samples and volumes were run under reducing and non-reducing conditions. All procedures were performed in parallel for both conditions. L- ladder +- exosomes. Image visualized with HRP linked to secondary antibody. Western blot image submitted by a verified customer review.

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/Immunofluorescence: CD63 Antibody (H5C6) [NBP2-42225] - HEK cells stained with CD63 antibody at a dilution of 1:50 followed by Donkey anti-mouse secondary antibody conjugated with Alexa Fluor 488 (1:500). Nuclei were stained with Hoechst 33342. ICC/IF image submitted by a verified customer review.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - Human peripheral blood cells were stained (2 x 10^6 cells/mL) using the anti-CD63 antibody (Blue) at a dilution of 1:1000. Signal was detected using a Gt x Ms DyLight 488 Secondary and gated to the monocyte/granulocyte cell populations. Isotype was Mouse IgG1 kappa (orange). Data collected on BD FACS Calibur flow cytometer.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - A431 cells were stained (1 x10^6 cells/mL) using the anti-CD63 antibody at a 1:1000 dilution (blue). Signal was detected with Gt x Ms DyLight 488 secondary. Isotype control (orange).

Flow (Intracellular): CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow (Intracellular): CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on HepG2 cells with CD63 Antibody (H5C6) NBP2-42225AF488 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 10 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to Alexa Fluor 488.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on HeLa cells with CD63 Antibody (H5C6) NBP2-42225APC (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 1 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to Allophycocyanin.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on SK-MEL-28 cells with CD63 Antibody (H5C6) NBP2-42225AF488 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 5 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to Alexa Fluor 488.

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) [NBP2-42225] - An intracellular stain was performed on HeLa cells with CD63 [H5C6] Antibody NBP2-42225AF488 (blue) and a matched isotype control (orange). Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Cells were incubated in an antibody dilution of 5 ug/mL for 30 minutes at room temperature. Both antibodies were conjugated to Alexa Fluor 488.

CD63 (H5C6) in MCF7 Human Cell Line -

CD63 (H5C6) was detected in immersion fixed MCF7 human breast cancer cell line using Mouse anti-CD63 (H5C6) Protein G Purified Monoclonal Antibody conjugated to Alexa Fluor® 488 (Catalog # NBP2-42225AF488) (green) at 5 µg/mL overnight at 4C. Cells were counterstained with DAPI (blue). Cells were imaged using a 100X objective and digitally deconvolved.

CD63 (H5C6) in A431 Human Cell Line -

CD63 (H5C6) was detected in immersion fixed A431 human skin carcinoma cell line using Mouse anti-CD63 (H5C6) Protein-G purified Monoclonal Antibody conjugated to DyLight 550 (Catalog # NBP2-42225R) (red) at 5 µg/mL overnight at 4C. Cells were counterstained with DAPI (blue). Cells were imaged using a 100X objective and digitally deconvolved.

Detection of CD63 (H5C6) in A431 Human Cell Line by Flow Cytometry.

An intracellular stain was performed on A431 human skin carcinoma cell line using Mouse anti- CD63 (H5C6) Protein-G purified Monoclonal Antibody conjugated to DyLight 550 (Catalog # NBP2-42225R, blue histogram) or matched control antibody (orange histogram) at 2.5 µg/mL for 30 minutes at RT.

CD63 (H5C6) in A431 Human Cell Line.

CD63 (H5C6) was detected in immersion fixed A431 human skin carcinoma cell line using Mouse anti-CD63 (H5C6) Protein-G purified Monoclonal Antibody conjugated to Alexa Fluor® 647 (Catalog # NBP2-42225AF647) (light blue) at 2 µg/mL overnight at 4C. Cells were counterstained with DAPI (blue). Cells were imaged using a 100X objective and digitally deconvolved.

Detection of CD63 (H5C6) in A431 Human Cell Line by Flow Cytometry.

An intracellular stain was performed on A431 human skin carcinoma cell line using Mouse anti- CD63 (H5C6) Protein-G purified Monoclonal Antibody conjugated to Alexa Fluor® 647 (Catalog # NBP2-42225AF647, blue histogram) or matched control antibody (orange histogram) at 2.5 µg/mL for 30 minutes at RT.

Western Blot: CD63 Antibody (H5C6) - BSA Free [NBP2-42225] -

Western Blot: CD63 Antibody (H5C6) - BSA Free [NBP2-42225] - NCAM as a neuronal exosome marker in plasma. (A–C) Quantitative analysis of plasma EVs using flow cytometry. Dot plots of fluorescent intensity for plasma EVs stained with PE-CY7-labeled CD63 antibody. EVs were unstained (A) & stained with either isotype control (B) or CD63 antibody (C). (D) A18945 iPSC-derived neurons cultured for 6 weeks & immunostaining with mature neuronal markers (DAPI, MAP2, NeuN, & merged image). Images were taken using Zeiss LSM confocal microscope at ×63 magnification. Scale bar, 20 μm. (E) Representative images of cortical organoids at day 90 of differentiation. Scale bar, 1 mm. (F) The organoids express cortical layer marker BRN2 (also called POU3F2). The nuclei were stained with DAPI. Scale bar, 100 μm. (Gi) Markers for proliferating neural progenitors (SOX2) & cortical neuron marker CTIP2 (also known as BCL11B) with nuclei DAPI staining (Gii). (H) Western blot analysis of NCAM & CD63 from EVs released from iPSCs, iPSC-derived cortical neurons, & iPSC-derived brain organoids. EVs were isolated using SEC from the cell culture media of each sample, & equal EV particle numbers (6 × 108) were subject to immunoblotting with NCAM & CD63 antibodies. (I–L) Flow cytometry dot plots of fluorescent intensity for plasma EVs double-stained with DiI & NCAM antibody. EV samples were unstained (I), single stained with DiI (J), & double-stained with DiI & NCAM antibody in the absence (K) & presence (L) of 0.1% Triton X-100. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/35655952), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

View 22 more images

Applications for CD63 Antibody (H5C6) - BSA Free

Application

Recommended Usage

Block/Neutralize

reported in scientific literature (PMID 35602933)

Electron Microscopy

reported in scientific literature (PMID 16735575)

Flow Cytometry

1:1000

Functional

reported in scientific literature (PMID 9811687)

Immunocytochemistry/ Immunofluorescence

1:50-1:100

Immunohistochemistry Whole-Mount

reported in scientific literature (PMID 21464080)

In vitro assay

reported in scientific literature (PMID 21464080)

Please Note: Optimal dilutions of this antibody should be experimentally determined.

Reviewed Applications

Read 6 reviews rated 3.7 using NBP2-42225 in the following applications:

Dot Blot (1 Review)
Immunocytochemistry (1 Review)
Immunocytochemistry/Immunofluorescence (2 Reviews)
Western Blot (2 Reviews)

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.05% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: CD63

CD63 (cluster of differentiation 63), also known as lysosome associated membrane protein 3 (LAMP-3), is a membrane spanning glycoprotein with a molecular mass ranging from 30-60 kDa that was the first characterized member of the tetraspanin superfamily (1,2). CD63 is ubiquitously expressed and largely present on the cell surface in the endosomal system (1-3). More specifically, it is generally present in multivesicular bodies (MVBs), also called late endosomes, and lysosomes (1). The CD63 molecule is a total of 238 amino acids (aa), has four hydrophobic membrane-spanning regions and three N-glycosylation sites, and the encoded protein has a theoretical molecular weight of 25 kDa (2,3). CD63 both directly and indirectly interacts with other proteins including integrins, cell surface receptors, other tetraspanins, kinases, and adapter proteins, to name a few (1). CD63 is involved in many cell processes including cell survival and activation, cell adhesion, invasion, and migration (1). Additionally, CD63 has been shown to interact with tissue inhibitor of metalloproteinase 1 (TIMP-1), which originally thought to be an inhibitor of cancer progression has recently been shown to have cancer promoting properties as well (1,4). The CD63-TIMP-1 interaction has been shown to activate the PI3K/AKT pathway in lung adenocarcinoma cells and also promote survival and invasion of acute myeloid leukemia cells (1).CD63 is a useful flow cytometry marker for basophil granulocytes and can be used for the basophil activation test (BAT) to assess IgE-mediated allergy response (5). As CD63 was initially discovered on activated blood platelets and is a lysosomal-associated protein it makes sense it would be involved in platelet or lysosomal-related disorders (1,2). More precisely, CD63 is associated with Hermansky-Pudlak syndrome, a disease characterized by albinism and platelet storage pool deficiency (6).

References

1. Pols, M. S., & Klumperman, J. (2009). Trafficking and function of the tetraspanin CD63. Experimental cell research. https://doi.org/10.1016/j.yexcr.2008.09.020

2. Metzelaar, M. J., Wijngaard, P. L., Peters, P. J., Sixma, J. J., Nieuwenhuis, H. K., & Clevers, H. C. (1991). CD63 antigen. A novel lysosomal membrane glycoprotein, cloned by a screening procedure for intracellular antigens in eukaryotic cells. The Journal of biological chemistry.

3. Horejsi, V., & Vlcek, C. (1991). Novel structurally distinct family of leucocyte surface glycoproteins including CD9, CD37, CD53 and CD63. FEBS letters. https://doi.org/10.1016/0014-5793(91)80988-f

4. Eckfeld, C., HauBler, D., Schoeps, B., Hermann, C. D., & Kruger, A. (2019). Functional disparities within the TIMP family in cancer: hints from molecular divergence. Cancer metastasis reviews. https://doi.org/10.1007/s10555-019-09812-6

5. Hoffmann, H. J., Santos, A. F., Mayorga, C., Nopp, A., Eberlein, B., Ferrer, M., Rouzaire, P., Ebo, D. G., Sabato, V., Sanz, M. L., Pecaric-Petkovic, T., Patil, S. U., Hausmann, O. V., Shreffler, W. G., Korosec, P., & Knol, E. F. (2015). The clinical utility of basophil activation testing in diagnosis and monitoring of allergic disease. Allergy. https://doi.org/10.1111/all.12698

6. Dell'Angelica, E. C., Shotelersuk, V., Aguilar, R. C., Gahl, W. A., & Bonifacino, J. S. (1999). Altered trafficking of lysosomal proteins in Hermansky-Pudlak syndrome due to mutations in the beta 3A subunit of the AP-3 adaptor. Molecular cell. https://doi.org/10.1016/s1097-2765(00)80170-7

Alternate Names

CD63, Granulophysin, Lamp-3, ME491, OMA81H, Tspan30

Entrez Gene IDs

967 (Human)

Gene Symbol

CD63

UniProt

P08962

Additional CD63 Products

Product Documents for CD63 Antibody (H5C6) - BSA Free

Product Datasheets

Download Product Datasheets

COA

SDS

Download SDS

Product Specific Notices for CD63 Antibody (H5C6) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations
Reviews

CD63 Antibody (H5C6) - BSA Free

Novus Biologicals, part of Bio-Techne | Catalog # NBP2-42225

Key Product Details

Species Reactivity

Validated:

Cited:

Applications

Validated:

Cited:

Label

Antibody Source

Format

Concentration

View all CD63 Antibodies »

Product Specifications

Immunogen

Marker

Clonality

Host

Isotype

Scientific Data Images for CD63 Antibody (H5C6) - BSA Free

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Western Blot: CD63 Antibody (H5C6)BSA Free [NBP2-42225]

Immunocytochemistry/ Immunofluorescence: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow (Intracellular): CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

Flow Cytometry: CD63 Antibody (H5C6) - BSA Free [NBP2-42225]

CD63 (H5C6) in MCF7 Human Cell Line -

CD63 (H5C6) in A431 Human Cell Line -

Detection of CD63 (H5C6) in A431 Human Cell Line by Flow Cytometry.

CD63 (H5C6) in A431 Human Cell Line.

Detection of CD63 (H5C6) in A431 Human Cell Line by Flow Cytometry.

Western Blot: CD63 Antibody (H5C6) - BSA Free [NBP2-42225] -

Applications for CD63 Antibody (H5C6) - BSA Free

Block/Neutralize

Electron Microscopy

Flow Cytometry

Functional

Immunocytochemistry/ Immunofluorescence

Immunohistochemistry Whole-Mount

In vitro assay

Reviewed Applications

Formulation, Preparation, and Storage

Purification

Formulation

Format

Preservative

Concentration

Shipping

Stability & Storage

Background: CD63

Alternate Names

Entrez Gene IDs

Gene Symbol

UniProt

Additional CD63 Products

Product Documents for CD63 Antibody (H5C6) - BSA Free

Product Datasheets

COA

Certificate of Analysis

SDS

Product Specific Notices for CD63 Antibody (H5C6) - BSA Free