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Application Note for CD63 monoclonal antibody (clone: 8A12)

Immunofluorescence staining of exosomes that are produced by cancer cell lines using CD63 monoclonal antibody (clone: 8A12)

Reported by Dr. Nobuyoshi Kosaka
Division of Molecular and Cellular Medicine, National Cancer Center Research Institute
Department of Translational Research for Extracellular Vesicles, Tokyo Medical University


Products

  • Anti CD63 for Exosome Isolation, Human (Mouse) Unlabeled, 8A12(Cat.#SHI-EXO-M02)

These products are the antibodies which can specifically detect CD9, CD63 and CD81 known as exosome markers.
These antibodies are suitable for isolation of exosome by immunoprecipitation.


Experimental Details

 Exosomes are membrane vesicles with a diameter of around 100nm. They are vesicle granules with a lipid bilayer membrane, and are released from cells. The ISEV (International Society for Extracellular Vesicles) has recommended using a term "Extracellular Vesicles" for various secretory granules. Among the various secretory granules, exosomes are defined as the one derived from the endosomes. Recent flourishing of exosome studies lead to a further understanding of exosome roles in various diseases.
However, 1) to understand physiological function of exosomes, and 2) to achieve treatment of diseases by targeting exosomes, it is necessary to understand the mechanisms of their generation. Since cancer cells secrete more exosomes than any other cells, there is a possibility that we can find novel cancer therapy by understanding their generation mechanisms. For this purpose, immunostaining of the CD63, one of the well known exosome marker proteins, was carried out to trace cellular dynamics of the exosome production.

Although several antibodies against human CD63 are commercially available, we've chosen to use Anti CD63, Human Unlabeled, 8A12(Cat. No. CAC-SHI-EXO-M02) from Cosmo Bio. Ltd.

As shown in Figs. 1 and 2, the CD63-positive glanular structures were observed in both the prostate cancer cell line PC-3M (Fig.1) and the colon cancer cell line HCT116 (Fig.2). We've used only human anti-CD63 antibody in this experiment, but still we found that cancer cell lines are rich in CD63 positive granules. As for the future work, the co-staining of the cancer cell lines with anti-CD63 antibody and other endosome markers will be performed to elucidate the genes that are involved in regulation of exosome production in the cancer cell lines.


Fig. 1 The localization of CD63 in the prostate cancer cell line PC-3M

Alexa Fluor® 488 labeled anti-mouse antibody was used as secondary antibody.


Fig.2 The localization of CD63 in the colon cancer cell line HCT116

Alexa Fluor® 488 labeled anti-mouse antibody was used as secondary antibody.



This product, anti-CD63 antibody (8A12; Cat. No. SHI-EXO-M02), is highly specific against human CD63, and had been shown to successfully suppress cancer metastasis through exosomes. Furthermore, this can be used in western blot analysis, immunoelectron microscopy, and ELISA. In this report, we showed that this product can also be used in immunostaining. Altogether, the anti-CD63 antibody (8A12) is a versatile antibody in exosome study.

Product List

Product Name Cat# Quantity Price

Anti CD63

CAC-SHI-EXO-M02 100UL

¥ 65,000