Human paraffin-embedded tonsil tissue slices were prepared with a standard protocol of deparaffinization and rehydration. Antigen retrieval was done with Citrate Buffered 1X (10mM, pH6.0) at 95°C for 40 minutes. Tissue was washed with PBS/0.05% Tween 20 twice for five minutes and blocked with 5% FBS and 0.2% gelatin for 30 minutes. Then, the tissue was stained with 10µg/mL of Alexa Fluor® 647 anti-CD8a (Clone C8/144B) antibody (green) and Alexa Fluor® 594 anti-CD45RA (Clone HI100) antibody (red) over night at 4°C. The image was scanned with a 10X objective and stitched with MetaMorph® software.
Human paraffin-embedded tonsil tissue slices were prepared with a standard protocol of deparaffinization and rehydration. Antigen retrieval was done with Citrate Buffered 1X (10mM, pH6.0) at 95°C for 40 minutes. Tissue was washed with PBS/0.05% Tween 20 twice for five minutes and blocked with 5% FBS and 0.2% gelatin for 30 minutes. Then, the tissue was stained with 10µg/mL of Alexa Fluor® 647 anti-CD8a (Clone C8/144B) antibody (green) and Alexa Fluor® 594 anti-CD45RA (Clone HI100) antibody (red) over night at 4°C. The image was scanned with a 10X objective and stitched with MetaMorph® software.
CD45RA is a 205-220 kD single chain type I glycoprotein. It is an exon 4 splice variant of the tyrosine phosphatase CD45. The CD45RA isoform is expressed on resting/naïve T cells, medullary thymocytes, B cells and monocytes. CD45RA enhances both T cell receptor and B cell receptor signaling. CD45 non-covalently associates with lymphocyte phosphatase-associated phosphoprotein (LPAP) on T and B lymphocytes. CD45 has been reported to be associated with several other cell surface antigens including CD1, CD2, CD3, and CD4. CD45 has also been reported to bind galectin-1. CD45 isoform expression can change in response to cytokines.
Each lot of this antibody is quality control tested by formalin-fixed paraffin-embedded immunohistochemical staining. For immunohistochemistry, a concentration range of 5 - 10 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.
Additional reported applications (for relevant formats of this clone) include: inhibition of CD45 functions2, immunohistochemical staining of frozen tissue sections3 and formalin-fixed paraffin-embedded tissue sections4, and immunocytochemistry15,16.
Application References
(PubMed link indicates BioLegend citation)
Knapp W, et al. 1989. Leucocyte Typing IV. Oxford University Press. New York.
Yamada T, et al. 2002. J. Biol. Chem. 277:28830. (WB, Block)
Weninger W, et al. 2003 J. Immunol. 170:4638. (IHC-F)
Imanguli MM, et al. 2009. Blood. 113:3620 (IHC-P)
Roque S, et al. 2007. J. Immunol. 178:8028. (FC) PubMed
Smeltz RB. 2007. J. Immunol. 178:4786. (FC) PubMed
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