Fluorescent Adrenergic Receptor Beta 2 (ADRB2) Internalization Assay Cell Line

Applications: Cell Screening
Categories: Cells

A novel CHOK1/tGFP-ADRB2 cell line has been developed through stable transfection to modulate the internalization of the adrenergic receptor beta 2 (ADRB2) complex in living cells and to assay for compounds inducing tGFP-ADRB2 distribution inside the cells. Fluorescent ADRB2 Internalization Assay cell line was obtained by transfection of an expression vector for a fusion protein of tGFP and human ADRB2, as well as G418 into CHOK1 cells. Single cells with strong green fluorescence were selected by flow cytometry, and allowed to expand. These cells constitutively express the tGFP-ADRB2 fusion protein.

Assay Details: Innoprot ADRB2 Internalizacion Cell Line has been designed to assay compounds or analyze stimuli for their ability to modulate adrenergic receptor beta 2 internalization process following and quantifying the fluorescence distribution inside the cells.

This highly reproducible assay allows monitoring ADRB2 receptor internalization process in High Content Analysis and fluorescence microscope applications.

Fig 1: Our expression plasmid containing the codingsequence of human Adrenergic receptor beta 2tagged in the C-terminal with tGFP protein. Our plasmid was transfected in CHO-K1 cells.Resistant clones were obtained by limitdilution, and receptor gene expression wastested by RT-PCR.

Fig 2: Redistribution of beta-2-adrenergicreceptor receptor stimulated with isoproterenol. Cells were treated with 100 ?M Isoproterenol for 7h.Activation and redistribution processes were detected andanalyzed using BD Pathway 855 High-ContentBioimager from BD Biosciences.

Fig 3: Isoproterenol concentrations response in thebeta-2-adrenergic receptor redistribution assay. Cells were treated with 6 log dilution series (n=8). The Ec50for the Isoproterenol was approx 4 ?M after a treatment of 7hwith agonist. Cells were fixed and the nucleus were stainedwith DAPI. Percent Activity was calculated relative to positive(100 uM). The internalization assay was validated withan average Z prime of 0.75+/- 0.01 for High ContentScreening.