Fluorescent Bradikinin Receptor 1 (BDKR1) Internalization Assay Cell Line
A novel CHOK1/tGFP-BDKR1 cell line has been developed through stable transfection to modulate the internalization of the bradykinin receptor 1 (BDKR1) complex in living cells and to a Betaay for compounds inducing tGFP-BDKR1 distribution inside the cells.
Fluorescent BDKR1 Internalization cell line was obtained by transfection of an expre Betaion vector for a fusion protein of tGFP and human BDKR1, 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 expre Beta the tGFP-BDKR1 fusion protein.
A Betaay Details: Innoprot BDKR1 Internalizacion Cell Line has been designed to a Betaay compounds or analyze stimuli for their ability to modulate bradykinin receptor 1 internalization proce Beta following and quantifying the fluorescence distribution inside the cells.
This highly reproducible assay allows monitoring BDKR1 receptor internalization process Beta in High Content Analysis and fluorescence microscope applications.
Fig 1: Our expre Betaion plasmid containing the codingsequence of human Bradikinin receptor 1tagged in the N-terminal with tGFP protein.Our plasmid was transfected in CHO-K1 cells,using calcium phosphate method. Resistantclones were obtained by limit dilution, andreceptor gene expre Betaion was tested by RTPCR.
Fig 2: Internalization of BDK1-tGFP stimulatedwith Bradykinin. Cells were treated with 10uMBradykinin for 5h. Activation and internalization proce Betaeswere detected and analyzed using BD Pathway 855 High-Content Bioimager from BD Biosciences.
Fig 3:BDK1R-tGFP internalization in response toBradykinin concentrations. Cells were treated with 8log dilution series (n=8). The Ec50 for the Bradykinin was 130nM after a treatment of 5h with agonist. Cells werefixed and the nuclei were stained with DAPI. % Activitywas calculated relative to positive (10uM). Theinternalization a Betaay was validated with an average of Z=0.76+/- 0.01 for High Content Screening.