OPTIMIZATION OF THREE-CUBE FRET ASSAY AND APPLICATION IN THE ANALYSIS OF SUBUNIT ASSEMBLING OF iGluR RECEPTORS IN LIVING CELLS
- VernacularTitle:三通道荧光共振能量转移分析法的优化及其对活细胞内受体复合物组成的分析
- Author:
Minzhen WANG
;
Shuang QIU
;
Yulin HUA
;
Fan YANG
;
Yi SHEN
;
Jianhong LUO
- Publication Type:Journal Article
- Keywords:
Fluorescence resonance energy transfer (FRET);
Cyan fluorescent protein (CFP);
Yellow fluorescence protein (YFP);
Ionotropic glutamate receptor (iGluR);
Protein interaction
- From:
Acta Anatomica Sinica
1957;0(04):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish and optimize three-cube FRET assay in living cells and analyze subunit assembling of iGluR receptors. Methods Taking HEK293 cells cotransfed with pECFP and pEYFP as negative control, and those transfected with pECFP-YFP as positive control,different calculation methods using fluorescence microscopy were compared. Results These calculation methods were all suitable for FRET measurement in the system. but the measurement results were affected by the ratio of Donor/Acceptor (D/A) in some degree,and different calculation methods have different optimized conditions. FRET measurement using FR value showed subunit specific assembly of iGluR subtypes.Conclusion There are different optimized conditions for these different calculation methods in the three-cube FRET measurement system,and a further evidence is provided for subunit specific assembling of iGluR subtypes from the FRET assay.
