In-capillary probing of quantum dots and fluorescent protein self-assembly and displacement using Förster resonance energy transfer.
Herein, a Förster resonance energy transfer system was designed, which consisted of CdSe/ZnS quantum dots donor and mCherry fluorescent protein acceptor. The quantum dots and the mCherry proteins were conjugated to permit Förster resonance energy transfer. Capillary electrophoresis with fluorescence detection was used for the analyses for the described system. The quantum dots and mCherry were sequentially injected into the capillary, while the real-time fluorescence signal of donor and acceptor was simultaneously monitored by two channels with fixed wavelength detectors. An effective separation of complexes from free donor and acceptor was achieved. Results showed quantum dots and hexahistidine tagged mCherry had high affinity and the assembly was affected by His6 -mCherry/quantum dot molar ratio. The kinetics of the self-assembly was calculated using the Hill equation. The microscopic dissociation constant values for out of- and in-capillary assays were 10.49 and 23.39 μM, respectively. The capillary electrophoresis with fluorescence detection that monitored ligands competition assay further delineated the different binding capacities of histidine containing peptide ligands for binding sites on quantum dots. This work demonstrated a novel approach for the improvement of Förster resonance energy transfer for higher efficiency, increased sensitivity, intuitionistic observation, and low sample requirements of the in-capillary probing system.
['Wang|Jianhao|J|', 'Fan|Jie|J|', 'Li|Jinchen|J|', 'Liu|Li|L|', 'Wang|Jianpeng|J|', 'Jiang|Pengju|P|', 'Liu|Xiaoqian|X|', 'Qiu|Lin|L|']
D002623:Chemistry Techniques, Analytical D019075:Electrophoresis, Capillary D031541:Fluorescence Resonance Energy Transfer D008024:Ligands D008164:Luminescent Proteins D010455:Peptides D011485:Protein Binding D045663:Quantum Dots
quantum dots fluorescent protein Förster resonance energy transfer capillary electrophoresis assembly