Surface plasmon resonance imaging for affinity analysis of aptamer-protein interactions with PDMS microfluidic chips.
We report on the use of PDMS multichannels for affinity studies of DNA aptamer-human Immunoglobulin E (IgE) interactions by surface plasmon resonance imaging (SPRi). The sensing surface was prepared with thiol-terminated aptamers through a self-assembling process in the PDMS channels defined on a gold substrate. Cysteamine was codeposited with the thiol aptamers to promote proper spatial arrangement of the aptamers and thus maintain their optimal binding efficiencies. Four aptamers with different nucleic acid sequences were studied to test their interaction affinity toward IgE, and the results confirmed that aptamer I (5'-SH-GGG GCA CGT TTA TCC GTC CCT CCT AGT GGC GTG CCC C-3') has the strongest binding affinity. Control experiments were conducted with a PEG-functionalized surface and IgG was used to replace IgE in order to verify the selective binding of aptamer I to the IgE molecules. A linear concentration-dependent relationship between IgE and aptamer I was obtained, and a 2-nM detection limit was achieved. SPRi data were further analyzed by global fitting, and the dissociation constant of aptamer I-IgE complex was found to be 2.7 x 10(-7) M, which agrees relatively well with the values reported in the literature. Aptamer affinity screening by SPR imaging demonstrates marked advantages over competing methods because it does not require labeling, can be used in real-time, and is potentially high-throughput. The ability to provide both qualitative and quantitative results on a multichannel chip further establishes SPRi as a powerful tool for the study of biological interactions in a multiplexed format.
Department of Chemistry, University of California, Riverside, CA 92521, USA.
Zhuangzhi
Z
Wang
Thomas
T
Wilkop
Danke
D
Xu
Yi
Y
Dong
Guangyu
G
Ma
Quan
Q
Cheng
D052157:Aptamers, Nucleotide / Q000032:analysis* / Q000378:metabolism; D001483:Base Sequence; D003543:Cysteamine / Q000032:analysis / Q000378:metabolism; D007073:Immunoglobulin E / Q000032:analysis / Q000378:metabolism; D007074:Immunoglobulin G / Q000032:analysis / Q000378:metabolism; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D011506:Proteins / Q000032:analysis* / Q000378:metabolism; D012680:Sensitivity and Specificity; D013438:Sulfhydryl Compounds / Q000032:analysis / Q000378:metabolism; D020349:Surface Plasmon Resonance / Q000379:methods*
D052157:Aptamers, Nucleotide; D007074:Immunoglobulin G; D011506:Proteins; D013438:Sulfhydryl Compounds; D007073:Immunoglobulin E; D003543:Cysteamine