Aptamer-based rolling circle amplification: a platform for electrochemical detection of protein.
Aptamer-based rolling circle amplification (aptamer-RCA) was developed as a novel versatile electrochemical platform for ultrasensitive detection of protein. This method utilized antibodies immobilized on the electrode surface to capture the protein target, and the surface-captured protein was then sandwiched by an aptamer-primer complex. The aptamer-primer sequence mediated an in situ RCA reaction that generated hundreds of copies of a circular DNA template. Detection of the amplified copies via enzymatic silver deposition then allowed enormous sensitivity enhancement in the assay of target protein. This novel aptamer-primer design circumvented time-consuming preparation of the antibody-DNA conjugate for the common immuno-RCA assay. Moreover, the detection strategy based on enzymatic silver deposition enabled a highly efficient readout of the RCA product as compared to a redox-labeled probe based procedure that might exhibit low detection efficiency due to RCA product distance from the electrode. With the platelet-derived growth factor B-chain (PDGF-BB) as a model target, it was demonstrated that the presented method was highly sensitive and specific with a wide detection range of 4 orders of magnitude and a detection limit as low as 10 fM. Because of the wide availability of aptamers for numerous proteins, this platform holds great promise in ultrasensitive immunoassay.
Zhou
Long
L
State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, P. R. China.
Ou
Li-Juan
LJ
Chu
Xia
X
Shen
Guo-Li
GL
Yu
Ru-Qin
RQ
D052157:Aptamers, Nucleotide; D001483:Base Sequence; D000077214:Becaplermin; D017931:DNA Primers; D004563:Electrochemistry / Q000379:methods*; D006801:Humans; D007118:Immunoassay / Q000379:methods; D010982:Platelet-Derived Growth Factor / Q000032:analysis*; D020574:Proto-Oncogene Proteins c-sis; D011994:Recombinant Proteins / Q000032:analysis; D012834:Silver / Q000737:chemistry
D052157:Aptamers, Nucleotide; D017931:DNA Primers; D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; D011994:Recombinant Proteins; D000077214:Becaplermin; D012834:Silver