DNA encapsulating liposome based rolling circle amplification immunoassay as a versatile platform for ultrasensitive detection of protein.
A novel rolling circle amplification (RCA) immunoassay based on DNA-encapsulating liposomes, liposome-RCA immunoassay, was developed for ultrasensitive protein detection. This technique utilized antibody-modified liposomes with DNA prime probes encapsulated as the detection reagent in the sandwiched immunoassays. The DNA prime probes were released from liposomes and then initiated a linear RCA reaction, generating a long tandem repeated sequences that could be selectively and sensitively detected by a microbead-based fluorescence assay. The developed technique offered very high sensitivity due to primary amplification via releasing numerous DNA primers from a liposome followed by a secondary RCA amplification. A biobarcode design was incorporated in the technique, which allowed the strategy to be directly implemented for multiplex assay of multiple proteins. Also, the technique allowed easy preparation of the DNA-carrying antibody reagent and the implementation with simple instrumentation. The technique was demonstrated for the determination of prostate-specific antigen (PSA), a highly selective biomarker associated with prostate cancer. The results revealed that the technique exhibited a dynamic response to PSA over a 6-decade concentration range from 0.1 fg mL(-1) to 0.1 ng mL(-1) with a limit of detection as low as 0.08 fg mL(-1) and a high dose-response sensitivity. The liposome-RCA immunoassay holds great promise as a versatile, sensitive, and robust platform to combine the nucleic acid amplification with immunoassay for ultrasensitive protein detection.
Ou
Li-Juan
LJ
State Key Laboratory of Chemo/Bio-Sensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, P. R. China.
Liu
Si-Jia
SJ
Chu
Xia
X
Shen
Guo-Li
GL
Yu
Ru-Qin
RQ
D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000378:metabolism; D001483:Base Sequence; D002417:Cattle; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007118:Immunoassay / Q000191:economics / Q000379:methods*; D008081:Liposomes / Q000737:chemistry*; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D011506:Proteins / Q000032:analysis* / Q000276:immunology
D000906:Antibodies; D015342:DNA Probes; D008081:Liposomes; D011506:Proteins