期刊
MICROCHIMICA ACTA
卷 186, 期 8, 页码 -出版社
SPRINGER WIEN
DOI: 10.1007/s00604-019-3594-3
关键词
Catalyzed hairpin assembly; Multiplexed detection assay; Aptamer functionalized magnetic beads; Biomarkers; Microfluidic chip electrophoresis
资金
- National Natural Science Foundation of China [31871825]
- Zhejiang Natural Science Foundation [LY19B050001, LY17C200007]
- Ningbo Science & Technology Project, China [2015C50059]
- K.C. Wong Magna Fund in Ningbo University
- Ningbo Natural Science Foundation [LY19B050001, LY17C200007]
An aptamer based assay is presented that is making use of a catalytic hybrid assembly and a microfluidic chip electrophoresis format. It enables simultaneous determination of thebiomarkers (BMs) alpha-fetoprotein (AFP), carbohydrate antigen 125 (CA125), andcarcinoembryonic antigen (CEA). The respective aptamers were covalently bound to Fe3O4@AuNPs (AuMPs) magnetic beads and then used to capture the biomarkers on their surface. Different single-stranded DNA primers were then labeled with various antibodies as encoding and signaling tags. The signal tags reacted with AuMPs-BMs to form different antibody-BM-aptamer complexes. After magnetic separation, three pairs of hairpins as substrates were introduced to trigger catalytic hybrid assembly by the primers in the complex. This will form many duplex DNA products of different length in the supernatant. The products can be magnetically separated by microfluidic chip electrophoresis and determined by fluorometry at excitation/emission wavelengths of 495/525nm. Several experimental conditions including the hairpin concentration, reaction time and temperature were systemically optimized. The method can simultaneously quantify AFP, CEA and CA125, respectively, with detection limits of 0.1, 0.2, 0.15 pg mL(-1) (at S/N = 3). The aptamer functionalized magnetic beads can be reused for at least 20 times with a recovery of up to 80% after heat treatment. The method was employed to simultaneously detect the three BMs in serum samples.
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