Aptamer/antibody sandwich method for digital detection of SARS-CoV2 nucleocapsid protein

Ge，Chenchen1; Feng，Juan1; Zhang，Jiaming2; Hu，Kai2; Wang，Dou3; Zha，Ling1; Hu，Xuejuan2; Li，Rongsong1

2022

10.1016/j.talanta.2021.122847

TALANTA   影响因子和分区

0039-9140

1873-3573

Nucleocapsid protein (N protein) is the most abundant protein in SARS-CoV2 and is highly conserved, and there are no homologous proteins in the human body, making it an ideal biomarker for the early diagnosis of SARS-CoV2. However, early detection of clinical specimens for SARS-CoV2 remains a challenge due to false-negative results with viral RNA and host antibodies based testing. In this manuscript, a microfluidic chip with femtoliter-sized wells was fabricated for the sensitive digital detection of N protein. Briefly, β-galactosidase (β-Gal)-linked antibody/N protein/aptamer immunocomplexes were formed on magnetic beads (MBs). Afterwards, the MBs and β-Gal substrate fluorescein-di-β-D-galactopyranoside (FDG) were injected into the chip together. Each well of the chip would only hold one MB as confined by the diameter of the wells. The MBs in the wells were sealed by fluorocarbon oil, which confines the fluorescent (FL) product generated from the reaction between β-Gal and FDG in the individual femtoliter-sized well and creates a locally high concentration of the FL product. The FL images of the wells were acquired using a conventional inverted FL microscope. The number of FL wells with MBs (FL wells number) and the number of wells with MBs (MBs wells number) were counted, respectively. The percentage of FL wells was calculated by dividing (FL wells number) by (MBs wells number). The higher the percentage of FL wells, the higher the N protein concentration. The detection limit of this digital method for N protein was 33.28 pg/mL, which was 300 times lower than traditional double-antibody sandwich based enzyme-linked immunosorbent assay (ELISA).

Aptamer/antibody sandwich COVID-19 Digital detection Microfluidic chip Nucleocapsid protein SARS-CoV2

SCI ; EI

英语

通讯

Special anti-COVID-19 Project Fund of Guangdong Provincial Education Department[2020KZDZX1214] ; Natural Science Foundation of Top Talent of SZTU[2020102] ; Shenzhen Natural Science Foundation[JCYJ20190813141001745] ; Special Fund for Science and Technology Innovation of Pingshan District, Shenzhen[PSKG202006]

Chemistry

Chemistry, Analytical

WOS:000697683600004

ELSEVIER

20213710887493

Antibodies ; Chemical detection ; Diagnosis ; Digital microfluidics ; Diseases ; Fluidic devices

Biological Materials and Tissue Engineering:461.2 ; Medicine and Pharmacology:461.6 ; Immunology:461.9.1 ; Hydraulic Equipment and Machinery:632.2 ; Microfluidics:632.5.1 ; Control Equipment:732.1 ; Chemistry:801

CHEMISTRY

2-s2.0-85114618074

Scopus

1.College of Health Science and Environmental Engineering,Shenzhen Technology University,Shenzhen,3002 Lantian Road, Pingshan District,518118,China
2.Sino-German College of Intelligent Manufacturing,Shenzhen Technology University,Shenzhen,3002 Lantian Road, Pingshan District,518118,China
3.Shenzhen Key Laboratory of Smart Healthcare Engineering,Department of Biomedical Engineering,Southern University of Science and Technology,Shenzhen,No. 1088, Xueyuan Rd., Xili, Nanshan District,518055,China

Ge，Chenchen,Feng，Juan,Zhang，Jiaming,et al. Aptamer/antibody sandwich method for digital detection of SARS-CoV2 nucleocapsid protein[J]. TALANTA,2022,236.

Ge，Chenchen.,Feng，Juan.,Zhang，Jiaming.,Hu，Kai.,Wang，Dou.,...&Li，Rongsong.(2022).Aptamer/antibody sandwich method for digital detection of SARS-CoV2 nucleocapsid protein.TALANTA,236.

Ge，Chenchen,et al."Aptamer/antibody sandwich method for digital detection of SARS-CoV2 nucleocapsid protein".TALANTA 236(2022).