A novel microfluidics PMMA/paper hybrid bioimmunosensor for laser-induced fluorescence detection in the determination of alpha-fetoprotein from serum

Low loss and high capture efficiency of target samples are prerequisites for sensitive detection by fluorescence immunosensors. For quantitative and ultrasensitive detection of alpha-fetoprotein (AFP), we designed a novel microfluidics pump-free PMMA/paper hybrid immunosensor. Here, the 2D planar structure of the detection area of the plastic microfluidic chip is improved to a 3D spatial structure, which increases the number of capture antibodies immobilized and the cross-section area of the intercept target protein. PMMA channels efficiently transport target proteins. The paper-based, three-dimensional detection area not only increases the amount of immobilization of the captured antibody but also improves the contact probability between the captured antibody and the target protein. This greatly improves the efficiency of immune binding. A laser-induced fluorescence detector is applied to detect the target protein on the paper-based detecting unit. Under optimal conditions, the immunosensor ensures that the reaction time is 12 min and the residual liquid is only 6.1% of the original volume, achieving ultra-sensitive detection of AFP in the range of 0.01 ng/mL to 100 ng/mL with a limit of quantitation (LOQ) as low as 4.5 pg/mL. The recoveries for AFP ranged from 96.6% to 101.5%, which means the hybrid microfluidics chip is reliable and has great potential for early screening.

Automated summary

We have designed a novel microfluidics pump-free PMMA/paper hybrid immunosensor for quantitative and ultrasensitive detection of alpha-fetoprotein (AFP). By improving the 2D planar structure of the plastic microfluidic chip to a 3D spatial structure, the capture antibody immobilization and the cross-section area of the intercepted target protein are increased. PMMA channels efficiently transport target proteins. The paper-based, three-dimensional detection area increases the amount of immobilized captured antibody and improves the contact probability between the captured antibody and the target protein, greatly improving the efficiency of immune binding. The immunosensor achieves ultra-sensitive detection of AFP in the range of 0.01 ng/mL to 100 ng/mL with a limit of quantitation as low as 4.5 pg/mL, under optimal conditions. The recoveries for AFP range from 96.6% to 101.5%, indicating the reliability and potential of the hybrid microfluidics chip for early screening.