Development Of Immunoassay On Paper-Based Microfluidic Devices For Determination Of Ractopamine, Alpha-Fetoprotein, Human Thyroid Stimulating Hormone

Abstract

In this dissertation, immunoassays on paper-based microfluidic devices have been developed using different types of label for various applications in agriculture safety and medical diagnosis, which can be divided into three parts. In the first part, the conjugation method of ractopamine (RAC) and protein carrier is developed for fabricating paper-based lateral flow strip test of RAC using gold nanoparticles (AuNPs) as a label. The developed conjugation method provided a simple and one-step method with the use of less chemicals. The strip test for RAC determination in animal feed offers a linearity of 0.075-0.750 ng g-1 and a limit of detection (LOD) of 0.1 ng g-1. In the second part, a wax-printed paper-based lateral flow device using enzyme label is developed to determine alpha-fetoprotein (AFP) in human serum by one-step sample loading. The proposed devices under the optimal conditions can be used for the AFP determination with the LOD of 1 ng mL-1 and a linearity of 1-100 ng mL-1. In the third part, the lateral flow immunoassay of human thyroid stimulating hormone (hTSH) is developed using a novel fluorescent label. The hybrid nanocomposite of the AuNPs and fluorophores of europium (III) doped silica (AuNPs@SiO2-Eu3+) was synthesized and used as the novel fluorescent label with dual signal readout through a colorimetric and a fluorescent signals. The LOD of 0.1 µIU mL-1 of the fluorescent signal with a linear range of 0.05-50 µIU mL-1 provided 50-fold lower than the LOD of colorimetric signal (5 µIU mL-1), which could be used for screening both hypothyroidism and hyperthyroidism in human serum. The above results demonstrate that the proposed devices using different types of label provide simple and rapid analysis, portability, high sensitivity and specificity, precision, reproducibility and low cost.