Inkjet Printing-Based Immobilization Method for a Single-Step and Homogeneous Competitive Immunoassay in Microchannel Arrays

Abstract

In this study, we report an inkjet printing-based method for the immobilization of different reactive analytical reagents on a single microchannel for a single-step and homogeneous solution-based competitive immunoassay. The immunoassay microdevice is composed of a poly(dimethylsiloxane) microchannel that is patterned using inkjet printing by two types of reactive reagents as dissolvable spots, namely, antibody-immobilized graphene oxide and a fluorescently labeled antigen. Since nanoliter-sized droplets of the reagents could be accurately and position-selectively spotted on the microchannel, different reactive reagents were simultaneously immobilized onto the same microchannel, which was difficult to achieve in previously reported capillary-based single-step bioassay devices. In the present study, the positions of the reagent spots and amount of reagent matrix were investigated to demonstrate the stable and reproducible immobilization and a uniform dissolution. Finally, a preliminary application to a single-step immunoassay of C-reactive protein was demonstrated as a proof of concept.

Homogeneous immunoassay utilizing fluorescence resonance energy transfer from quantum dots to tyramide dyes deposited on full immunocomplexes

“…Our approach demonstrated superior sensitivity compared to other approaches (ref. [10][11][12][13][14] and was comparable to the sensitivity of the approach based on signal amplification (ref. 22) (Table 1).…”

“…Immunocomplexes bring the RET donor and acceptor into close proximity, resulting in energy transfer. Various forms of RET have been utilized, including bioluminescence RET, 8,9 FRET, [9][10][11][12][13][14][15][16][17][18] upconversion FRET, [19][20][21] and chemiluminescence RET. 22,23 RET-based homogeneous immunoassays offer userfriendly procedures and hold great potential for routine applications, particularly in on-site testing scenarios.…”

“…To enhance the sensitivity of RETbased homogeneous immunoassays, researchers have made various efforts. Some attempts involved reducing the donoracceptor distance by employing a leucine zipper to bring the donor and acceptor into close proximity [10][11][12][13][14] or utilizing a smaller antibody fragment (4 nm × 2.5 nm × 3 nm) 25 instead of the full antibody. However, these approaches did not significantly improve the LODs and yielded results comparable to those presented in previous studies (Table 1).…”