A single-step enzyme immunoassay capillary sensor composed of functional multilayer coatings for the diagnosis of marker proteins

Abstract

A single-step, easy-to-use enzyme immunoassay capillary sensor, composed of functional multilayer coatings, was developed in this study. The coatings were composed of substrate-immobilized hydrophobic coating, hydrogel coating, and soluble coating containing an enzyme-labeled antibody. The response mechanism involved a spontaneous immunoreaction triggered by capillary action-mediated introduction of a sample antigen solution and subsequent separation of unreacted enzyme-labeled antibodies and antigen-enzyme-labeled antibody complexes by the molecular sieving effect of the hydrogel. An enzyme reaction at the substrate-immobilized hydrophobic coating/hydrogel coating interface resulted in a protein-selective fluorescence response. An antigen concentration-dependent response was obtained for diagnostic marker protein samples (hemoglobin A1c (HbA1c), 7.14-16.7 mg mL(-1); alpha-fetoprotein (AFP), 1.4-140 ng mL(-1); C-reactive protein (CRP), 0.5-10 μg mL(-1)) that cover a clinically important concentration range. The successful measurement of CRP in diluted serum samples demonstrated the application of this capillary sensor.

Fabrication and packaging of a mass-producible capillary-assembled microchip for simple and multiplexed bioassay

“…As an initial demonstration, we have successfully attained the multiplexed glycosidase enzyme assay. In addition to its use for enzyme activity assays, the CAs-CHIP-3G microdevice could also be used for other bioassays by integrating other capillary biosensors that we have developed previously for various analytes such as ions [27,28], metabolites [30,31], and proteins [37]. This inexpensive, reliable, and easy-to-use bioanalytical microdevice could be extended for colorimetric assay with the smartphone camera as the detector which has more potential for point-of-care or point-of-use applications.…”

Sensitive “capillary ELISA” via vapor-phase surface modification

“…However, the capillary tube was processed by a complex surface treatment to equip chemically linked biomolecules and physically adsorbed peroxidase in polyethyelene glycol membrane. In another report, LOD for CRP was 0.5 g/mL even with complex surface treatment including multiple coatings composed of soluble, hydrogel and hydrophobic coatings [22]. Using the well-known surface-chemistry based on APTES and GA, a square capillary tube reported to produce LOD of 10 ng/mL [24].…”

“…Also in the POC applications, various versions of capillary tube-based immunoassay systems were developed taking advantage of its simple structural feature [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25]. Rectangular-shaped capillary tubes were used to perform enzyme immunoassay for various types of biomarkers [18,[22][23][24]. When the well-known surface chemistry based on 3-aminoprolytriethoxysilane (APTES) and glutaraldehyde (GA) were used to install antibody in the capillary tube, the immunoassay results produced 10 ng/mL level detection capability [24].…”

A Simple and Rapid Immunoassay Based on Microchip Electrophoresis Using a Reagent-Release Cartridge

“…Our group has developed simple and rapid assays using reagent-release capillaries (RRCs) [18][19][20][21][22][23][24][25][26][27][28][29][30], in which the reagents are adsorbed onto the inner surface of the capillary as a soluble coating. When the sample solution is spontaneously introduced into the capillary by a capillary action, the adsorbed reagents dissolve again and mix with the sample solution to give a fluorescent signal.…”