Gravity-Driven Microfluidic Siphons: Fluidic Characterization and Application to Quantitative Immunoassays

Abstract

A range of biosensing techniques including immunoassays are routinely used for quantitation of analytes in biological samples and available in a range of formats, from centralized lab testing (e.g., microplate enzyme-linked immunosorbent assay (ELISA)) to automated point-of-care (POC) and lateral flow immunochromatographic tests. High analytical performance is intrinsically linked to the use of a sequence of reagent and washing steps, yet this is extremely challenging to deliver at the POC without a high level of fluidic control involving, e.g., automation, fluidic pumping, or manual fluid handling/pipetting. Here we introduce a microfluidic siphon concept that conceptualizes a multistep ″dipstick″ for quantitative, enzymatically amplified immunoassays using a strip of microporous or microbored material. We demonstrated that gravity-driven siphon flow can be realized in single-bore glass capillaries, a multibored microcapillary film, and a glass fiber porous membrane. In contrast to other POC devices proposed to date, the operation of the siphon is only dependent on the hydrostatic liquid pressure (gravity) and not capillary forces, and the unique stepwise approach to the delivery of the sample and immunoassay reagents results in zero dead volume in the device, no reagent overlap or carryover, and full start/stop fluid control. We demonstrated applications of a 10-bore microfluidic siphon as a portable ELISA system without compromised quantitative capabilities in two global diagnostic applications: (1) a four-plex sandwich ELISA for rapid smartphone dengue serotype identification by serotype-specific dengue virus NS1 antigen detection, relevant for acute dengue fever diagnosis, and (2) quantitation of anti-SARS-CoV-2 IgG and IgM titers in spiked serum samples. Diagnostic siphons provide the opportunity for high-performance immunoassay testing outside sophisticated laboratories, meeting the rapidly changing global clinical and public health needs.

Smartphone multiplex microcapillary diagnostics using Cygnus: Development and evaluation of rapid serotype-specific NS1 detection with dengue patient samples

“…In contrast, previous studies used syringes to aspirate samples/wash/reagents. A full stepwise illustration of operation is provided in S2 Fig , and fluid mechanics described in [ 34 ].…”

“…For simpler assays requiring only a sample plus single reagent, MCF test strips used capillary action achieved by coating the inner surface with hydrophilic polymer [ 33 ]; this hydrophilic polymer coating is compatible with antibody coating for immunoassays [ 31 , 32 , 33 ]. Recently, we further reduced the complexity of MCF immunoassay devices by using gravity to flow the sample followed by wash and detection reagents sequentially through microcapillaries [ 34 ].…”

Merging microfluidics with luminescence immunoassays for urgent point-of-care diagnostics of COVID-19

“…(Reprinted with permission from Ref. [ 152 ]). (C) Schematic diagram of electro-driven ICA integrated with UCNPs.…”

Mechanical Timer-Actuated Fluidic Dispensing System: Applications to an Automated Multistep Lateral Flow Immunoassay with High Sensitivity

“…This, in turn, results in low signal-to-noise ratios due to unbound enzyme/conjugate via excessive washing. Microfluidic capillary systems using trigger valves realize sophisticated fluidic control in capillary networks at the expense of reagent waste and complicated system orchestration and structure. − Despite several studies addressing sequential assay steps via pressing, folding, , sliding, and siphoning mechanisms, these types of approaches continue to require manual operations to introduce reagents in each step. Thus, a simple platform that can sequentially perform multistep assays in a highly sensitive, paper-independent, and automated manner should be designed.…”