title: Enabling a microfluidic immunoassay for the developing world by integration of on-card dry reagent storage.
abstract: As part of an effort to create a point-of-care diagnostic system for the developing world, we present a microfluidic flow-through membrane immunoassay with on-card dry reagent storage. By preserving reagent function, the storage and reconstitution of anhydrous reagents enables the devices to remain viable in challenging, unregulated environmental conditions. The assay takes place on a disposable laminate card containing both a porous membrane patterned with capture molecules and a fibrous pad containing an anhydrous analyte label. To conduct the assay, the card is placed in an external pumping and imaging instrument capable of delivering sample and rehydrated reagent to the assay membrane at controlled flow rates to generate quantitative results. Using the malarial antigen Plasmodium falciparum histidine-rich protein II (PfHRP2) as a model, we demonstrate selection of dry storage conditions, characterization of reagent rehydration, and execution of an automated on-card assay. Gold-antibody conjugates dried in a variety of sugar matrices were shown to retain 80-96% of their activity after 60 days of storage at elevated temperatures, and the release profile of the reconstituted reagent was characterized under flow in microfluidic channels. The system gave a detection limit in the sub-nanomolar range in under nine minutes, showing the potential to expand into quantitative, multi-analyte analysis of human blood samples.
authors.affiliation: Department of Bioengineering, University of Washington, Box 355061, Foege N530J, 1705 NE Pacific Street, Seattle, WA 98195-5061, USA. yasuo@u.washington.edu
authors.forename: Dean Y
authors.initials: DY
authors.lastname: Stevens
authors.forename: Camille R
authors.initials: CR
authors.lastname: Petri
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Osborn
authors.forename: Paolo
authors.initials: P
authors.lastname: Spicar-Mihalic
authors.forename: Katherine G
authors.initials: KG
authors.lastname: McKenzie
authors.forename: Paul
authors.initials: P
authors.lastname: Yager
mesh_terms: D000818:Animals; D003890:Desiccation; D003906:Developing Countries; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods; D007202:Indicators and Reagents / Q000737:chemistry*; D044085:Microfluidics / Q000295:instrumentation / Q000379:methods; D008954:Models, Biological; D010961:Plasmodium
chemical_list: D007202:Indicators and Reagents