Rapid quantification of the malaria biomarker hemozoin by improved biocatalytically initiated precipitation atom transfer radical polymerizations

The fight against tropical diseases such as malaria requires the development of innovative biosensing techniques. Diagnostics must be rapid and robust to ensure prompt case management and to avoid further transmission. The malaria biomarker hemozoin can catalyze atom transfer radical polymerizations (ATRP), which we exploit in a polymerization-amplified biosensing assay for hemozoin based on the precipitation polymerization of N-isopropyl acrylamide (NIPAAm). The reaction conditions are systematically investigated using synthetic hemozoin to gain fundamental understanding of the involved reactions and to greatly reduce the amplification time, while maintaining the sensitivity of the assay. The use of excess ascorbate allows oxygen to be consumed in situ but leads to the formation of reactive oxygen species and to the decomposition of the initiator 2-hydroxyethyl 2-bromoisobutyrate (HEBIB). Addition of sodium dodecyl sulfate (SDS) and pyruvate results in better differentiation between the blank and hemozoin-containing samples. Optimized reaction conditions (including reagents, pH, and temperature) reduce the amplification time from 37 +/- 5 min to 3 +/- 0.5 min while maintaining a low limit of detection of 1.06 ng mL(-1). The short amplification time brings the precipitation polymerization assay a step closer to a point-of-care diagnostic device for malaria. Future efforts will be dedicated to the isolation of hemozoin from clinical samples.