Multiplexed enrichment and detection of malarial biomarkers using a stimuli-responsive iron oxide and gold nanoparticle reagent system.
There is a need for simple yet robust biomarker and antigen purification and enrichment strategies that are compatible with current rapid diagnostic modalities. Here, a stimuli-responsive nanoparticle system is presented for multiplexed magneto-enrichment and non-instrumented lateral flow strip detection of model antigens from spiked pooled plasma. The integrated reagent system allows purification and enrichment of the gold-labeled biomarker half-sandwich that can be applied directly to lateral flow test strips. A linear diblock copolymer with a thermally responsive poly(N-isopropylacrylamide) (pNIPAm) segment and a gold-binding block composed of NIPAm-co-N,N-dimethylaminoethylacrylamide was prepared by reversible addition-fragmentation chain transfer polymerization. The diblock copolymer was used to functionalize gold nanoparticles (AuNPs), with subsequent bioconjugation to yield thermally responsive pNIPAm-AuNPs that were co-decorated with streptavidin. These AuNPs efficiently complexed biotinylated capture antibody reagents that were bound to picomolar quantities of pan-aldolase and Plasmodium falciparum histidine-rich protein 2 (PfHRP2) in spiked pooled plasma samples. The gold-labeled biomarker half-sandwich was then purified and enriched using 10 nm thermally responsive magnetic nanoparticles that were similarly decorated with pNIPAm. When a thermal stimulus was applied in conjunction with a magnetic field, coaggregation of the AuNP half-sandwiches with the pNIPAm-coated iron oxide nanoparticles created large aggregates that were efficiently magnetophoresed and separated from bulk serum. The purified biomarkers from a spiked pooled plasma sample could be concentrated 50-fold into a small volume and applied directly to a commercial multiplexed lateral flow strip to dramatically improve the signal-to-noise ratio and test sensitivity.
Department of Bioengineering, University of Washington, Seattle, Washington 98195, USA.
Michael A
MA
Nash
John N
JN
Waitumbi
Allan S
AS
Hoffman
Paul
P
Yager
Patrick S
PS
Stayton
D000953:Antigens, Protozoan / Q000097:blood* / Q000276:immunology / Q000302:isolation & purification*; D015415:Biomarkers / Q000032:analysis; D015374:Biosensing Techniques / Q000295:instrumentation; D002846:Chromatography, Affinity / Q000295:instrumentation*; D004586:Electrophoresis / Q000295:instrumentation; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006046:Gold / Q000737:chemistry*; D006801:Humans; D018189:Immunomagnetic Separation / Q000295:instrumentation*; D007202:Indicators and Reagents / Q000737:chemistry; D058185:Magnetite Nanoparticles / Q000737:chemistry* / Q000528:radiation effects / Q000648:ultrastructure; D008288:Malaria / Q000097:blood* / Q000276:immunology; D036103:Nanotechnology / Q000295:instrumentation; D015800:Protozoan Proteins / Q000097:blood* / Q000276:immunology / Q000302:isolation & purification*; D011934:Reagent Strips
D000953:Antigens, Protozoan; D015415:Biomarkers; C514457:HRP-2 antigen, Plasmodium falciparum; D007202:Indicators and Reagents; D058185:Magnetite Nanoparticles; D015800:Protozoan Proteins; D011934:Reagent Strips; D006046:Gold