An embedded microretroreflector-based microfluidic immunoassay platform

Abstract

We present a microfluidic immunoassay platform based on the use of linear microretroreflectors embedded in a transparent polymer layer as an optical sensing surface, and micron-sized magnetic particles as light-blocking labels. Retroreflectors return light directly to its source and are highly detectable using inexpensive optics. The analyte is immuno-magnetically pre-concentrated from a sample and then captured on an antibody-modified microfluidic substrate comprised of embedded microretroreflectors, thereby blocking reflected light. Fluidic force discrimination is used to increase specificity of the assay, following which a difference imaging algorithm that can see single 3 μm magnetic particles without optical calibration is used to detect and quantify signal intensity from each sub-array of retroreflectors. We demonstrate the utility of embedded microretroreflectors as a new sensing modality through a proof-of-concept immunoassay for a small, obligate intracellular bacterial pathogen, Rickettsia conorii, the causative agent of Mediterranean Spotted Fever. The combination of large sensing area, optimized surface chemistry and microfluidic protocols, automated image capture and analysis, and high sensitivity of the difference imaging results in a sensitive immunoassay with a limit of detection of roughly 4000 R. conorii per mL.

Salmonella Typhimurium Sensing Strategy Based on the Loop-Mediated Isothermal Amplification Using Retroreflective Janus Particle as a Nonspectroscopic Signaling Probe

“…This phenomenon can be easily induced by the various light source, such as nonmonochromatic light, and the reflected light has a significantly bright intensity. Based on these features, a number of studies have been introduced that apply the retroreflection principle to the development of biosensors, such as retroreflective Janus particle, , retroreflector cube, and photolithographically fabricated linear microreflector . In particular, RJPs can be observed with simple optical instruments such as a white LED and a CMOS camera; the newly developed platform can measure the concentration of the target gene by simply counting the number of observed RJPs.…”

A low-cost smartphone-based platform for highly sensitive point-of-care testing with persistent luminescent phosphors

“…The phosphors were then washed 3 times in pure anhydrous ethanol, then dried for 3 hours under reduced pressure and elevated temperature (≈37°C) with the SpeedVac concentrator. Monoclonal mouse anti-β hCG antibodies (Arista Biologicals, Item # ABBCG-0402) were oxidized to introduce aldehydes 49,50 for coupling by first transferring the antibodies into pH 5.4 sodium acetate buffer with a 7k Zeba Spin Desalting Column (ThermoFisher, Catalog # 89882). A 100 mM stock solution of sodium periodate was prepared in sodium acetate buffer, then added to the buffer-exchanged antibodies so that the final sodium periodate concentration was 10 mM with an IgG concentration of ≈1 mg/mL.…”