Development of a functional point-of-need diagnostic for myeloperoxidase detection to identify neutrophilic bronchitis

Abstract

With over 4.8 million Canadians suffering from chronic airway diseases, respiratory exacerbations are currently the leading cause of hospitalization in Canada. In cases of bacterial infection, neutrophil cell density increases from ∼10 million cells per gram to over 15 million cells per gram. As sputum is a direct discharge from the primarily affected areas of respiratory diseases, quantification of granulocytes (including neutrophils) can be used to effectively determine a course of patient treatment. Unfortunately this quantification is currently limited to labour-intensive and time-consuming cell counts. In the present study, we describe a simple one-step lateral flow test (LFT) that can semi-quantitatively determine myeloperoxidase (MPO), a biomarker found in neutrophils, in minimally-processed sputum samples. This point-of-need (PON) diagnostic device provides positive results observable to the naked eye after 15 minutes. 37 human sputum samples were quantified for MPO using the developed LFT and compared to neutrophil levels quantified through traditional cell counting. A trend between sputum MPO concentration and total neutrophils was observed, suggesting that the LFT has the potential to replace cell counting for neutrophil approximation to aid in directing therapies quickly at the point of need.

Rapid quantification of sputum eosinophil peroxidase on a lateral flow test strip

“…3 Herein, we describe a rapid LFT for the detection of eosinophil peroxidase (EPX), a biomarker that is eosinophil-specific, and demonstrate its utility for testing of processed patient sputum samples. Our group has recently shown that a rapid, paper-based, lateral flow test (LFT) for myeloperoxidase (MPO) provides a reliable, rapid and potentially cost-effective alternative to cytometry for the identification of neutrophilic bronchitis.…”

Microfluidic and Paper-Based Devices for Disease Detection and Diagnostic Research

“…The most widely used and commercially available LFSA is the home pregnancy test, which identifies hormones in urine. This same concept can be applied for recognizing other biomarkers to diagnose a patient with a disease or illness such as the detection of myeloperoxidase as a neutrophilic bronchitis precursor [ 29 ]. These assays work by taking a liquid patient sample (e.g., blood, urine, or saliva) and applying that sample to an absorbent pad on one side of the strip.…”

“…Many LFSAs utilize antibodies to detect specific biomolecules [ 29 , 31 , 32 , 33 ]. Before applying the sample to the strip, it must be mixed with detection antibodies that bind to the target molecule that is conjugated to a secondary molecule to provide an analytical output for qualitative analysis.…”

“…The second line, or test line, contains immobilized antibodies that bind to the target molecule. Current LFSA results can be interpreted as a fluorescent signal present on the test line [ 32 ], as a visually present test line [ 29 , 30 , 33 ], or by measuring a pressure output from a chemical reaction [ 31 ].…”

“…A recently developed LFSA uses a combination of gold nanoflowers conjugated to antibodies and quantum dot nanobeads to produce a fluorescent signal in the presence of tetrodotoxin (TTX), a potent neurotoxin present within marine life, as a detection method for patients who have potentially consumed contaminated food or to test food sources [ 32 ]. Oliveira-Rodriguez et al used a strip assay by conjugating antibodies to nanoparticles for the quantitative detection of extracellular vesicles [ 33 ], whereas Wolfe et al used a similar approach to detect myeloperoxidase from sputum samples to diagnose neutrophilic bronchitis [ 29 ]. While still providing a visual result, Raston et al developed an LFSA that incorporates aptamers conjugated to nanoparticles to bind to specific target molecules such as Vaspin as a biomarker for diabetes-related diseases [ 30 ].…”