DEVELOPMENT OF A PROTEASETAG® LATERAL FLOW DEVICE FOR DETECTION OF ACTIVE NEUTROPHIL ELASTASE IN THE CLINIC

Darragh McCafferty, Charlene Robb, Timothy Ferguson, Kelly Moffitt, Brian Walker, Lorraine Martin

Research output: Contribution to journalMeeting abstractpeer-review

7 Citations (Scopus)

Abstract

Introduction: Neutrophil elastase (NE) levels are frequently elevated in cystic fibrosis (CF) sputum and correlate with inflammatory markers and measurements of lung impairment such as decreased FEV1. NE is there-fore considered a potential biomarker of exacerbation however no tools are available to monitor protease levels at home or in the clinic. The routine monitoring/measurement of NE in sputum could assist the proactive man-agement of CF patients by identifying early signs of exacerbation which could facilitate pre-emptive medical intervention to avoid potentially seri-ous complications. ProAxsis has developed an activity-based immunoassay (ABI) for NE quantification that incorporates a ProteaseTag® molecule capable of the rapid and selective binding of active NE. The translation of this ProteaseTag® technology to a point of care (POC) test, in the form of a lateral flow device (LFD), should provide a valuable tool to monitor disease activity.Aim: To develop and validate a prototype LFD that can rapidly detect active NE in airways samples.Methods: Experiments were performed using a prototype LFD, com-posed of conjugate pad containing the detector component; with appropriate test and control reagents striped onto nitrocellulose membrane. CF Sol/NE-Tag/running buffer were transferred sequentially to the reaction tube (total volume 100 μL), and left to stand for 15 minutes; after which sig-nal intensity was scored between 0-10. Other closely related proteases ie, cathepsin G (CATG), human airways trypsin (HAT), were also examined to evaluate assay specificity.Results: Following initial quantification using the ABI, ProteaseTag® Active NE Immunoassay (ProAxsis Ltd), a subset of CF sol samples con-taining differing levels of active NE, were selected for inclusion in the study. LFD analysis revealed an excellent correlation between these two assay platforms; with a signal for NE, detectable in less than 60 seconds. The assay was also found to be highly sensitive with a positive signal noted for biological samples containing ~90 ng/mL of active NE. Specificity anal-ysis revealed negligible cross-reactivity upon testing with proteases such as CATG and HAT.Conclusion: The development of a highly sensitive and specific LFD for rapid detection of active NE in complex clinical samples could prove to be a vital tool in CF management; on account of this device’s capacity to measure NE, an established biomarker of infection. It is anticipated that a positive test would prompt such individuals to seek medical attention, to mitigate their risk of exacerbation leading to hospitalisation.
Original languageEnglish
Pages (from-to)280
Number of pages1
JournalPediatric Pulmonology
Volume51
Issue numberS45
Publication statusPublished - Oct 2016
EventNorth American Cystic Fibrosis Conference - Orlando, United States
Duration: 25 Oct 201629 Oct 2016

Keywords

  • point of care test
  • cystic fibrosis
  • neutrophil elastase
  • biomarker

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