NOVEL ASSAY DEVELOPMENT METHODS FOR RAPID DETECTION OF GENOME INSTABILITY BIOMARKERS

Abstract

Background: Accumulation of genomic damage in the form of mutation events, chromosomal abnormalities, and DNA damage are all associated with cancer, aging, and different degenerative diseases, and are all significant contributors to genome instability. However, valid genome instability biomarkers must be detected in a timely and accurate fashion to inspire diagnosis, prognosis and precision medicine applications. Traditional diagnostic methods, however, tend to be time consuming, labor intensive and less sensitive.

Objective: Within this study, advanced molecular diagnostic technologies have been used to develop quick and very sensitive assays for detecting genome instability biomarker.

Methods: Novel biosensor platforms designed for detection of genome instability biomarkers such as γ-H2AX, 8-OHdG and micronuclei were designed. Analytical tools utilizing the CRISPR system, fluorescence biosensors and electrochemical detection assays were designed and tested with cell and clinical samples. The detection sensitivity and specificity as well as the reproducibility of the assay were comparatively analyzed.

Findings: The developed assays were highly sensitive with detection limit at 0.5 to 2.0 ng/mL, and had the capability of detecting the biomarkers in 20 to 30 minutes. The average diagnostic accuracy achieved by the CRISPR-based assays was about 95% and 92%, respectively, and the fluorescence biosensors had better signal amplification and rapid response performance. Clinical validation revealed a very consistent detection of biomarkers in oxidative stress and genome instability conditions.

Conclusion: New assay development methods significantly enhance the speed, sensitivity and accuracy of the detection of genome instability biomarkers; these are useful for early disease diagnosis, genomic monitoring, and precision diagnostic applications.