Homogeneous Immunochemical Assay on the Lateral Flow Strip for Measurement of DNase i Activity

Deoxyribonuclease I (DNase I) is an important enzyme that cleaves both double-stranded and single-stranded DNA at their phosphate backbone. DNase I is a useful biomarker. Previous studies have shown that patients with prostate cancer and systemic lupus erythematosus exhibit reduced DNase I activity, and patients with myocardial infarction exhibit increased DNase I activity. Current methods of measuring DNase I relies either on an immunochemical assay, which requires multiple washing steps, or on a single radial enzyme diffusion assay, which requires a long digestion time and an expensive fluorescence detection system. We have developed a lateral flow immunochemical assay for the measurement of DNase I activity on the test strip. The assay utilized a dually labeled double-stranded DNA as the reporter probe. The biotin-labeled terminal of the probe bound to the streptavidin immobilized on the lateral flow test strip, and the fluorescein-labeled terminal bound to the antibody-conjugated gold nanoparticles, resulting in a visible test line. The presence of DNase I would cleave the reporter probe and lead to reduced test line intensity. Using the DNase I test strip, we have successfully measured the DNase I activity and determined the factors that influence the sensitivity and linear dynamic range of the assay. We have also investigated the conditions that inhibited the DNase I activity. The combined advantage of a wash-free assay format and colorimetric readout would make the lateral flow DNase I test strip a suitable platform for point-of-care diagnostics.