Colorimetric Detection of DNA Strands on Cellulose Microparticles Using ZZ-CBM Fusions and Gold Nanoparticles.

Abstract

Nucleic acid testing requires skilled personnel and expensive instrumentation. A method for the colorimetric detection of oligonucleotides that combines cellulose microparticles with biomolecular recognition is presented. DNA sequences from Trypanosoma brucei and dengue are used as model targets. Cellulose microparticles (≈20 µm) are bioactived by anchoring anti-biotin antibodies via fusions that combine a carbohydrate-binding module (CBM) with the ZZ fragment of protein A. Samples are prepared by incubating DNA probes immobilized on ≈14 nm gold nanoparticles (AuNPs) with biotin-labeled targets and mixed with bioactive microparticles. The presence of unlabeled targets could also be probed by introducing a second, biotinylated DNA probe. The target:probe-AuNP hybrids are mixed with and captured by the microparticles, which change color from white to red. Depletion of AuNPs from the liquid is also signaled by a decrease in absorbance at 525 nm. It was possible to detect targets with concentrations as low as 50 n m. In the presence of noncomplementary targets, microparticles remain white and the liquid remains red. The system is able to discriminate targets with a high degree of homology (≈53%). Overall, it is demonstrated that simple systems for the visual detection of nucleic acids can be set up by combining cellulose microparticles with biomolecular recognition agents based on CBMs and AuNPs.