Detection of DNA using gold nanoparticle-coated silica nanoparticles.

We report a sensitive lateral flow assay (LFA) in which the assay colour change originated from reporter labels constructed from silica spheres (radius = 450 nm) coated with approximately 3.9 × 103 gold nanoparticles (radius = 8.5 nm). These reporter labels were modified with DNA and deposited in the conjugation area of an LFA device assembled on wax-patterned Fusion 5 paper. Test and control zones of the device were pre-loaded with capture probe formed by avidin-coated mesoporous silica nanoparticles attached with biotin-tagged DNA sequences. Proof-of-concept was demonstrated by the detection of a partial sequence of the actin gene of Colletotrichum truncatum. The DNA target could be detected with an LOD of 46 pM, which was 5 times lower than a comparative assay using gold nanoparticles alone. The assay showed good selectivity against the Colletotrichum species C. scovillei and C. gloeosporioides, as well as against DNA from the fungal genera Aspergillus niger and Alternaria alternata. There was negligible change in sensor response over storage for one month at room temperature. The LFA was used to detect PCR products following extraction from mycelium.

A new and high-performance microfluidic analytical device based on Fusion 5 paper for the detection of chili pepper anthracnose pathogen Colletotrichum truncatum.

A microfluidic analytical device based on wax-patterned Fusion 5 paper was designed and fabricated to facilitate early detection and improve control of anthracnose disease. Here, a rapid, specific, on-site, and low operational cost nucleic acid biosensor (ACT-Ct-PAD) based on the actin gene (ACT) and wax-patterned Fusion 5 paper was used to detect the PCR products of Colletotrichum truncatum (Ct), the main causal agent of chili anthracnose in Asia. The sensor was developed by using DNA conjugated gold nanoparticles (AuNPs-DNA) as a detection probe, which will hybridize to a complementary target sequence. Avidin coated mesoporous silica particles were attached to biotin-tagged DNA sequences forming capture probes, which were immobilized on the test and control zones of the device. The hybridization complex (MSP-dsDNA-AuNPs) produces an intense red color, which provides a platform for colorimetric detection. By targeting an actin gene sequence, the ACT-Ct-PAD device allows the detection of Ct DNA within 15 min. The specificity of the sensor was confirmed by the absence of a positive signal for DNA from non-target Colletotrichum species and two different fungal genera. Our wax-patterned Fusion 5 sensor provides a simple tool for the rapid nucleic acid diagnosis with a detection limit down to 17.42 femtomoles. This method has the potential to be applied for protein assay as well; hence, it has a considerable impact on on-site diagnostics.