Hindawi Publishing Corporation Journal of Nanomaterials Volume 2011, Article ID 781098, 11 pages doi:10.1155/2011/781098 Research Article Nanobiosensor for Detection and Quantification of DNA Sequences in Degraded Mixed Meats M. E. Ali, 1 U. Hashim, 1 S. Mustafa, 2 Y. B. Che Man, 2 M. H. M. Yusop, 2 M. Kashif, 1 Th. S. Dhahi, 1 M. F. Bari, 3 M. A. Hakim, 4 and M. A. Latif 5 1 Institute of Nano Electronic Engineering (INNE), Universiti Malaysia Perlis, 01000 Kangar, Perlis, Malaysia 2 Halal Products Research Institute, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia 3 School of Materials Engineering, Universiti Malaysia Perlis, 01000 Kangar, Perlis, Malaysia 4 Institute of Tropical Agriculture, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia 5 Department of Crop Science, Faculty of Agriculture, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia Correspondence should be addressed to U. Hashim, uda@unimap.edu.my Received 9 November 2010; Accepted 10 April 2011 Academic Editor: Claude Estournes Copyright © 2011 M. E. Ali et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. A novel class of nanobiosensor was developed by integrating a 27-nucleotide AluI fragment of swine cytochrome b (cytb) gene to a 3-nm diameter citrate-tannate coated gold nanoparticle (GNP). The biosensor detected 0.5% and 1% pork in raw and 2.5- h autoclaved pork-beef binary admixtures in a single step without any separation or washing. The hybridization kinetics of the hybrid sensor was studied with synthetic and AluI digested real pork targets from moderate to extreme target concentrations and a sigmoidal relationship was found. Using the kinetic curve, a convenient method for quantifying and counting target DNA copy number was developed. The accuracy of the method was over 90% and 80% for raw and autoclaved pork-beef binary admixtures in the range of 5–100% pork adulteration. The biosensor probe identified a target DNA sequence that was several-folds shorter than a typical PCR-template. This offered the detection and quantitation of potential targets in highly processed or degraded samples where PCR amplification was not possible due to template crisis. The assay was a viable alternative approach of qPCR for detecting, quantifying and counting copy number of shorter size DNA sequences to address a wide ranging biological problem in food industry, diagnostic laboratories and forensic medicine. 1. Introduction Hybrid biomaterials composed of functionalized nanopar- ticles, covalently linked to biomolecules such as peptides, proteins, and polynucleotides, are especially interesting for their size-dependent properties and dimensional similarities to biomacromolecules [1–8]. These nanobioconjugates are potential agents for multiplexed bioassays [9, 10], materials synthesis [1, 11–13], ultrasensitive optical detection and im- aging [14–16], in vivo magnetic resonance imaging (MRI) [17, 18], long-circulating carriers for targeted drug release [19], and structural scaffold for tissue engineering [20]. Nanoscaffolding and nanoquenching properties of thiol- capped gold nanocrystals (GNCs), covalently linked to fluo- rophore-lebeled oligonucleotide through metal-sulfur bond, were extensively studied for decades to detect specific se- quences and single-nucleotide mismatches [1–3, 21, 22]. Un- fortunately, such studies were limited to the laboratory level model experiments with synthetic oligo-targets. No studies so far practically explored the sequence and mismatch detect- ing power of the fluorophore-labeled-oligo-nanoparticle conjugates in heterogeneous biological samples. Hybridiza- tion kinetics of such nanobio-conjugates is also remained unknown. In this paper, we have structurally and functionally inte- grated a 3-nm diameter citrate-tannate-coated gold nano- crystal to fabricate a novel class of species-specific nanobio- sensor to determine pork adulteration in raw as well as proc- essed mixed meats. Several reports have highlighted the sig- nificance and importance of species determination in meat and meat products for health and religious concern to certain food ingredients as well as promotion of fair-trade with