nanoKAP^sm 2008

 

The utilization of functional nanoparticles for biosensing has generated a great deal of interest because they are found to greatly enhance sensitivity of various given methods. Electrochemical biosensors based on nanoparticle labels are very attractive for bioassays, due to their high sensitivity, inherent simplicity, miniaturization, and low cost. In this presentation, we will give an overview on recent development of nanoparticles based biosensors for selective and sensitive detection of protein biomarkers.

Arnold Kell, PhD, and Benoit Simard, PhD, Principal Research Officer, Steacie Institute for Molecular Sciences, National Research Council, Canada*

Infectious and chronic diseases and bioterrorism are of tremendous global concern. The National Research Council of Canada is dedicated to developing nanoparticle platforms to aid in the rapid detection of these deadly diseases and security threats. Specifically, this presentation will highlight our efforts to utilize unique surface ligands anchored to magnetic nanoparticles, dye-doped silica nanoparticles and quantum dots to enable the selective labeling, magnetic confinement and optical detection of a variety of potentially harmful bacteria and cells in biological samples.

*In collaboration with: Kui Yu

We describe a new concept using combinatorial self-assembly of DNA nanotiles into micrometer-sized two-dimensional arrays that carry nucleic acid probes and barcoded fluorescent dyes to achieve multiplexed detection. The specificity and sensitivity of the arrays by detecting multiple DNA sequences and aptamer binding molecules were demonstrated. Accurate control of the interprobe distances and solution-based binding reactions ensures fast target binding kinetics. The detection sensitivity can be future improved using hybridization chain reaction (HCR) as a signal amplifier.

Principles and examples of electronic microarray applications for electric-field assisted (nano-)assembly of molecules and components for sensors and detection of pathogens will be summarized. Electric field and hydrodinamically enhanced biochemical reactions will be demonstrated in the array format such as rolling circle amplification for isothermal on-chip amplification with sensitivity approaching few molecules. Attempts toward miniaturization of the electronic microarray platform will be summarized with intended point-of-care applications.

Staphylococcal enterotoxins (SEs) are amajor cause of food-borne diseases. Traditionally, SEs assayed immunologically with ELISA. Carbon nanotubes’ (CNT) uniquemechanical and electronic properties combined with a large specific surface areamake them attractive for biosensing.We applied CNT to increase the sensitivity of a simple and portable point-of-care immunosensor based on the detection of Enhanced chemiluminescence (ECL) by a cooled Charge- Coupled Device (CCD) detector. This combination of ECL, CNT and CCD technologies is used to improve the detection of Staphylococcal Enterotoxin B (SEB) in food. Anti-SEB primary antibodies were immobilized onto the CNT surface and the antibody-nanotube mixture was immobilized onto a polycarbonate surface. SEB was then detected by a sandwich-type ELISA assay on the CNTpolycarbonate surface with an ECL assay. SEB in buffer, soymilk, apple juice, andmeat baby food was assayed with a limit of SEB with CCD detector is 0.01 ng/mL similar to the detection limit of a fluorometric detector, which is farmore sensitive than the conventional ELISA detection limit of ~1 ng/ml. Our simple, versatile and inexpensive CCD based Point-of-care detector combined with the CNT-ECL immunosensormethod can be used to simplify and increase sensitivity formany other types of diagnostics and detection assays.

We report the construction of a novel biosensing nanodevice to detect single molecules of target DNA, RNA, or protein on the same platform at the same time. The combination of reliable specificity, high sensitivity, and low cost should make this technology a viable option for widespread clinical use. Early diagnosis of diseases such as cancer can have a profound impact on a patient’s prognosis. The diagnoses are dependent on rapid, sensitive detection of specific protein and/or nucleic acid biomarkers. Rapid, sensitive detection and identification of pathogens is also critically important to minimize the transfer and spread of disease as well as to devise and evaluate effective treatment strategies.

Clark Tibbetts, PhD, Executive Vice President and CTO, Tessarae, LLC*

Oligonucleotide probes that are used to amplify target gene sequences are nanoscale information transducers. A single 25-nucleotide probe (8 nm length) represents extraordinary sequence specificity, as only one of 10¹⁵ different sequences of the four bases A, G, C and T. Such probes are noisy channels, due to idiosyncratic interactions of similar gene sequences from distinct organisms as well as specific mutations within a species. This accounts for unacceptably high rates of false negative and false positive results from PCR-based tests in critical applications. Claude Shannon and Edward Moore demonstrated that reliable (error-free) communications circuits result from networks comprised of less reliable (“crummy”) components or relays. This principle underlies the superior sensitivity, specificity and multiplicity of the TessArray Resequencing Pathogen Microarray (RPM) for simultaneous detection and identification many different viruses and bacteria in a single specimen. An RPM detector array network of almost one million different oligonucleotide probes detects and specifies almost 125,000 bases of virtually error-free pathogen gene nucleotide sequences. A real-world TessArray RPM application example illustrates rapid and unequivocally accurate detection and identification of any HN subtype of human and/or avian influenza virus.

*In collaboration with: K.Schafer and M.Lorence, TessArae; D.Swayne and Colleen Thomas, U.S. Dept of Agriculture

Oliver Lung, PhD, Research Scientist, Canadian Food Inspection Agency, Animal Diseases Research Institute, Canada*

Avian influenza virus (AIV) and Foot-and-Mouth Disease virus (FMDV) outbreaks can result in severe economic losses. We developed two electronic microarrays that can subtype all 16 H (hemagglutinin) subtypes of AIV, and all 7 serotypes of FMDV. Probes were also designed against the conserved regions of the AIV matrix (M) gene and FMDV polymerase coding region to detect all AIV and FMDV, respectively. Thirty-two influenza isolates representing all 16 H subtypes, and 23 FMDV isolates representing all 7 serotypes were correctly identified and typed using the two arrays.

*In collaboration with: A.Beeston, J.Geddes, K.Burton Hughes, J.Pasick, A.Clavijo, T.Furukawa-Stoffer, W.Mauro, and D.Deregt, Canadian Food Inspection Agency; and D.Hodko, Nanogen

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Chemical detection equipment is becoming more and more sensitive as the particles are smaller, yet not all chemical sampling equipment is in this range yet. Nano sized particles have different properties than the regular chemicals. There are already studies finding carbon nanotubes have asbestos like qualities and the bioaccumulative effects found in fish brains. Come discuss nanotoxicology, how the industry is changing. Resources on PPE, which MSDS chemical databases track at this level, and a survey of field sampling equipment that work on the nano scale. The Nano Safety Consortium is working with the EPA to have companies voluntarily come forward with safety data.