Metallic Striped Nanowires as Multiplexed Immunoassay Platforms for Pathogen Detection

Abstract

The rapid growth and development in biodetection technology has largely been driven by the emergence of new and deadly infectious diseases and the realization of biological warfare as new means of terrorism. [1,2] To address the need for portable, multiplex biodetection systems, we report here a novel biosensing platform using engineered nanowires as an alternative substrate for sandwich immunoassays (Figure 1 A). The nanowires are built through submicrometer layering of different metals by electrodeposition within a porous alumina template. [3,4] A variety of metals can be deposited: in this study, we employed stripes of gold, silver, and nickel. Owing to the permutations in which the metals can be deposited, a large number of unique yet easily identifiable encoded nanowires can be included in a multiplex array format.Image processing of an optical reflectance image can enable the stripe pattern to be identified rapidly, while fluorescence images report information on the degree of binding between the antibody-conjugated nanowires and a fluorophore-tagged antigen target. Such nanowires have been utilized to efficiently detect and report both DNA hybridization and immunoassay processes. [5,6] Herein, we demonstrate the feasibility of using multistriped metallic nanowires (Figure 1 A) in a suspended format to enable rapid and sensitive single and multiplex immunoassays for biowarfare agent simulants.Both the hybridization and kinetics of the capture of the target analyte in solution favor the nanowires over conventional fixed array-based formats. The incorporation of an appropriate ferromagnetic metallic component, for example, Ni, enables the nanoparticles to be manipulated by using magnetic fields. [7][8][9][10] To demonstrate the capability of directly detecting potential biological warfare agents in both clinical and environmental samples, a reagent set of three antigens generally accepted for use in simulating actual biothreat agents was chosen. The three nonpathogenic simulants include 1) Bacillus globigii (Bg) spores to simulate Bacillus anthracis and other bacterial species, 2) RNA MS2 bacteriophage to simulate Variola (virus for smallpox) and other pathogenic viruses, and 3) ovalbumin (Ova) protein to simulate protein toxins such as ricin or botulinum toxin. Besides the relative handling safety of these simulants, they were also chosen to reflect the variation in target sizes, ranging from large bacterial spores ( % 2 mm) to small protein molecules ( % 2 nm). Figure 1. A) Analogy between a conventional barcode and a metallic stripeencoded nanowire (diameter % 250 nm; length % 6 mm). Ni segments (50 nm) are deposited at both ends on the magnetic nanowire (not drawn to scale). B) Schematic of the sandwich immunoassay performed on a nanowire. C) Post-assay reflectance and fluorescence readout of the nanowires. The identity of the antigen present can be easily identified from the stripe pattern of the nanowires; for example, the fluorescently lit nanowire to which anti-Bg spore Ab was attached has a stripe pa...

Towards development of chemosensors and biosensors with metal-oxide-based nanowires or nanotubes

“…Also metal nanowires can be used for virus detection. For example, copper nanowires for the central channel of tobacco mosaic virus particles (Balci et al, 2006(Balci et al, , 2007 and Au/Ag striped nanowires as multiplexed immunoassay platforms for pathogen detection are shown (Tok et al, 2006). Lieber and co-workers reported a direct, real-time electrical detection of single virus particles with high selectivity by using nanowire FETs (Patolsky and Lieber, 2005;Patolsky et al, 2004).…”

Synthesis and Characterization of Noble Metal Nanowires

“…For instance, a mercaptoundecanoic acid (HS(CH 2 ) 10 CO 2 H) was used to coat Au-based nanowires resulting in surfaces functionalized with carboxylate groups that allowed the use of carbodiimide chemistry to conjugate primary amine groups of a capture antibody to the carboxylate groups on the nanowires. (Tok et al, 2006;Hermanson, 2008).…”

Heightened sense for sensing: recent advances in pathogen immunoassay sensing platforms

“…The multi-striped nanowires, when coated with specific antibodies, have been reported by Tok and co-workers to efficiently and accurately allow multiplex detection of biowarfare simulants (Figs 1a & b). 8 In addition, the metallic multi-striped nanowires have also been used by Sha and co-workers to detect and differentiate nucleic acid strands which contain single nucleotide polymorphisms. 9 The ability to fabricate addressable nanowire arrays as sensing platforms in either micro-or nanoscale has achieved tremendous progress over the last few years.…”

“…The nanowire is ~250 nm in diameter and ~6 µm in length. 8 [b] An image of a post-assay reflectance and fluorescence readout of metallic-stripe encoded nanowires. The identity of the antigen present can be easily identified from the nanowires striping pattern (0=Au; 1=Ag) and their corresponding fluorescence images.…”