Nucleic acid-based assays for pathogen detection and identification offer sensitivity, specificity and resolution. Unfortunately, the manipulations for these assays have traditionally relied on nucleic acid isolation and amplification methods poorly suited for use outside of an appropriately equipped laboratory. Nonetheless, a significant need exists for more comprehensive, specific and facile diagnostics for use at the point-of-care. The recent emergence of several isothermal nucleic acid amplification strategies promise to alleviate the need for thermocycling hardware to achieve sensitive nucleic acid detection. Liberated from demanding hardware requirements, nucleic acid amplification becomes suitable for use under circumstances where access to a laboratory infrastructure is limited or unavailable. To enable sensitive, multiplexed detection of isothermally amplified nucleic acids without costly or complex instrumentation, we developed a rapid lateral flow chromatographic approach to DNA microarray fabrication and hybridization. In highly miniaturized embodiments, we refer to as lateral flow microarrays (LFM), the technology enables the detection of 250 amol of nucleic acid analyte in 2 minutes. Coupled with isothermal amplification, the method addresses many of the hurdles to the translation of nucleic acid assays from the laboratory to the field. However, a significant remaining challenge is presented by the need to render a complex biological sample, containing enzyme inhibitors and nucleases, suitable for isothermal amplification. Further, the integration of sample preparation, amplification and detection into a low cost, yet easily used, system is required to realize a facile and robust point-of-care diagnostic tool. The over arching hypothesis of this proposal is that a highly simplified and integrated nucleic acid analysis device can be realized using low cost lateral flow chromatography technologies. Making use of easily fabricated lateral flow devices offers an approach to sample preparation that is facile and familiar to the end user and provides a solid-phase support to capture and concentrate diagnostic targets from confounding sample matrix constituents. Leveraging our prior advances in LFM technology and the availability of sensitive detection schemes compatible with low cost instrumentation, we will develop an integrated nucleic acid analysis system making use of LFM and supporting field deployable sample preparation and amplification methods.

Public Health Relevance

The proposed work will result in the development of a device capable of rapidly detecting nucleic acid sequences in clinical specimens. The suitability of the device for use outside of a laboratory infrastructure will provide a unique ability to conduct informative DNA or RNA-based diagnostic assays at the point-of-care.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Specialized Center--Cooperative Agreements (U54)
Project #
5U54AI057156-10
Application #
8440793
Study Section
Special Emphasis Panel (ZAI1-DDS-M)
Project Start
Project End
2015-02-28
Budget Start
2013-03-01
Budget End
2015-02-28
Support Year
10
Fiscal Year
2013
Total Cost
$290,555
Indirect Cost
$42,944
Name
University of Texas Medical Br Galveston
Department
Type
DUNS #
800771149
City
Galveston
State
TX
Country
United States
Zip Code
77555
Pandey, Aseem; Lin, Furong; Cabello, Ana L et al. (2018) Activation of Host IRE1?-Dependent Signaling Axis Contributes the Intracellular Parasitism of Brucella melitensis. Front Cell Infect Microbiol 8:103
Russell-Lodrigue, Kasi E; Killeen, Stephanie Z; Ficht, Thomas A et al. (2018) Mucosal bacterial dissemination in a rhesus macaque model of experimental brucellosis. J Med Primatol 47:75-77
Matz, L M; Kamdar, K Y; Holder, M E et al. (2018) Challenges of Francisella classification exemplified by an atypical clinical isolate. Diagn Microbiol Infect Dis 90:241-247
Langsjoen, Rose M; Haller, Sherry L; Roy, Chad J et al. (2018) Chikungunya Virus Strains Show Lineage-Specific Variations in Virulence and Cross-Protective Ability in Murine and Nonhuman Primate Models. MBio 9:
Raja, B; Goux, H J; Marapadaga, A et al. (2017) Development of a panel of recombinase polymerase amplification assays for detection of common bacterial urinary tract infection pathogens. J Appl Microbiol 123:544-555
Nunes, Marcio R T; Contreras-Gutierrez, María Angélica; Guzman, Hilda et al. (2017) Genetic characterization, molecular epidemiology, and phylogenetic relationships of insect-specific viruses in the taxon Negevirus. Virology 504:152-167
Rossetti, Carlos A; Drake, Kenneth L; Lawhon, Sara D et al. (2017) Systems Biology Analysis of Temporal In vivo Brucella melitensis and Bovine Transcriptomes Predicts host:Pathogen Protein-Protein Interactions. Front Microbiol 8:1275
Paterson, Andrew S; Raja, Balakrishnan; Mandadi, Vinay et al. (2017) A low-cost smartphone-based platform for highly sensitive point-of-care testing with persistent luminescent phosphors. Lab Chip 17:1051-1059
Park, Arnold; Yun, Tatyana; Vigant, Frederic et al. (2016) Nipah Virus C Protein Recruits Tsg101 to Promote the Efficient Release of Virus in an ESCRT-Dependent Pathway. PLoS Pathog 12:e1005659
Pandey, Aseem; Cabello, Ana; Akoolo, Lavoisier et al. (2016) The Case for Live Attenuated Vaccines against the Neglected Zoonotic Diseases Brucellosis and Bovine Tuberculosis. PLoS Negl Trop Dis 10:e0004572

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