Analytical and Bioanalytical Chemistry

, Volume 402, Issue 1, pp 315–323

Micro-total analysis system for virus detection: microfluidic pre-concentration coupled to liposome-based detection

  • John T. Connelly
  • Sowmya Kondapalli
  • Marc Skoupi
  • John S. L. Parker
  • Brian J. Kirby
  • Antje J. Baeumner
Original Paper

Abstract

An integrated microfluidic biosensor is presented that combines sample pre-concentration and liposome-based signal amplification for the detection of enteric viruses present in environmental water samples. This microfluidic approach overcomes the challenges of long assay times of cell culture-based methods and the need to extensively process water samples to eliminate inhibitors for PCR-based methods. Here, viruses are detected using an immunoassay sandwich approach with the reporting antibodies tagged to liposomes. Described is the development of the integrated device for the detection of environmentally relevant viruses using feline calicivirus (FCV) as a model organism for human norovirus. In situ fabricated nanoporous membranes in glass microchannels were used in conjunction with electric fields to achieve pre-concentration of virus–liposome complexes and therefore enhance the antibody–virus binding efficiency. The concentrated complexes were eluted to a detection region downstream where captured liposomes were lysed to release fluorescent dye molecules that were then quantified using image processing. This system was compared to an optimized electrochemical liposome-based microfluidic biosensor without pre-concentration. The limit of detection of FCV of the integrated device was at 1.6 × 105 PFU/mL, an order of magnitude lower than that obtained using the microfluidic biosensor without pre-concentration. This significant improvement is a key step toward the goal of using this integrated device as an early screening system for viruses in environmental water samples.

Keywords

Biosensor Microfluidic Pre-concentration FCV Liposomes 

Abbreviations

CPE

Cytopathogenic effects

DMEM

Dulbecco’s modified Eagle’s media

ELISA

Enzyme-linked immunosorbent assay

FBS

Fetal bovine serum

HEPES

4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid

HSS

HEPES–saline–sucrose

IMS

Immunomagnetic separation

LIA

Liposome immunoassay

mAb

Monocolonal antibody

μTAS

Micro-total analysis system

OG

Octyl-β-d-glucopyranoside

pAb

Polyclonal antibody

PBS

Phosphate-buffered saline

PBSS

Phosphate-buffered saline sucrose

PCR

Polymerase chain reaction

PFU

Plaque forming unit

RT-PCR

Reverse transcriptase-PCR

SERS

Surface-enhanced Raman spectroscopy

SNR

Signal-to-noise ratio

SRB

Sulforhodamine B

US EPA

United States Environmental Protection Agency

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Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • John T. Connelly
    • 1
  • Sowmya Kondapalli
    • 2
  • Marc Skoupi
    • 1
  • John S. L. Parker
    • 3
  • Brian J. Kirby
    • 2
  • Antje J. Baeumner
    • 1
  1. 1.Department of Biological and Environmental EngineeringCornell UniversityIthacaUSA
  2. 2.Sibley School of Mechanical and Aerospace EngineeringCornell UniversityIthacaUSA
  3. 3.Baker Institute of Animal Health, College of Veterinary MedicineCornell UniversityIthacaUSA

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