A fully integrated microfluidic genetic analysis system with sample-in-answer-out capability.
We describe a microfluidic genetic analysis system that represents a previously undescribed integrated microfluidic device capable of accepting whole blood as a crude biological sample with the endpoint generation of a genetic profile. Upon loading the sample, the glass microfluidic genetic analysis system device carries out on-chip DNA purification and PCR-based amplification, followed by separation and detection in a manner that allows for microliter samples to be screened for infectious pathogens with sample-in-answer-out results in < 30 min. A single syringe pump delivers sample/reagents to the chip for nucleic acid purification from a biological sample. Elastomeric membrane valving isolates each distinct functional region of the device and, together with resistive flow, directs purified DNA and PCR reagents from the extraction domain into a 550-nl chamber for rapid target sequence PCR amplification. Repeated pressure-based injections of nanoliter aliquots of amplicon (along with the DNA sizing standard) allow electrophoretic separation and detection to provide DNA fragment size information. The presence of Bacillus anthracis (anthrax) in 750 nl of whole blood from living asymptomatic infected mice and of Bordetella pertussis in 1 microl of nasal aspirate from a patient suspected of having whooping cough are confirmed by the resultant genetic profile.
Easley
Christopher J
CJ
Department of Chemistry, University of Virginia, Charlottesville, VA 22904, USA.
Karlinsey
James M
JM
Bienvenue
Joan M
JM
Legendre
Lindsay A
LA
Roper
Michael G
MG
Feldman
Sanford H
SH
Hughes
Molly A
MA
Hewlett
Erik L
EL
Merkel
Tod J
TJ
Ferrance
Jerome P
JP
Landers
James P
JP
D004247:DNA / Q000302:isolation & purification*; D046209:Electrophoresis, Microchip / Q000379:methods; D005821:Genetic Techniques; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA