title: Fast on-site diagnosis of influenza A virus by Palm PCR and portable capillary electrophoresis.
abstract: A method combining Palm polymerase chain reaction (PCR) and portable capillary electrophoresis (CE) was developed for rapid on-site analysis of influenza A (H1N1) virus. The portable CE system was suitable for rapid diagnosis which was able to detect a sample in ∼4 min after sample loading, while the 'Palm PCR' system allowed for high-speed nucleic acid amplification in ∼16 min. The analysis time from DNA sample to analysis of amplified target DNA molecule was only ∼20 min, which was significantly less than slab gel electrophoresis with other commercially available PCR machine. When the 100-bp DNA ladder was separated, the relative standard deviation values (n=5) for the migration times and peak areas of the 100 and 200-bp DNA molecules were 0.26 and 8.9%. The detection limits were 6.3 and 7.2 pg/μL, respectively. The combined method was also able to identify two influenza A-associated genes (the HA and NP genes of the novel H1N1 influenza). CE separation was achieved with a sieving matrix of 1% poly(vinylpyrrolidone) (Mr=1,300,000) in 1× TBE buffer (pH 8.45). The combined Palm PCR-portable CE system should provide an improved, fast on-site molecular genetic diagnostic method.
authors: Department of Applied Chemistry, College of Applied Science, Kyung Hee University, Yongin-si, Gyunggi-do 446-701, Republic of Korea.
authors: Seoyeon
authors: S
authors: Lim
authors: Department of Applied Chemistry, College of Applied Science, Kyung Hee University, Yongin-si, Gyunggi-do 446-701, Republic of Korea.
authors: He
authors: H
authors: Nan
authors: Ahram Biosystems Inc., Seoul 133-120, Republic of Korea.
authors: Min-Jun
authors: MJ
authors: Lee
authors: Department of Applied Chemistry, College of Applied Science, Kyung Hee University, Yongin-si, Gyunggi-do 446-701, Republic of Korea. Electronic address: shkang@khu.ac.kr.
authors: Seong Ho
authors: SH
authors: Kang
mesh_terms: D004279:DNA, Viral / Q000032:analysis; D019075:Electrophoresis, Capillary / Q000191:economics / Q000295:instrumentation*; D004867:Equipment Design; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000302:isolation & purification*; D007251:Influenza, Human / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000191:economics / Q000295:instrumentation*; D015203:Reproducibility of Results
chemical_list: D004279:DNA, Viral
keywords: Fast diagnosis; H1N1 influenza; On-site analysis; Palm PCR; Portable CE