Identification of Adenovirus, Influenza Virus, Parainfluenza Virus, and Respiratory Syncytial Virus by Two Kinds of Multiplex Polymerase Chain Reaction (PCR) and a Shell Vial Culture in Pediatric Patients with Viral Pneumonia

Lee, Jong-Han; Chun, Jin-Kyong; Kim, Dong Soo; Park, Yongjung; Choi, Jong Rak; Kim, Hyon-Suk

Labopass TM RV detection kit (http://www.cosmo4.com/index_eng.html ) Nucleic acids from nasopharyngeal aspirates were also used for the Labopass TM kit. This kit was also designed to detect 12 types of viruses. The final reaction volume was 50 µL composed of 40 µL premixture and 10 µL nucleic acid. Two kinds of PCR protocols were used. AdV and human bocavirus (HboV) operated at 95˚C for 3 min, then 35 cycles of 95˚C for 1 min, 55˚C for 1 min, and 72˚C for 1 min, followed by a 5 min final extension at 72˚C and preserved at 4˚C. HCoV, EnV, PIFV, RhV, and RSV all operated at 42˚C for 60 min, 95˚C for 3 min, then 35 cycles of 95˚C for 1 min, 55˚C for 1 min, and 72˚C for 1 min, followed by a 5 min final extension at 72˚C and preserved at 4˚C. The PCR products were analyzed by identifying bands with a 2% agarose gel stained with ethidium bromide.