2.2. Detection of IgG and IgM antibodies
Anti-HEV IgG and IgM antibodies were detected by
enzyme-linked immunosorbent assays (ELISAs) by using of
rat HEV-LPs as antigen as described previously (Li et al.,
2011).
2.3. Nested broad-spectrum RT-PCR
RNA extraction was performed using the MagNA Pure
LC system with MagNA Pure LC Total Nucleic Acid isolation
kit (Roche Applied Science, Mannheim, Germany) according
to the manufacturer’s recommendations. Reverse
transcription (RT) was performed at 42 8C for 50 min
followed by 70 8C for 15 min in a 20ml reaction mixture
containing 1ml of SuperscriptTM II RNase H reverse
transcriptase (Invitrogen Inc., Carlsbad, CA), 1ml of the
oligo(dT) primer, 1 ml of RNaseOUTTM, 2 ml of 0.1 M
dithiothreitol, 4 ml of 5 RT buffer, 1ml of 10 mM
deoxynucleoside triphosphates, 5m1 of RNA, and 5m1
of distilled water.