These suspensions were centrifuged at 3,000 x g for 20 minutes, and 100 [micro]l was used for RNA extraction by addition of a high-molarity solution of guanidinium
Each such instrument must be disinfected to the maximum extent possible, for example by washing repeatedly with detergent/proteinase solutions and exposing the washed instruments to less harsh chemicals (e.g., 6 M urea or 4 M guanidinium
thiocyanate) that have shown moderate to good disinfection of TSE tissue extracts (42-44).
Viral RNA was extracted by binding to size-fractionated silica beads (Sigma, Roosendaal, the Netherlands) in the presence of guanidinium
Total cellular RNA was isolated from mononuclear cells by Isogen-LS (Nippon Gene), with a modified acid guanidinium
In addition, [ADH2.sup.2]/[ADH2.sup.2] results obtained by SSCP analysis may be mistaken for [ADH2.sup.1]/[ADH2.sup.2] results obtained by standard RFLP analysis because of incomplete digestion of PCR products by the restriction enzyme, MsII, because of contamination by various substances (e.g., guanidinium
thiocyanate), which may inhibit the activity of MsII.
RNA was extracted from serum, PBMC, or infected cell pellets by the guanidinium
acid-phenol method (16).
Total RNA was extracted by the guanidinium
thiocyanate method (10).
In this method, DNA was combined in an equal volume of a solution of 6 mol/L guanidinium
isothiocyanate with biotinylated oligonucleotide capture probe sequences (33 pmol; Midland Certified Reagent Company) that were complementary to the regions of the DNA fragments that were outside of the regions that were amplified in subsequent PCR amplification reactions.
Total cellular RNA was extracted from infected B95a cells or directly from clinical specimens, if virus isolation was not successful, by the guanidinium
acid-phenol method (25).
 Nonstandard abbreviations: RT-PCR, reverse transcription-PCR; GERD, gastro-esophageal reflux disease; GIT, guanidinium
isothiocyanate; QRT-PCR, quantitative reverse transcription-PCR; [beta]-gal, galactosidase; [beta]-GUS, [beta]-glucuronidase; SDS, sodium dodecyl sulfate; and CK, cytokeratin; FAM, 6-carboxyfluorescein; and TET, tetrachloro-6-carboxyfluorescein.
The method is based on the use of guanidinium
isothiocyanate and adsorption of the nucleic acids to silica particles.
The cells were rinsed in PBS and total RNA was isolated by guanidinium
isothiocyanate using the TRIzol reagent (MBI Fermentas, Lithuania) according to the manufacturer's instructions.