Improved detection of viral RNA isolated from liquid-based cytology samples.
BACKGROUND
Molecular diagnosis requires the ability to obtain high-quality nucleic acids that are representative of the disease state. We evaluated the recovery and detection of limiting amounts of viral oncogenic RNA from cells fixed in liquid-based cytology media.


RESULTS
Serial dilutions of a human papillomavirus (HPV)-positive cell line fixed in a liquid media was used as a model system. Total nucleic acid (TNA) extraction produced RNA with clearly visible ribosomal bands even after one year of storage. These TNA extracts, treated with DNase-I, were used in an RT-PCR assay for HPV-16 E6-E7 oncogenic transcripts. With chemiluminscent Southern blot detection, samples with one HPV-positive cell in 30,000 were consistently detected.


CONCLUSIONS
PreservCyt-fixed cells can yield RNA suitable for molecular assays even after one year of storage.
Tarkowski
T A
TA
Viral Exanthems and Herpesvirus Branch, Division of Viral and Rickettsial Diseases, Centers for Disease Control and Prevention, 1600 Clifton Road, Atlanta, GA 30333, USA.
Rajeevan
M S
MS
Lee
D R
DR
Unger
E R
ER
D015139:Blotting, Southern; D002624:Chemistry, Clinical / Q000379:methods*; D004247:DNA / Q000032:analysis*; D004587:Electrophoresis, Agar Gel; D005260:Female; D006801:Humans; D008163:Luminescent Measurements; D017720:Molecular Epidemiology; D009856:Oncogene Proteins, Viral / Q000096:biosynthesis; D027383:Papillomaviridae / Q000235:genetics; D050725:Papillomavirus E7 Proteins; D012367:RNA, Viral / Q000032:analysis*; D012097:Repressor Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured; D002583:Uterine Cervical Neoplasms / Q000821:virology
C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D012367:RNA, Viral; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16; D004247:DNA