Liquid-based cytology in DNA-based molecular research: viability and potential application.
OBJECTIVE
To evaluate the effectiveness of UCM and PreservCyt liquid media in molecular analysis using exfoliated oral cells and analyze Tp53 codon 72 extracted from these cells.


METHODS
Exfoliated oral cells were placed in UCM and PreservCyt preservation media. DNA was isolated by proteinase K/phenol-chloroform protocol. Quantity and quality was measured by Nanodrop 1000. To evaluate the applicability of the isolated DNA to molecular techniques, DNA from oral cells was used to analyze Tp53 codon 72 polymorphism and GADPH genes.


RESULTS
DNA was successfully isolated from all samples, with an average concentration of 56.52 +/- 34.68 ng/microL for UCM and 42.13 +/- 25.50 ng/microL for PreservCyt. Average 260/280 ratios were 1.83 +/- 0.06 for UCM and 1.74 +/- 0.11 for PreserveCyt. Analysis of Tp53 codon 72 by polymerase chain reaction-restriction fragment length polymorphism was performed for all samples, with 4% of Pro/Pro, 60% of Arg/Arg and 36% of Arg/Pro genotype. In the analysis of GADPH multiplex polymerase chain reaction, fragments varying from 100 to 400 bp were obtained from all samples.


CONCLUSIONS
Cytologic liquid media are suitable for preserving cell samples for posterior DNA isolation and molecular analysis and could become reliable research tools.
Filho
Adhemar Longatto
AL
Institute of Investigation in Health and Life Sciences, School of Health Sciences, University of Minho, Braga, Portugal. longatto@ecsaude.uminho.pt
Gonçalves
Alberto Emanuel Pinheiro
AE
Martinho
Olga
O
Schmitt
Fernando C
FC
Reis
Rui Manuel
RM
D000293:Adolescent; D004252:DNA Mutational Analysis / Q000379:methods*; D004847:Epithelial Cells / Q000737:chemistry / Q000166:cytology*; D005260:Female; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics*; D006801:Humans; D008297:Male; D009061:Mouth Mucosa / Q000737:chemistry / Q000166:cytology*; D012150:Polymorphism, Restriction Fragment Length; D014021:Tissue Preservation / Q000379:methods*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*
C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases