Detection of biomarker gene expression by real-time polymerase chain reaction using amplified ribonucleic acids from formalin-fixed random periareolar fine needle aspirates of human breast tissue.
OBJECTIVE
To address the detection of breast cancer biomarker gene expression in formalin-fixed random periareolar fine needle aspiration (RPFNA) samples of benign breast tissue collected during breast cancer prevention trials by quantitative real-time polymerase chain reaction (qPCR).


METHODS
Formalin-fixed breast epithelial cells collected by RPFNA and processed as thin layer preparations were isolated by laser capture microdissection (LCM). Ribonucleic acid (RNA) was extracted and amplified using a single round of T7-based linear amplification followed by quality assessment and biomarker assay using TaqMan chemistry.


RESULTS
More than 80% of RPFNA samples yielded RNA of sufficient quantity and quality for measurement of a panel of biomarker genes following a single round of linear amplification. RNA and protein expression for estrogen receptor alpha, as assessed by LCM/qPCR and immunohistochemistry, were correlated. Amplification plots were similar for cDNA standards and cDNA derived from RPFNA samples.


CONCLUSIONS
Assessment of gene expression using amplified RNA from microdissected formalin-fixed RPFNAs can increase the number of biomarkers used during breast cancer chemoprevention trials.
Petroff
Brian K
BK
Breast Cancer Prevention Center, University of Kansas Medical Center, Kansas City, Kansas 66160, USA. bpetroff@kumc.edu
Phillips
Teresa A
TA
Kimler
Bruce F
BF
Fabian
Carol J
CJ
D015415:Biomarkers; D044963:Biopsy, Fine-Needle; D001940:Breast / Q000378:metabolism*; D018076:DNA, Complementary / Q000032:analysis; D005557:Formaldehyde; D020869:Gene Expression Profiling; D006801:Humans; D007150:Immunohistochemistry; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013997:Time Factors; D016707:Tissue Fixation; D014158:Transcription, Genetic / Q000235:genetics*
D015415:Biomarkers; D018076:DNA, Complementary; D012333:RNA, Messenger; D005557:Formaldehyde