Successful application of microarray technology to microdissected formalin-fixed, paraffin-embedded tissue.
The establishment of a reliable method for using RNA from formalin-fixed, paraffin-embedded (FFPE) tissue would provide an opportunity to obtain novel gene expression data from the vast amounts of archived tissue. A custom-designed 22,000 oligonucleotide array was used in the present study to compare the gene expression profile of colonic epithelial cells isolated by laser capture microdissection from FFPE-archived samples with that of the same cell population from matched frozen samples, the preferred source of RNA. Total RNA was extracted from FFPE tissues, amplified, and labeled using the Paradise Reagent System. The quality of the input RNA was assessed by the Bioanalyzer profile, reverse transcriptase-polymerase chain reaction, and agarose gel electrophoresis. The results demonstrate that it is possible to obtain reliable microarray data from FFPE samples using RNA acquired by laser capture microdissection. The concordance between matched FFPE and frozen samples was evaluated and expressed as a Pearson's correlation coefficient, with values ranging from 0.80 to 0.97. The presence of ribosomal RNA peaks in FFPE-derived RNA was reflected by a high correlation with paired frozen samples. A set of practical recommendations for evaluating the RNA integrity and quality in FFPE samples is reported.
Coudry
Renata A
RA
Division of Population Science, Fox Chase Cancer Center, 333 Cottman Ave., Philadelphia, PA 19111, USA.
Meireles
Sibele I
SI
Stoyanova
Radka
R
Cooper
Harry S
HS
Carpino
Alan
A
Wang
Xianqun
X
Engstrom
Paul F
PF
Clapper
Margie L
ML
D003106:Colon / Q000166:cytology; D017931:DNA Primers; D004587:Electrophoresis, Agar Gel; D004847:Epithelial Cells / Q000737:chemistry; D005069:Evaluation Studies as Topic; D005404:Fixatives; D005557:Formaldehyde; D006801:Humans; D007834:Lasers; D042282:Microdissection; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016612:Paraffin Embedding; D012313:RNA / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014021:Tissue Preservation / Q000379:methods*
D017931:DNA Primers; D005404:Fixatives; D005557:Formaldehyde; D012313:RNA