Sonification as a means of enhancing the detection of gene expression levels from formalin-fixed, paraffin-embedded biopsies.
We describe a novel approach for the isolation and quantification of mRNA from archival specimens by using oligo(dT)25 paramagnetic beads and the reverse transcription polymerase chain reaction (RT-PCR). A preliminary study is conducted to evaluate experimental conditions and the effects of varied periods of sonification on the yield of mRNA isolated from formalin-fixed, paraffin-embedded human biopsy samples. Validation of this approach will involve the comparison of observed beta-actin gene expression levels among different samples from the same patient after various periods of sonification. The novelty of this method is that it offers a means of analyzing archival samples using semiquantitative RT-PCR following sonification-enhanced mRNA extraction. The utility of this protocol allows for gene expression studies to be done effectively using the formalin-fixed, paraffin-embedded archival samples that are commonly available in all hospital pathology departments.
Houze
T A
TA
Ostra University Hospital, University of Göteborg, Sweden. thomas.houze@mailbox.swipnet.se
Gustavsson
B
B
D000199:Actins / Q000235:genetics; D001706:Biopsy; D015179:Colorectal Neoplasms / Q000737:chemistry; D017931:DNA Primers; D005557:Formaldehyde; D015870:Gene Expression / Q000235:genetics*; D006801:Humans; D008280:Magnetics; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D012334:RNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification*; D013010:Sonication
D000199:Actins; D017931:DNA Primers; D012334:RNA, Neoplasm; D005557:Formaldehyde