T3,DITPA and CGS

Rat ventricular cardiomyocytes were prepared from 18 day old fetus. Approximately 6 million cells were plated on 100 mm culture plate. The serum used in the medium was depleted from thyroid hormone by passing through anion exchange resin column. After 48 hours of growth with 70% confluency, cells were exposed to either T3, DITPA or CGS for another 48 hours before harvesting. Final concentrations of T3, DITPA and CGS in the medium were 6nM, 218nM and 1nM respectively. RNA was prepared from cells with Triazol. Twenty micrograms of total RNA was reverse transcribed and labeled with Alexa Fluor 546 and 647. Labeled probes were hybridized with Qiagen's rat unigene oligo library. After 2 low and 1 high stringency washes Prolong antifade was added to the slides to prevent bleaching effect. Slides were scanned in Array Worx scanner with dual wave length. The data ratio was normalized using a location and intensity dependent Lowess formula. Keywords: equivalent probe