Livers from winter flounder collected at monthly intervals throughout the year were analyzed for their content of antifreeze protein mRNA. Putative mRNA was detected in liver RNAs from summer fish by liquid hybridization to kinetically purified antifreeze protein cDNA. These mRNA sequences were shown by Northern blot analysis to be of the same length as mature antifreeze protein mRNA isolated from winter fish, and were able to direct the incorporation of alanine into a translation product which comigrated with antifreeze preproprotein. The build-up of antifreeze protein mRNA levels in the autumn and their decline in the spring to summer levels were measured by liquid hybridization. These seasonal fluctuations match closely, but slightly precede, the rise and fall in plasma osmolality due to the presence of antifreeze protein. In mid-winter 0.5% of the total liver RNA is antifreeze protein mRNA, and although by late August the level of this mRNA has declined to 0.0007% of the total RNA, at no time during the summer is the mRNA undetectable. These results suggest that antifreeze protein production is more likely to be regulated by changing the rate of transcription of their genes than by switching them between active and inactive states.