Lowering culture temperature has been reported as a significant factor in the improvement of mammalian cell productivity. To determine the physiological changes which take place at different temperature cultivations, an NS0 cell line producing human-mouse chimeric antibody was cultured at 22, 34 and 37 C. Various cellular parameters such as viability, productivity, metabolism, apoptosis and cell cycle were studied and notable changes were shown to be accompanied by changes in metabolic rates. Reduction of the temperature to 22 C resulted in a decrease in the growth rate, inhibition of antibody production, arrest of cell cycle in G2 phase and delay in apoptosis. A slight increase in antibody production was observed at 34 C due to the increase of growth rate and prolonged stationary phase. To better understand and explore the mechanisms underpinning these biological alterations and to identify the genes involved in the genetic reprogramming, genome-wide analyses were performed using GeneChip Mouse Genome arrays. The examination of differential gene expression induced by temperature reduction demonstrated a specific pattern of gene expression in NS0 cells in response to temperature stress. The effect of temperature on transcription induced changes within a wide range of genes involved in metabolic and signalling pathways. Most deregulated genes involved in essential metabolic pathways (i.e. glycolysis/gluconeogenesis, pentose phosphate pathway and inositol metabolism) were repressed in cells cultured at 22 C. By combining gene expression and physiological changes at different temperatures it was possible to provide greater understanding of cell response to hypothermic conditions.