AbstractCold hardening treatment – a brief exposure to low temperatures – can protect certain insects against subsequent exposure to temperatures sufficiently low to cause damage or lethality. Microarray analysis to examine the changes in transcript abundance associated with cold hardening treatment (0 °C for 2 h followed by 30 min recovery at 25 °C) was undertaken in Drosophila melanogaster in order to gain insight into this phenomenon. Transcripts associated with 36 genes were identified, a subset of which appeared to be also differentially expressed after heat shock treatment. Quantitative RT‐PCR was used to independently determine transcript abundance of a subset of these sequences. Taken together, these assays suggest that stress proteins, including Hsp23, Hsp26, Hsp83 and Frost as well as membrane‐associated proteins may contribute to the cold hardening response.