Maturation of cells of the mononuclear phagocyte lineage from bone marrow precursors to tissue macrophages (MAC) via circulating blood monocytes (MO) is a multistep process only partially understood. Similarly, MAC differentiation can be observed if MO are cultured in vitro. In an attempt to further characterize molecular changes occurring during this process we carried out differential screening of a MO‐derived MAC cDNA library using MO and MAC cDNA. After subcloning and confirmation by a second round of screening, partial sequencing of 41 cDNA clones was performed. In 33 clones the sequences of 7 different previously identified cDNAs were found. The mRNA expression of two of the corresponding genes (apolipoprotein E, ferritin light chain) is already known to be up‐regulated during MAC maturation. For one gene (cathepsin B), a specific up‐regulation of mRNA expression could be shown corresponding to previous protein data. For four genes [human cartilage glycoprotein (HC‐gp39), osteopontin, type IV collagenase, and tryptophanyl‐tRNA synthetase] the specific expression in MAC versus MO was previously unknown but could be confirmed by the use of Northern blot analysis. Of these genes, HC‐gp39 is especially interesting because it is only expressed during the late stages of MAC differentiation. J. Leukoc. Biol. 60: 540–545; 1996.