Transcription assays are important for studying molecular mechanisms that control the initiation of transcription. It is often desirable or necessary to use a cell-free in vitro transcription assay, rather than transfection of a promoter and a reporter gene into a cell line, to study the regulation of transcription initiation. However, the preparation of transcriptionally active nuclear extracts from mammalian tissues can be a difficult and frustrating process. We report in this paper that the additional step of concentrating a nuclear extract preparation with a microconcentrator at the end of a previously described procedure (3) results in nuclear extracts that are consistently transcriptionally active. The promoter of the pulmonary surfactant protein A (SP-A) gene is the focus of the experiments in this study. Surfactant is a lipoprotein complex that is essential for normal lung function. SP-A is the major surfactant-associated protein and is known to be important in the structure, function and metabolism of surfactant.