Five different samples of Drosophila melanogaster Kc167 cells were stained with Hoechst 33342, a DNA stain. The last sample (labeled nodsRNA, CIL 21789) is of wild-type cells. Each of the other four samples (labeled 48, 340, Anillin, and mad2, CIL 21749, 21739, 21759, and 21773, respectively) has a different gene knocked down by RNAi. This is the set of wild-type cells. There are 10 images of each sample, for a total of 50 images. The images were acquired on a Zeiss Axiovert 200M microscope. A recommended use for these five image sets is to algorithmically compute the number of cells in each image. A tab-delimited text file containing this information can be found at the URL indicated in the attribution box. The recommended citation when using these images is: "We used the Drosophila Kc167 1 image set (Carpenter, et al., Genome Biology, 2006) available from the Broad Bioimage Benchmark Collection (www.broad.mit.edu/bbbc)."