A continuous cell line (HBL-100) was obtained from primary cultures of cells derived from an early lactation sample of human milk. There was no evidence of a breast lesion in the milk donor. Karyotype analysis showed that all metaphases contained human chromosomes including a large acrocentric marker chromosome. Both desmosomes and cytoplasmic tonofibrils were observed during early passage. HBL-100 cells exhibited several characteristics of transformation including the ability to form colonies in soft agar, an aneuploid chromosome complement, and continuous growth.