Primary chicken kidney (CK) and chicken embryo kidney (CEK) cells were evaluated as possible substrates for growth of the cold-adapted attenuated influenza vaccine master strain A/Ann Arbor/6/60 (A/AA/6/60-ca). Yields of 10(6)-10(7) TCID50 per ml of culture fluid were obtained in either cell type. Yields from the human diploid strain MRC-5 were approximately 100-fold less. More reproducible cultures were obtained from CEK cells, using an overnight trypsinization step at 4 degrees C, than from CK cells. Comparable yields per embryo were obtained from CEK cells grown in roller cultures to those grown on the surface of microcarriers. These yields were less than those obtained from the allantoic fluids of whole embryos. Frozen storage of CEK or CK cells, after primary trypsinization, dispersal from a cultured CK primary monolayer or culture on microcarriers, was unsuccessful. The cold-adapted phenotype of A/AA/6/60-ca was retained after growth in CEK cultures and no differences in immunogenicity were detectable in mice between CEK- and allantoic-grown virus. Allantoic-grown preparations of A/AA/6/60-ca contained a lower protein concentration per infectious unit than those grown in CEK.