AbstractClones have been isolated from the heterogeneousSpodoptera frugiperdaIPLB‐SF21‐AE insect cell population. Five of these clones, in addition to the parent cell line and the SF9 cell line (another clonal isolate of the parent cell line), have been compared in regards to morphology, growth, budded virus synthesis, and recombinant protein synthesis. No significant differences in cell morphology were found among these cell lines. There was, however, a significant difference in the average cell size, with diameters ranging from 9.30 ± 0.184 to 11.11 ± 0.22 μm and from 9.17 ± 0.05 to 11.25 ± 0.24 μm for cells growing in Excell 401 serum‐free medium in spinner flask cultures and in TNM‐FH medium supplemented with 10 FBS in tissue flask cultures, respectively. While no significant differences in the growth rates were found in TNM‐FH medium containing 10 calf serum, significant differences were found in Excell 401 serum‐free medium, with population doubling times ranging from 38.5 ± 6.6 to 64.5 ± 6.4 h in spinner flask studies. Significant differences in expression levels ofEscherichia coliβ‐galactosidase (β‐gal) were also found in both 12‐well plates and spinner flasks. In the 12‐well plate studies, the peak levels of β‐galactosidase obtained by these cell lines ranged from 0.332 ± 0.091 to 0.805 ± 0.117 mg/106cells and from 0.580 ± 0.130 to 1.458 ± 0.132 mg/106cells in Excell 401 and Hyclone Hy‐Q serum‐free media, respectively. In the spinner flask studies, peak expression levels ranged from 0.128 ± 0.053 to 0.573 ± 0.215 mg/106cells in Excell 401 serum‐free medium. Significant differences were also found in the expression levels of budded virus, which ranged from 64 ± 29 to 1125 ± 521 plaque‐forming units (pfu)/cell and from 67 ± 31 to 233 ± 95 pfu/cell for the wildtype and recombinant (β‐gal)Autographa
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