Effect of conditioned medium factors on productivity and cell physiology in Trichoplusia ni insect cell cultures.
The influence of conditioned medium (CM) on cell physiology and recombinant protein production in Trichoplusia ni insect cells (T. ni, BTI-Tn-5B1-4) has been investigated. Cell cycle analysis showed that a high proportion of the cell population (80-90%) was in G1 during the whole culture, indicating that the S and G2/M phases are short relative to the G1 phase. Directly after inoculation, a rapid decrease of the S-phase population occurred, which could be observed as a lag-phase. The following increase in the number of cells in S occurred after 7 h of culture for cells in fresh medium, whereas for cells with the addition of CM it occurred at an earlier time point (5 h) and these cells had therefore a shorter lag-phase. The initial changes in the S-phase population were also affected by the inoculum cell density, as higher seeding cell densities resulted in a more rapid increase in the S-phase population after inoculation. These changes in cell cycle distribution were reflected in the cell size, and the CM-cells were smaller than the cells in fresh medium. Recombinant protein production in T. ni cells was improved by the addition of CM. The specific productivity was increased by 30% compared to cells in fresh medium. This beneficial effect was seen between 20 and 72 h of culture. In contrast, the highest specific productivity was obtained already at 7 h for the cells in fresh medium and then decreased rapidly. The total product concentration was around 30% higher in the culture with CM compared to the culture in fresh medium, and the maximum product concentration was obtained on day 2 compared to day 3 for the cells in fresh medium. Our results indicate that the positive effect on productivity by CM is related to its growth-promoting effect, suggesting that the proliferation potential of the culture determines the productivity.
Calles
Karin
K
Karo Bio AB, Huddinge, SE-141 57 Huddinge, Sweden.
Eriksson
Ulrika
U
Häggström
Lena
L
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002453:Cell Cycle / Q000187:drug effects; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology*; D019476:Insect Proteins / Q000096:biosynthesis* / Q000187:drug effects; D009036:Moths / Q000166:cytology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000187:drug effects; D013997:Time Factors
D017077:Culture Media, Conditioned; D019476:Insect Proteins; D011994:Recombinant Proteins