Genomic Analysis of a Hybridoma Batch Cell Culture Metabolic Status in a Standard Laboratory 5 L Bioreactor Bhargavi Kondragunta Center for Advanced Sensor Technology and Dept. of Chemical and Biochemical Engineering, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250 Div. of Cellular and Gene Therapy, Center for Biologics Evaluation and Research, Food and Drug Administration, 29 Lincoln Dr, Bethesda, MD 20892 Jing Han, and Bharat H Joshi Div. of Cellular and Gene Therapy, Center for Biologics Evaluation and Research, Food and Drug Administration, 29 Lincoln Dr, Bethesda, MD 20892 Kurt A. Brorson Div. of Monoclonal Antibodies, Center for Drug Evaluation and Research, Food and Drug Administration, 10903 New Hampshire, Silver Spring, MD 20903 Raj K. Puri Div. of Cellular and Gene Therapy, Center for Biologics Evaluation and Research, Food and Drug Administration, 29 Lincoln Dr, Bethesda, MD 20892 Shaunak Uplekar, Antonio R. Moreira, and Govind Rao Center for Advanced Sensor Technology and Dept. of Chemical and Biochemical Engineering, University of Maryland, Baltimore County, 1000 Hilltop Circle, Baltimore, MD 21250 DOI 10.1002/btpr.1605 Published online in Wiley Online Library (wileyonlinelibrary.com). Currently, there is a gap in the knowledge of the culture responses to controlled bioreac- tor environment during the course of batch cell culture from early exponential phase to sta- tionary-phase. If available, such information could be used to designate gene transcripts for predicting cell status and as a quality predictor for a controlled bioreactor. In this study, we used oligonucleotide microarrays to obtain baseline gene expression profiles during the time-course of a hybridoma batch cell culture in a 5 L bench-scale bioreactor. Gene expres- sion changes that were up or down modulated from early-to-late in batch culture, as well as invariant gene profiles with significant expression were identified using microarray. Typical cellular functions that seemed to be correlated with transcriptomics were oxidative stress response, DNA damage response, apoptosis, and cellular metabolism. As confirmatory evi- dence, microarray findings were verified with a more rigorous semiquantitative gene-specific Reverse transcriptase-polymerase chain reaction (RT-PCR). The results of this study suggest that under predefined bioreactor culture conditions, significant gene changes from lag to log to stationary phase could be identified, which could then be used to track the culture state. V V C 2012 American Institute of Chemical Engineers Biotechnol. Prog., 000: 000–000, 2012 Keywords: cell culture, genomic, time-course, sparged bioreactor, microarrays Introduction It is well recognized that in order to ensure consistent prod- uct quality (e.g. glycosylation 1,2 ), critical cell culture environ- mental parameters such as dissolved oxygen (DO), pH, CO 2 levels are controlled using set-points during a bioreactor pro- cess. Several changes in the culture environment can occur owing to varying requirement for aeration, and possibly agita- tion in cultures, in order to maintain set-point control for tem- perature, %DO and pH (CO 2 gas for acid control) in a bioreactor. For example, as cultures grow and become more dense, the culture’s aeration and agitation requirements increase, and remain high until death phase. In addition, there could be other bioreactor environmental stress factors 3–8 that could result in apoptosis. 9,10 These include gas sparging in culture media, high gas entrance velocity at the sparger site, increased hydrodynamic forces by air bubble bursting at the liquid surface or foaming, insufficient mixing at high cell den- sities, energy dissipation of impeller stream and possible reac- tive oxygen species formation, during the course of bioreactor culture. Increasing levels of apoptosis in cell culture has been associated with lower productivity, product quality, and cell Additional Supporting Information may be found in the online ver- sion of this article. Correspondence concerning this article should be addressed to G. Rao at grao@umbc.edu or R.K. Puri at Raj.Puri@fda.hhs.gov. V V C 2012 American Institute of Chemical Engineers 1