Control of redox potential in hybridoma cultures: effects on MAb production, metabolism, and apoptosis

Abstract

Culture redox potential (CRP) has proven to be a valuable monitoring tool in several areas of biotechnology; however, it has been scarcely used in animal cell culture. In this work, a proportional feedback control was employed, for the first time, to maintain the CRP at different constant values in hybridoma batch cultures for production of a monoclonal antibody (MAb). Reducing and oxidant conditions, in the range of −130 and +70 mV, were maintained in 1-l bioreactors through automatic control of the inlet gas composition. Cultures at constant DOT, in the range of 3 and 300 %, were used for comparison. The effect of constant CRP on cell concentration, MAb production, metabolism of glucose, glutamine, thiols, oxygen consumption, and programmed cell death, was evaluated. Reducing conditions resulted in the highest viable cell and MAb concentrations and thiols production, whereas specific glucose and glutamine consumption rates remained at the lowest values. In such conditions, programmed cell death, particularly apoptosis, occurred only after nutrient exhaustion. The optimum specific MAb production rate occurred at intermediate CRP levels. Oxidant conditions resulted in a detrimental effect in all culture parameters, increasing the specific glucose, glutamine, and oxygen consumption rates and inducing the apoptotic process, which was detected as early as 24 h even when glutamine and glucose were present at non-limiting concentrations. In most cases, such results were similar to those obtained in control cultures at constant DOT.

Generation of Antibodies Selectively Recognizing Epitopes in a Formaldehyde-Fixed Cell-Surface Antigen Using Virus-like Particle Display and Hybridoma Technology

“…1 Ratio of number of reactive mAbs and total number of mAbs identified as FF90-3T3/Gag/trNGFR binders. 2 Ratio of number of NGFR-unspecific mAbs and total number of mAbs identified as FFPE-3T3/Gag/trNGFR binders.…”

“…Hits, i.e., clones producing Ag-specific mAbs, are identified in one or multiple screening rounds for binding against target and non-target Ags employing enzyme-linked immunosorbent assay (ELISA) or flow cytometry. Isolated hybridoma cell clones can subsequently also be employed for the production of the cognate mAbs on a larger scale [ 2 ].…”

“…Ratio of number of positive hybridoma supernatants to total number of screened supernatants 2. Ratio of number of NGFR-unspecific hybridoma supernatants to 1st screening positive ones.…”

Online control of cell culture redox potential prevents antibody interchain disulfide bond reduction

“…The effects of cell culture redox potential have previously been investigated for their effect on cell growth and productivity in hybridoma cultures (Meneses‐Acosta, Gomez, & Ramirez, 2012). However, to the authors' knowledge, this is the first report demonstrating the use of controlling cell culture redox potential as a means to improve product quality.…”