Thermobiochemical evidence for the rapid metabolic rate in hybridoma cells genetically engineered to overexpress the anti-apoptotic protein bcl-2 in batch culture

H...............H

Show simple item record

dc.contributor.author Kemp, Richard B.
dc.contributor.author Olomolaiye, D.
dc.contributor.author Feng, Y.
dc.date.accessioned 2009-10-06T14:10:04Z
dc.date.available 2009-10-06T14:10:04Z
dc.date.issued 2004-07-30
dc.identifier.citation Kemp , R B , Olomolaiye , D & Feng , Y 2004 , ' Thermobiochemical evidence for the rapid metabolic rate in hybridoma cells genetically engineered to overexpress the anti-apoptotic protein bcl-2 in batch culture ' Thermochimica Acta , pp. 207-216 . , 10.1016/j.tca.2003.07.020 en
dc.identifier.issn 1872-762X
dc.identifier.other PURE: 122186
dc.identifier.other dspace: 2160/3159
dc.identifier.uri http://hdl.handle.net/2160/3159
dc.description Feng, Y., Olomolaiye, D., Kemp, R. B. (2004). Thermobiochemical evidence for the rapid metabolic rate in hybridoma cells genetically engineered to overexpress the anti-apoptotic protein bcl-2 in batch culture. Thermochimica Acta, 417,(2),207-216. Sponsorship: BBSRC grant 2/E10985. en
dc.description.abstract A serious problem in the culture of animal cells to produce therapeutic proteins is apoptosis (programmed cell death) because it restricts the ability of the cell culture to yield the heterologous material. One means to delay apoptosis is genetically to engineer the anti-apoptotic gene bcl-2 into the genome. This had been adopted elsewhere to give TB/C3 hybridoma cells that overexpressed bcl-2 (pEF bcl-2-MC1neopA plasmid) and a control, pEF, which contained a nonsense sequence (pEF-MC1neopA). In the spinner cultures used in the present investigation, bcl-2 cells grew for 60 h to give greater cell density at a faster specific rate and with prolonged better viability than the controls in which, after 36 h, the membrane integrity and the apoptotic index of the cells declined rapidly leading to death. Prior to 36 h, production of the monoclonal antibody was only slightly higher in the pEF control giving little evidence for the metabolic burden of its production on antibody synthesis. Bcl-2 synthesis, however, was associated with 125% increase in heat flow rate (HFR) measured in the chemically (triacetin) calibrated batch microcalorimeter. HFR is a function of the metabolic rate and it is suggested that its greater level in the bcl-2 cells than the control was caused by the increased protein production due to the bcl-2 gene expression. The bcl-2 cells had an increased lactate flux compared with the control. The calorimetric–respirometric (CR) ratio confirmed the likelihood that glycolysis rather than glutaminolysis was the primary reason for this increase. It may be due to the limitation in mitochondrial capacity and/or the paucity of biosynthetic precursors leading to production of them from glucose. The stationary liquid phase balance (SLPB) was shown to be the appropriate on-line method to measure the oxygen uptake rate (OUR) in the dilute cell suspensions grown in the tank bioreactor. The metabolic rate expressed as HFR was measured in a customised flow microcalorimeter, which had been calibrated using the enthalpy of hydrolysis produced by the new test reaction using methyl paraben. Both HFR and OUR declined before the decrease in cell density in batch culture, indicating that they were sensitive indicators of deteriorating environmental conditions. However, neither proved to be an early detector of the onset of apoptosis. en
dc.format.extent 10 en
dc.language.iso eng
dc.relation.ispartof Thermochimica Acta en
dc.title Thermobiochemical evidence for the rapid metabolic rate in hybridoma cells genetically engineered to overexpress the anti-apoptotic protein bcl-2 in batch culture en
dc.type Text en
dc.type.publicationtype Article (Journal) en
dc.identifier.doi http://dx.doi.org/10.1016/j.tca.2003.07.020
dc.contributor.institution Aberystwyth University en
dc.contributor.institution Institute of Biological, Environmental and Rural Sciences en
dc.description.status Peer reviewed en


Files in this item

Files Size Format View

There are no files associated with this item.

This item appears in the following Collection(s)

Show simple item record

Search Cadair


Advanced Search

Browse

My Account