Navigation Links
Gram Quantities of MAb Produced with Simple Bioreactor in Serum-Free Perfusion Culture Replacing Ascitic Fluid Production

By J. Doyle, L. Johnstone and L. Cottis
AGEN Biomedical Ltd. Queensland, Australia
http://www.agen.com.au


The CelliGen Plus immobilized-bed perfusion was investigated as a means for producing large quantities of MAb from a hybridoma, as an alternative to in vivo production. This system best represented a production facility that was easy to operate, flexible in its use, small enough to enable autoclaving and yet large enough to produce the required quantities of MAb. Three cell culture trials were performed using serum-free medium. Each run proceeded in the same manner, with MAb yields of 39.3, 33.8 and 30.3 grams. The perfusion system enabled us to scale down the size of the reactor needed, as the yields were significantly higher than that obtainable in batch fermentation which would have necessitated a bioreactor of several hundred liters.


Materials and Methods
All runs were performed in a 7.5 Liter NBS CelliGen Plus bioreactor with the basket impeller (New Brunswick Scientific, Edison, NJ). Working volume was 5 L and the bed volume was 2.6 L. Cells were inoculated at a cell density of 2 x 105 cells/mL at > 80% viability. Temperature was maintained at 37C, pH 7.20 and top dissolved oxygen (DO) at 50% of air saturation. The pH was controlled via the interactive pH-DO controller with CO2 at the initial stage and then with an 8% NaHCO3 solution when perfusion reached 2 volumes/day. Agitation rate was initiated at 80 rpm and increased throughout the run to maintain the bottom DO concentration at > 35% air saturation. The maximal rate did not exceed 150 rpm. Gas supply was via the ring sparger at a rate not exceeding 0.13 LPM and this was only increased when the top DO setpoint could not be maintained.

Gibcos Hybridoma Serum-Free Medium was used, supplemented with between 2 - 4 mM L-glutamine and 100 U/mL penicillin and 100 Ug/mL streptomycin. A separate 5% glucose solution was fed to the cultures when the perfusion rate reached 2 volumes/day to maintain the residual glucose concentration at > 1 g/L. The hybridoma cell line used produces an IgG1 MAb. Perfusion at 1 volume/ day (based on the bed volume), was initiated 24 hours after the commencement of the run, with increases to 2 then 3 volumes/day also 24 hours apart. The perfusion rate was maintained at 3 volumes/day for the duration.


Results and Discussion
The MAb produced in these cultivations has been traditionally produced from ascitic fluid with an average yield after purification of 5 mg/mouse. Yields produced using a contract service in 10 & 250 L batch reactors produced ~35 mg/L before purification. Using the CelliGen Plus perfusion system, yields of between 4 - 5 times that of the batch fermentation were obtained, based on purified material from the perfusion system but unpurified material from the batch system ( see table ).

All AGEN staff found the CelliGen Plus bioreactor very user friendly, easily simplifying an otherwise complicated and laborious procedure. The bioreactors display and associated BioCommand software provided fermentation information at the touch of a fingertip and trend graphing of process controls could also be viewed via BioCommand software. These features made the data handling, interpreting and report writing much simpler, something much appreciated by all staff. For CelliGen Plus literature contact your local NBS sales representative or write us at bioinfo@nbsc.com.



'"/>

Source:


Page: All 1 2

Related biology technology :

1. Simple, Sensitive, and Rapid Detection of FLAG -Tagged Fusion Proteins
2. A Simple and Rapid Technique to Purify High-Quality Midiprep DNA
3. Simple Purification of DNA from Plasmid Minipreps, PCR Amplifications and Agarose Gels
4. Simple, Sensitive Isotyping of Mouse Monoclonal Antibodies
5. Simple Isolation of RNA from Tissue and Cultured Cells
6. HSVision Molecular Beacon Detection Module Rapidly Detects Herpes Simplex Virus DNA
7. Simple and Rapid Isolation of Plant RNA Using the Eppendorf Perfect RNA, Eukaryotic, Maxi Kit
8. The Fastest, Simplest, and Most Effective Way to Remove DNA Contamination
9. Simple Approaches for Optimization of RT-PCR
10. Detection of Morphine and Morphine-Glucuronide in Saliva or Urine with a Rapid and Simple Method Without Derivatization on the Varian 500-MS
11. CHO Cell Culture Application Using a BioFlo 110 Benchtop Bioreactor
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:2/24/2017)... ... ... FireflySci, Inc is an explosive small business that continues to grow at ... their powerful cuvette and spectrophotometer calibration to the spectroscopy world. Now ... as they add yet another mark on the global map. , With distributors in ...
(Date:2/23/2017)... RAFAEL, Calif., Feb. 23, 2017 ... U.S. dollars, except per share data, unaudited)Three Months Ended ... ChangeTotal BioMarin Revenue $   ...   22832%$ 1,117$   89026%Aldurazyme Net Product Revenue ... 906538%34823946%Naglazyme Net Product Revenue  ...
(Date:2/23/2017)... Feb. 23, 2017  Seattle,s upscale Capitol Hill neighborhood, with its ... place for a head lice treatment salon to set up ... Tuscan restaurant and a French bistro on E Madison Ave, ... aren,t just any old lice clinic, we pride ourselves on ... and release some of the stigma associated with lice. Everyone ...
(Date:2/23/2017)... ... February 23, 2017 , ... ... insulin, cortisol, CRP, adiponectin, uric acid, and/or other biomarkers or SNPs of interest) ... from Salimetrics’ SalivaLab , the relationship between insulin and other relevant biomarkers ...
Breaking Biology Technology:
(Date:2/8/2017)... About Voice Recognition Biometrics Voice recognition biometrics ... a stored voiceprint template. Acoustic features of an ... are compared to distinguish between individual voices. Voice ... PCs already have a microphone and can authenticate ... are most likely to be deployed in telephone-based ...
(Date:2/8/2017)... -- Report Highlights The global biosurgery market ... in 2016 at a compound annual growth rate (CAGR) ... - An overview of the global market for biosurgery. ... 2015 and 2016, and projections of compound annual growth ... on the basis of product type, source, application, and ...
(Date:2/6/2017)... Feb. 6, 2017 According to Acuity ... driving border authorities to continue to embrace biometric ... are 2143 Automated Border Control (ABC) eGates and ... at more than 163 ports of entry across ... 2016 achieving a combined CAGR of 37%. APC ...
Breaking Biology News(10 mins):