ACC PRESENTATION

INTRODUCTION SCALE UP IN SUSPENSION CONTINUOUS CULTURES ADHERENT CELLS SCALE AND COMPLEXITY

SCALE UP IN SUSPENSION CULTURE It involves- 1. An increase in the volume of the culture medium 2. Agitation is done when the depth exceeds 5mm and above 5-10 cm. 3.Sparging with C02 is done to maintain adequate gas exchange. 4.Stirring should be between 30-100rpm as:- (i) Stirring at higher speed creates shear forces that would damage the cells. (ii) It should be sufficient enough to prevent cell sedimentation. 5. Antifoam or Pluronic F68 is added at conc. 0.01- 0.1% when serum conc. is 2%. 6. Carboxymethylcellulose(1-2%) is added to inc. the viscosity of the medium.

CONTINUOUS CULTURE A technique used to grow microorganisms or cells continually in a particular phase of growth. For example, if a constant supply of cells is required, a cell culture maintained in the log phase is best The conditions must therefore be continually monitored and adjusted accordingly so that the cells do not enter the stationary phase. The culture can be maintained at chemostat or biostat

CHEMOSTAT The cells are grown to mid log phase(monitored by daily cell counts). A measured volume of cells is removed each day and replaced with an equal volume of medium. Requirements:- 1.Stirrer vessel 2.Four ports- 2 for CO2 inlet and outlet One for medium inlet and one for spent medium outlet. Flow rate is regulated by peristaltic pump.

ADHERENT CELLS Transformed cells(such as HeLa-S3 are able to grow in liquid suspension where as anchorage dependent cells are not. The culture vessels need to be coated with a water repellent silicone e.g. Repelcote, Ca2+ conc. may need to be reduced. S-MEM medium is a variation of Eagle’s MEM with no calcium in formulation and has been used for the culture of HeLa-S3

SCALE AND COMPLEXITY A controlled fermentor or bioreactor should be used The culture vessels should have 1. Regulated input of medium and gas. 2.Provide the capability for data collection from oxygen,CO2 3. Glucose electrodes in the culture vessel They are regulated by a programmable control unit that helps in:- 1. Records and outputs data. 2. Used to regulate Gas and Liquid input and output. 3. Stirring speed 4. Temperature

THANK YOU