What is a Bioreactor Vessels or containers designed to support the optimal growth and metabolic activity of cells producing a product of interest Can be classified in several ways including Type of mixing ID: 635668
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Slide1
Bioreactor Basics
Mini-BIOMAN, July 2017Slide2
What is a Bioreactor?
Vessels or containers designed to support the optimal growth and metabolic activity of cells producing a product of interest
Can be classified in several ways including:
Type of mixing
Stirred Tank
Airlift
Mode of operation
Batch
culture
Fed batch
Perfusion
Type of vessel
Stainless steel/glass
Single use/ disposable Slide3
Ways to Prepare Bioreactors
CIP/SIP Systems
CIP (Clean
I
n
P
lace)
automatic cleaning of processing equipment, vessels, piping and in-line devices
m
inimal manual setup or shutdown
l
ittle or no operator intervention during cleaning
s
trong base, rinse, strong acid, final rinse with WFI
c
onductivity tests to monitor content of cleaning solutions and rinse water
SIP (Steam
I
n
P
lace)
e
quipment and vessels sterilized with clean steam
After sterilization system must remain pressurized to maintain sterility Slide4
Cell Culture Media for Bioreactors
Provides all the nutrition cells need within a narrow window of environmental conditions for
optimal expression of the target protein
Major
media components
Carbohydrate
energy source
– glucose
Nitrogen source
such as amino acids
Lipids
– often in the form of fatty acid
Cells also require
Trace minerals in the form of electrolytes (salts
)
fetal
bovine
serum supplements
–
not so common due to risk of animal
viruses
Chemically defined, serum free media commonly used – reduces threat of adventitious animal virus contaminants
Selective agents that cells require for optimal expression of the target protein Slide5
Bioreactor- Mixing Types
Stirred Tank
R
equired gases (e.g. 0
2
) nutrient media, cells are continuously stirred by agitator impellor (stirrer) at the bottom or top of the vessels
Baffles in the center of vessels ensures proper mixing and prevents formation of vortexes that might shear cells
Airlift
G
as is pumped from below through
a
sparge
tube within the
bioreactor creating bubbles which mixes the contents of the vessels
C
ontains
baffle that guides
gas
up through bioreactor
on one side of the baffle and then over and down the other side Slide6
Bioreactor Types – Stainless Steel
Made of durable material that can accommodate high volumes (up to 20,000L) of culture
Double walled, glycol jacketed with 4 layers that provide insulation/temperature control and sterile contact for cell cultures
Very large industrial sized reactors bioreactors have fixed vessel configurations with predefined port assemblies that can not be easily reconfigured
Expensive and time consuming cleaning procedures
high
costs to produce purified
water and steam for cleaning (CIP/SIP)
Slide7
Types of Bioreactors- Glass and Stainless Steel Slide8Slide9
Bioreactor and utilities and pipingSlide10
Top of 20,000 liter final bioreactor
10Slide11
Stainless Steel BioreactorsSlide12
Bioreactor Types – Single Use
Disposable bioreactors- intended
for one time
use
Components
are typically made of plastic and are disposed of after
use
Generally used for mammalian cultures
Cultivation chamber is inflated plastic bag
S
ingle
use technology in biomanufacturing is
becoming widespread - has advantages
and
disadvantagesSlide13
Advantages of Single Use Technology
Reduction or elimination of cleaning, sanitization, and sterilization steps. This reduces:
consumption of water – pure water is extraordinarily expensive to produce
energy used to produce purified water
consumption of cleaning and sanitizing chemicals (CIP)
eliminates the need to sterilize bioreactors (the vendor has done this)
eliminates the need to generate clean steam
need for cleaning and sterilization validation (and moves it to the vendor) – eases regulatory complianceSlide14
Single Use Advantages Continued
l
ower upfront capital costs – this becomes a major advantage for a small company or a new company wanting to start production quickly
faster cycle times
and faster, less expensive changeover between campaigns – less fear of cross-contamination (by eliminating the need for cleaning)
l
ower
r
isk – lower probability of cross-contamination with another product or microbial contaminationSlide15
Single Use Disadvantages
Scale Limitations (only up to 2000 liter cell culture)
increasing product titers and cell densities are making this less important
Limited to mammalian cell culture - low oxygen transfer coefficient rates excludes use of bacteria
Increased reliance on outside vendors – potential supply chain problems
Concerns over leachables/extractables from the plastics
Additional consumables cost
Environmental impact – increase in solid waste that currently goes to landfillSlide16
Disposable wave bioreactor and its mechanics Slide17
Single Use
B
ioreactors Slide18Slide19
Applikon eZ Control BioreactorSystem
Jacketed Stirred Tank Reactor + Controller
Controller - Front view
eZ Controller
c
ombines and supports actuators, pumps, and valves to optimize bench space
Controller for measurement and control of process variables –
pH
,
temp
,
DO, agitation
Control
Setpoints
Temperature
................37°C
pH ...........................
7.2
DO
...........................50
%
Agitation
.....................75
rpm
Slide20
No foam parameter button on our model
Control Panel Slide21
Bioreactor VesselSlide22Slide23Slide24Slide25Slide26
Cell Density & Viability
Automated Cell Counter; Trypan Blue Assay
Temperature
– 37°C
Monitored by temperature probeDissolved oxygen
– cells need oxygen to live
Controlled by DO probe and computer system based upon the rate at which oxygen molecules diffuse a membrane covering a set of electrodes
Glucose and Lactate
Monitored by Biochemistry Analyzer or spectrophotometric method
Cell Culture ParametersSlide27
Critical process parameter
Media contains bicarbonate; when combined with CO
2
from air, makes a buffering system to control pH
Culture Parameters- pHSlide28
Culture Parameters-Glucose
Glucose
+ O
2
CO
2
+ H
2
O
cells
Main energy source for cells to grow, metabolizeSlide29
Why measure Lactate?
Lactate is a product of glucose break down
During
cell proliferation glucose
concentration
decreases and lactate concentration increasesSlide30
BioanalyzerSlide31
YSI Analyzer 2900
User
Interface
2700 SELECT
Reagent Bottle
Compartment
Buffer Bottle
Waste Bottle
Calibration Standard
Manual Sample
Printer
Keypad/Display
Test Tube Holder
BUFFER
WASTESlide32
WASTE
BUFFER
STD
Sipper Assembly
Buffer Pump
Sipper Pump
Cal Pump
Buffer Bottle
Waste Bottle
Cal Standard Bottle
Sample Chamber
w/ Electrodes
Inside the
2900