Course Outline
Laboratory Sessions
Fermentation Engineering Module: Production of a recombinant protein
Concepts studied:
- direct and indirect methods for studying cell growth kinetics using experiments and simple mathematical models
- measurement of consumption or production rates for substrates, waste products and recombinant product (example: collagen)
- determination of volumetric mass transfer coefficient experimentally using the dynamic and steady-state methods, as well as from published correlations
- cell metabolism using on-line measurements of oxygen uptake and carbon dioxide production rates; respiratory quotient calculations and control of the process
- construction of a carbon mass balance
- process performance: production rate, yield coefficients, productivity
- mathematical and practical basis of P-I-D controllers
Skills taught:
- plasmid manipulation and E. coli cell transformation
- design of a stirred-tank microbial bioreactor and its controls
- medium design, preparation and bioreactor autoclaving
- aseptic techniques, sampling and data monitoring
- making sterile connections using traditional techniques
- operating the bioreactor in the batch and fed-batch mode
- feedback control strategies for high-cell density fermentations
- process scale-up
- assays:
- vis-spectrophotometry and dry cell weight (to measure cell growth)
- HPLC (to measure substrate and waste product concentrations)
- polarographic probes (to measure dissolved oxygen levels)
- UV-spectrophotometry (to measure total proteins)
- enzymatic (to measure a product such as collagen)
- mass spectrometry (to determine gas composition on-line)
Cell Culture Engineering Module: Production of recombinant antibodies and vaccines
Concepts studied:
- direct and indirect methods for studying cell growth kinetics using experiments and simple mathematical models
- measurement of consumption or production rates for substrates and waste products (example: heat-shock protein 96)
- determination of volumetric mass transfer coefficient from cell-free experiments
- connection of cell metabolism, yield coefficients and productivity to process control
- delta P and crossflow
- transmembrane pressure and flux
- gel layer and membrane fouling
- process yield, recovery and mass balance
Skills taught:
- aseptic techniques and cell passaging
- design of the traditional (cell culture) stirred-tank reactor and the novel Wave bioreactor and their controls
- oxygen measurement with invasive Clark electrode or non-invasive optical sensor, and mass transfer coefficient determination in a cell-free bioreactor
- inoculum preparation in T-flasks
- making sterile connections using a novel tube-fusing technique
- design of a microfiltration process for maximizing cell recovery
- mass transport in ultrafiltration and the design of an ultrafiltration process for product concentration and purification
- cell harvesting by filtration
- design and implementation of a cell disruption process
- cell debris removal by centrifugation and crossflow filtration
- assays:
- automated dye-exclusion assay (Trypan blue) and microscopy (to measure cell growth and cell viability)
- enzymatic (to measure substrate and waste products)
- Western blot and densitometry for quantitation of desired product
Communication (CI)
There are several session throughout the semester which are entirely devoted to a written assignmnent, and the preparation and practice of an oral presentation.
The written assignment for course 10.28/10.28L is called Technical Analysis Paper (TAP). The TAP considers a current topic of intense interest to chemical-biological engineering researchers.
Lectures
- Technical: 1 hour/week, on average
- Communication: 1 hour/week, on average
Assignments
* one individual report called the Technical Analysis Paper (TAP). It is an original assignment, which the student develops incrementally over the entire term. The student is taught how to critique selected publications on a research topic chosen by the student to how to propose novel experiments and new directions. It is the major written Communication (CI) assignment and students receive significant teaching from both the technical and the communication staff on how to produce such a document.
Examples of Publications List and Potential Research Topics for the Technical Analysis Paper (TAP) Assignment
Upstream Processing and Protein Production
Example: “Production of Human a-1-Antitrypsin from Transgenic Rice Cell Culture in a Membrane Bioreactor” (McDonald et al.)
- Transgenic Plant Cell Culture Production of Recombinant Proteins
- Compartmentalized Bioreactors
Downstream Processing
Example: “Capture of Human Monoclonal Antibodies from Cell Culture Supernatant by Ion Exchange Media Exhibiting High Charge Density” (Necina et al.)
- Evolution of Monoclonal Antibody Purification
- Media Optimization in Bioprocessing
Instrumentation and Analytical Methods
Example: “Monitoring Green-Fluorescent Protein-Operon Fusion Protein Expression During High Cell Density Cultivation of Escherichia coli Using an On-line Optical Sensor” (DeLisa et al.)
- Genomic Analysis of High Cell Density E. coli Cultivations
- GFP as a Quantitative Reporter for Recombinant Protein Production
* short team technical memoranda for the Independent Project
* one individual oral presentation - all the students present towards the end of the term (CI requirement)
* a series of individual lab assignments on the material presented in lecture and in the manual
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