Biomedical Engineering Reference
In-Depth Information
54.
Protein Expr Purif
. 2003 Jun;
29(2):311-20. A less laborious
approach to the high-throughput
production of recombinant
proteins in
Escherichia coli
using
2-liter plastic bottles. Millard CS,
Stols L, Quartey P, Kim Y,
Dementieva I, Donnelly MI.
Environmental Research
Division, Argonne National
Laboratory, Bldg. 202/Rm.
BE111, 9700 South Cass Avenue,
Argonne, IL 60439.
Contemporary approaches to biology often call for the
high-throughput production of large amounts of
numerous proteins for structural or functional
studies. Even with the highly efficient protein
expression systems developed in
Escherichia coli
,
production of these proteins is laborious and time
consuming. We have simplified established protocols
by the use of disposable culture vessels: common 2 L
polyethylene terephthalate beverage bottles. The
bottles are inexpensive, fit conveniently in commonly
available flask holders, and, because they are
notched, provide sufficient aeration to support the
growth of high-density cultures. The use of
antibiotics and freshly prepared media alleviates the
need for sterilization of media and significantly
reduces the labor involved. Uninoculated controls
exhibited no growth during the time required for
protein expression in experimental cultures. The
yield, solubility, activity, and pattern of
crystallization of proteins expressed in bottles were
comparable to those obtained under conventional
culture conditions. After use, the bottles are
discarded, reducing the risk of cross-contamination
of subsequent cultures. The approach appears to be
suitable for high-throughput production of proteins
for structural or functional studies.
55.
Biotechnol Prog
. 2003 Jan-Feb;
19(1):2-8. Fluid mechanics, cell
distribution, and environment in
CellCube bioreactors. Auniņs JG,
Bader B, Caola A, Griffiths J,
Katz M, Licari P, Ram K, Ranucci
CS, Zhou W. Merck Research
Laboratories, West Point,
Pennsylvania.
Cultivation of MRC-5 cells and attenuated hepatitis A
virus (HAV) for the production of VAQTA, an
inactivated HAV vaccine (1), is performed in the
CellCube reactor, a laminar flow fixed-bed bioreactor
with an unusual diamond-shaped, diverging-
converging flow geometry. These disposable
bioreactors have found some popularity for the
production of cells and gene therapy vectors at
intermediate scales of operation (2, 3). Early testing
of the CellCube revealed that the fluid mechanical
environment played a significant role in nonuniform
cell distribution patterns generated during the cell
growth phase. Specifically, the reactor geometry and
manufacturing artifacts, in combination with certain
inoculum practices and circulation flow rates, can
create cell growth behavior that is not simply
explained. Through experimentation and
computational fluid dynamics simulations, we can
account for practically all of the observed cell growth
behavior, which appears to be due to a complex
mixture of flow distribution, particle deposition
under gravity, fluid shear and, possibly, nutritional
microenvironment.
