Flexcell Tissue Train Culture System

FLEXCELL ® INTERNATIONAL CORPORATION
1. INTRODUCTION
Flexcell ® ’s Tissue Train ® culture system
utilizes a novel method to culture and
mechanically load cells in a three-dimensional
(3D) hydrogel or cell-assembled matrix.
Mechanical load may be applied using the
Flexcell ® Tension System with a user defined
program. Cell-gel constructs or cell-assembled
matrices may be mechanically loaded
uniaxially or equibiaxially using special
Arctangle ® or cylindrical Loading Stations ,
respectively. A program defining the %
elongation, frequency and duration of
mechanical load can be regulated by the user
in the FX-5000 FlexSoft ® software.
Flexcell’s Tissue Train ® Culture System can
be used as a stand-alone device to create
three-dimensional geometries (i.e., linear,
trapezoidal, or circular) for cell culture in a
matrix gel or allow the cells to build a selfassembled
matrix that connects to the anchors
in the Tissue Train ® culture plate. The
investigator can also apply regulated strain to
the growing tissues. A user can define a
frequency, elongation and duration of strain in
a regimen that simulates the strain
environment of a tissue in the body. The strain
regimen is controlled by the Flexcell Tension
System (see Part B, pg. 7) that can apply static
or cyclic equibiaxial or uniaxial strain. The
following is a method by which to culture
cells in a 3D matrix.
2. METHOD TO CREATE 3D CELL-SEEDED HYDROGEL
2.1 PREPARATION OF CELLS IN 3D
LINEAR OR TRAPEZOIDAL GELS IN A
TISSUE TRAIN ® CULTURE PLATE
1. Prepare cells according to the laboratory’s
established protocol for primary cultures
or continuous cell lines in the medium of
choice.
2. Release cells from their substrates with
0.05% trypsin, trypsin-EDTA, 0.05%
bacterial collagenase or other means. Add
serum containing media to the cells to
neutralize the trypsin or collagenase.
3. Count cells and determine the number of
cells needed:
Linear gel: approximately 50,000-
200,000 cells in 150-200 L for each
well of a 6-well Tissue Train ® culture
plate.
Trapezoidal gel: approximately
700,000 cells in 700 l for each well
of a 6-well Trapezoidal Tissue Train ®
culture plate.
NOTE: Cell seeding density will vary depending
on cell type. We recommend testing cell seeding
densities to determine the best cell number for
your application and cell type.
4. Wash cells 2 times with medium to
remove trypsin or collagenase.
5. Before adding cells, the matrix protein gel
solution should be neutralized to pH 7.0
using 1 M sodium hydroxide.
6. Cells may be reconstituted in one volume
of media containing 10% fetal calf serum,
70% matrix protein gel fluid and 20% 5X
MEM. The objective is to achieve an
overall gel-MEM concentration of 1X.
The suggested formula for the cell/matrix
protein gel combination is as follows:
2