13
12
10. Place the flasks back onto the tray with the corner notch of each flask in the
upper right position once medium has completely drained into the harvest
bottle.
NOTE
: Corning® MicroDEN® flasks can be harvested individually by unclamping
one of the tubes from the pump to stop flow. Liquid will flow through the
clamped tube.
Cold Buffer Washes
1. Add 26 mL of cold buffer (4°C) to the left side inlet storage bottle.
◗
–
To unscrew the cap, turn the bottle while keeping the cap and tubing set
in place so the tubes don’t twist (Figure 1). Take care not to touch the dip
tubes to any non-sterile surface.
2. Insert tubing set and place the cap back onto the bottle. While pressing
down on the tubing set, rotate the orange cap to tighten the assembly to
the bottle (Figure 1). Place the bottle back into the thermoform tray.
3. Select “Harvest” using the function button on the Corning® MicroDEN®
unit
to perfuse cold buffer into the flasks.
4. Remove the flasks from the thermoform tray and slide the flasks side-to-
side as buffer fills the flasks (Figure 6).
5. Allow perfusion to continue when the inlet bottle is empty of buffer. Air will
perfuse into the flasks.
6. When air starts to perfuse, rotate the flasks so that the outlet port is down
and the inlet port is up (Figure 5). The corner notch of each flask should be
at the lower left position.
◗
–
Reversing flask orientation allows the liquid to drain into the harvest bottle.
7. Continue to slide the flasks side-to-side as liquid drains from the flasks into
the harvest bottle and air is perfused into the flasks.
8. Stop perfusion by pressing the MicroDEN
unit function button when buffer is
drained from the flasks.
9. Place the flasks back onto the thermoform tray with the corner notch of each
flask in the upper right position.
10. Repeat Steps 1-9 for a second cold buffer wash.
NOTE:
Flasks can be harvested individually by unclamping one of the tubes from
the pump to stop flow. Liquid will flow through the clamped tube.
Non-adherent and most adherent iDCs are transferred from the MicroDEN flasks
to the harvest bottle after two cold buffer washes. Additional washes can be
incorporated if the remaining strongly adherent cells are desired. Additionally,
the flasks, consumable kit, or the entire unit with consumable kit can be placed
on a platform shaker to aid detaching strongly adherent cells. Alternatively, the
flasks or consumable kit can be gently vortexed. Do not place the MicroDEN
unit
on the vortex.
The dendritic cell product is collected in the harvest bottle after washing
with cold buffer. The harvest bottle can then be removed from the rest of the
consumable by disconnecting the MPCs.
Helpful Harvest Tips
◗
◗
When beginning to perfuse liquid, or when the tubing is filled with air, angle
the left side inlet MPCs upwards so that air is completely removed from the
MPCs (Figure 4). If this is not performed, air will be trapped in the MPCs and
can cause an air bubble to perfuse into the flask.
◗
◗
When beginning to perfuse air, angle the left side inlet MPCs downwards so
that liquid is drained from the MPCs (Figure 7).
Disposing of Consumables
The entire consumable kit (thermoform tray, flasks, tubing, connectors, and bottles)
are designated as single-use and should be appropriately discarded after use.
NOTE:
Tubing
connections
not shown
Corner notch on
flask in lower
left position
Outlet ports
are down
Inlet ports
are up
Slide the flasks
side to side
Figure 6.
Slide the flasks side-to-side as medium drains into the harvest bottle.
Figure 7.
MPCs angled downward to drain liquid.
