eBook: Optimizing Cell Culture Techniques - 22

Select the custom program 'M_ASP_384_MAN'. The
pipette aspirates 70 µl at speed 1. This low speed
minimizes the shearing force that the spheroid
could be subjected to. The aspiration volume is set
higher than the volume of the well (50 µl) to ensure
complete removal of the medium. The special design
of the well protects the spheroids and leaves
a defined residual volume of 2-3 µl. A prompt on
the pipette screen informs the user that the medium
needs to be purged to the waste reservoir or
into a collection plate. The last step in the program
is a loop, which means that the required pipetting
steps can be easily repeated for multiple plates
without the need to restart the program. Abort the
program, eject, and discard the tips once all the
plates have been processed.
4. Medium dispensing - Akura 384
Fresh medium is dispensed into two Akura 384
plates from the reservoir
Discard the used tips and load new 125 µl sterile,
filter GRIPTIPS onto the VIAFLO 384. Place an automation
friendly reagent reservoir with fresh medium
on position B. Allow up to 10 ml of extra medium
to prevent aspiration of air.
Place an Akura 384 plate onto an Akura 384 lid in
position A. Adjust the plate manually towards the
user ('Bottom of plate') to create a small horizontal
offset (1-2 mm). Select and run the custom program
'M_DISP_384_MAN'. Move the pipette head to the
reservoir on position B. The pipette aspirates 110 µl
fresh medium, which is sufficient to fill two plates
with 50 µl medium plus 10 µl of excess volume.
Move the pipette head to position A and gently
immerse the pipette tips into the wells of the Akura
384 plate until the minimum height is reached
(Figure 4b). The pipette dispenses 50 µl medium
into each well at slow speed. The dispensing step
Figure 5. Nine consecutive medium exchanges were performed
with InSphero's Human Islet Spheroids in Akura 384 plates using
INTEGRA's VIAFLO 384. Three independent experiments aspirating
>90 % of the medium showed <1 % spheroid loss.
Remarks
Partial plates: Programs can be adapted to a
different number of samples at any time, giving
laboratories total flexibility to meet current and
future demands.
is repeated once more for a second well plate. The
pipette will prompt the user to purge the medium
that is remaining in the tip. The dispensing process
for two plates is repeated up to 100 times due to a
loop at the end of the program.
Tip: Z-heights may vary from system to system,
depending on the calibration. Check the correct
height using a dummy plate. Pipette tips need to be
positioned 0.2-0.3 mm above the spheroid. Lower
pipette positioning could damage the spheroids.
Three independent experiments showed <1 %
spheroid loss from Akura 384 plates during nine
consecutive medium exchanges (Figure 5).
22
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