BUCHI Encapsulator B-395 Pro User Manual

Page 58

6 Operation

B-395 Pro

Operation Manual, Version C

6 .10

Practicing with the Encapsulator, working with the complete reaction
vessel

After getting comfortable with Ca-alginate bead formation, perform test runs with the complete reac-
tion vessel to simulate sterile working conditions, but by using non-sterile alginate solution.
Sterile working conditions have the additional difficulty that it is often impossible to modify something
in the autoclaved reaction vessel without losing sterility, like if a piece was forgotten or assembled
in the wrong way. Therefore, it is important that the reaction vessel is properly prepared before
sterilization. Follow section 5 for the assembly and sterilization of the reaction vessel. If you change
something, note it in a separate procedure.

1. Prepare all the needed encapsulation reagents (as an example, for animal cell encapsulation, see

section 6.14).

For this run:

60 mL 1.5 % alginate solution

500 mL 100 mM CaCl

polymerization solution

600 mL 0.9 % NaCl + 10 mM CaCl

washing solution

Assemble a pressure bottle.

2. Take the autoclaved reaction vessel and attach it to the control unit.

Verify that the silicone tubing at the liquid drain port of the reactor base plate is closed with a
clamp. Make sure that the bead collecting valve at the reactor vessel is closed, that the magnetic
stirrer is below the reactor and that the stirrer bar is above the magnetic stirrer.

–

Connect the electrostatic dispersion unit with the red wire to the reaction vessel.

–

Move the collection cup of the bead bypass below the nozzle.

3. Place 500 mL of polymerization solution in the pressure bottle. Attach the silicone tubing to the

liquid membrane filter.
Connect the pressure bottle to the air outlet of the control unit. Switch on the control unit. Set the
air pressure at 0.3 to 0.7 bar (4 to 10 psi). When the desired amount of liquid is pumped, release
the air pressure.

NOTE
The amount of hardening solution should be 8 to 10 times the volume of the polymer mixture. The
polymerization solution should have a height of at least 2 cm (approx. ¾“) in the reaction vessel (a
minimum of 200 mL).

4. Set the vibration frequency, the electrostatic tension and – if the syringe pump is used – the

pumping rate to the appropriate values as previously determined. Set the speed of the magnetic
stirrer so that a vortex is just visible.

5. Fill a 60 mL syringe with the alginate/sample solution and attach it to the Encapsulator. Activate

the vibration control system and set the vibration frequency at values predetermined in section
6-9. Activate the syringe pump and set the pumping speed as previously determined. Press the
“turbo” button until a continuous liquid jet is formed. Activate the electrostatic dispersion system. If
needed, adjust the pumping speed and/or the frequency to obtain a clear bead chain down to the
collection cup.