BUCHI Encapsulator B-395 Pro - Manual (Page 63)

6 Operation
63 B-395 Pro Operation Manual, Version C
6 .14 Encapsulation protocol for alginate-PLL-alginate membranes
The Alginate-Polylysine-Alginate-membrane is a well established encapsulation system for animal cells
first described by Lim and Sun
1 .Below is a well tried protocol.
Required solutions
1. 1.5 % Alginate solution: 1.5 % low viscositiy alginate in MOPS washing-buffer
adjust pH to 7.0 at 25°C
sterile filtration through a 0.2 µm filter
2. Polymerization solution: 10 mM MOPS (Morpholinopropanesulfonic acid)
100 mM CaCl
2
pH = 7.2 at 25°C
3. PLL solution: 0.05 % Poly-L-lysine MG 15’000-30’000 in MOPS washing buffer
4. MOPS washing buffer: 10 mM MOPS (Morpholinopropanesulfonic acid)
0.85 % NaCl
pH = 7.2 at 25°C
5. 0.03 % alginate solution: 2 mL of 1.5 % alginate solutio.
+ 98 mL MOPS washing-buffer.
6. Depolymerization solution: 50 mM Na
3
-Citrate
0.45 % NaCl
10 mM MOPS
pH = 7.2 at 25°C
For one encapsulation of 12 mL polymer-product mixture the following is needed:
• 12 mL 1.5 % alginate solution (sterile filtered)
• 100 mL 0.03 % alginate solution (not sterile)
• 225 mL Polymerization solution (not sterile)
• 75 mL PLL Solution (not sterile)
• 900 mL MOPS Washing buffer (not sterile)
• 200 mL Depolymerization solution (not sterile)
1 Lim F. and Sun A.M. 1980. Microencapsulated Islets as Bioartificial Pancreas. Science 210: p.908-910.