11. Place the RBC bag on the shaker platform, and secure with the
three shaker magnets. For optimum performance the shaker
should be placed at the same level as the machine. Unclamp the
ratchet clamp on the glycerol line.
12. Check to confirm that the set is installed properly and press
YES. After doing this, the machine guides the user through a
set of verification questions to ensure the proper loading of the
disposable set. Upon the question “Sterile dock RBC bag,”
open the weld between the RBC bag and the
glycerolization set.
13. The ACP215 prompts the operator to enter his/her initials (see
Note 2).
14. Press “Modify Program” and fill in the net weight of the RBC
unit using the YES/UP or NO/DOWN arrows.
15. Press START to begin the glycerolization process. The glycerol
solution will be added to the RBC at a computed flow rate
(see Note 3). The glycerol flow rate will progressively increase
during the process assuring that the osmolarity rate will not
increase more than 500 mOsm/kg.min.
16. When the computed volume of glycerol has been processed,
the pump stops. The shaker stops 30 s later. The message
“Glycerolization complete” will be displayed.
17. The printer will print out information related to the glycerolization procedure.
18. Heat-seal the tubing between the RBC bag and the
glycerolization set.
19. Detach the RBC bag from the rest of the glycerolization set.
20. Remove the disposable set from the machine and discard the
disposable set.
21. Sterilely dock a 1000 mL freezing bag to the bag containing
the glycerolized red blood cells. Leave about 10 cm of tubing
between the bags.
22. Open the weld and transfer the cells to the freezing bag by
gravity. Weigh and record the net weight of the unit.
3.1.4 Removal
of Supernatant Glycerol
Originally, the HGM described the removal of supernatant removal
before freezing [31]. However, recently it has been shown that
removal of supernatant glycerol before freezing is not necessary
[32]. To decrease the volume of the RBC unit to be frozen,
supernatant glycerol could be removed (see Note 4):
1. Sterilely dock a 500 mL transfer bag to the freezing bag. Do
not open the weld.
2. Spin the unit at 1248 Â g in a 22
C centrifuge for 10 min. To
minimize red cell mixing, the brake should be set low.
528
Johan W. Lagerberg
three shaker magnets. For optimum performance the shaker
should be placed at the same level as the machine. Unclamp the
ratchet clamp on the glycerol line.
12. Check to confirm that the set is installed properly and press
YES. After doing this, the machine guides the user through a
set of verification questions to ensure the proper loading of the
disposable set. Upon the question “Sterile dock RBC bag,”
open the weld between the RBC bag and the
glycerolization set.
13. The ACP215 prompts the operator to enter his/her initials (see
Note 2).
14. Press “Modify Program” and fill in the net weight of the RBC
unit using the YES/UP or NO/DOWN arrows.
15. Press START to begin the glycerolization process. The glycerol
solution will be added to the RBC at a computed flow rate
(see Note 3). The glycerol flow rate will progressively increase
during the process assuring that the osmolarity rate will not
increase more than 500 mOsm/kg.min.
16. When the computed volume of glycerol has been processed,
the pump stops. The shaker stops 30 s later. The message
“Glycerolization complete” will be displayed.
17. The printer will print out information related to the glycerolization procedure.
18. Heat-seal the tubing between the RBC bag and the
glycerolization set.
19. Detach the RBC bag from the rest of the glycerolization set.
20. Remove the disposable set from the machine and discard the
disposable set.
21. Sterilely dock a 1000 mL freezing bag to the bag containing
the glycerolized red blood cells. Leave about 10 cm of tubing
between the bags.
22. Open the weld and transfer the cells to the freezing bag by
gravity. Weigh and record the net weight of the unit.
3.1.4 Removal
of Supernatant Glycerol
Originally, the HGM described the removal of supernatant removal
before freezing [31]. However, recently it has been shown that
removal of supernatant glycerol before freezing is not necessary
[32]. To decrease the volume of the RBC unit to be frozen,
supernatant glycerol could be removed (see Note 4):
1. Sterilely dock a 500 mL transfer bag to the freezing bag. Do
not open the weld.
2. Spin the unit at 1248 Â g in a 22
C centrifuge for 10 min. To
minimize red cell mixing, the brake should be set low.
528
Johan W. Lagerberg