Procedure number: 275
Page 1 of 2
(Blood bank name and address)
Subject: 40% Glycerol Citrate Freezing Solution
40% GLYCEROL CITRATE FREEZING SOLUTION
PURPOSE
To prepare 40% glycerol for use as a cryo-protective agent in the freezing and storage of red cells at
temperatures of –20oC or colder.
PRINCIPLE
Pure 99.9% glycerol or “glycerine” is diluted with distilled water in the presence of potassium citrate
monohydrate, sodium phosphate monobasic monohydrate, sodium phosphate dibasic anhydrous, and sodium
azide. This allows for incorporation of glycerol into the red cells without damage to their structural integrity.
POLICIES
1.
The solution should be clear. Do not use if it is cloudy, turbid, or discolored.
2.
Freezing solution outdates in six months from date prepared when stored at 2-6oC.
MATERIALS AND EQUIPMENT
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
12.
13
14.
Potassium citrate monohydrate (K3C6H5O7H2O)
Sodium phosphate monobasic monohydrate (NaH2PO4H2O)
Sodium phosphate dibasic anhydrous (Na2HPO4)
99.9 % pure Glycerol (Glycerin)
10% Sodium Azide (NaN3) stored in a brown bottle.
Distilled water
1000 mL graduated cylinder
Pipette capable of accurately measuring 1 mL.
1000 mL beaker
2, 500 mL storage bottles with tight caps.
Magnetic stirrer with stirring bar or glass stirring rod
Balance sensitive to 0.1 gm
Weighing dishes or weighing (slick-surfaced) paper
Reagent Preparation Logbook
PROCEDURE
1.
Measure 600 mL of distilled water in the 1000 mL graduated cylinder and pour it into the 1000 mL beaker.
2.
Weigh out the following into a weighing dish or weighing paper, and pour each into the water-filled beaker:
A. 19.4 gm of potassium citrate monohydrate (K3C6H5O7H2O)
B.
2.4 gm sodium phosphate monobasic monohydrate (NaH2PO4H2O)
C.
2.8 gm sodium phosphate dibasic anhydrous (Na2HPO4)
3.
Stir until dissolved and well mixed on a magnetic stir plate or using a clean glass rod.
4.
Measure 400 mL of 99.9% glycerol (glycerin) into the 1000mL graduated cylinder and transfer it
to the beaker.
Procedure number: 275
Page 2 of 2
(Blood bank name and address)
Subject: 40% Glycerol Citrate Freezing Solution
5.
Using the pipette measure 1 mL of 10% sodium azide and add it to the beaker.
6.
Stir until well mixed using the magnetic stir plate or clean glass rod.
7.
Pour the 40% glycerol solution into the two 500-mL bottles.
8.
Prepare labels with the following information and place them on the bottles:
A.
B.
C.
D.
E.
F.
G.
9.
Chemical name: “40% Glycerol citrate”
Common name: “40% Glycerol freezing solution”
Manufacturer of the glycerol
“Store at 2 – 6oC”
Date prepared
Expiration date [6 months from preparation date]
Initials of the individual preparing the solution
Store at 2-6C.
RECORD KEEPING
Record the reagent preparation information in Reagent Preparation Log.
REFERENCES
Adapted from the Blood Center of Wisconsin Immunohematology Reference Laboratory procedure manual.
Adopted
[initials]
[date]
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed
Reviewed