Platelets can be stored at room temperature for up to 7 days. Critical factors affecting platelet storage include temperature, availability of metabolic fuels like glucose and oxygen, and respiratory capacity. Over time in storage, platelets undergo a storage lesion characterized by shape change, activation, decreased function. Tests are used to monitor storage lesions and ensure platelet quality is maintained over storage duration. Platelet additive solutions provide synthetic alternatives to plasma as storage media and have advantages but also risks like lower efficacy and enhanced bacterial growth.

1. Platelet preservation and storage
Presented by: Manuru Shekhar Sameer Moderated by: Dr Kshitija Mittal

2. Room temperature storage
 Room temperature storage:
• Slow metabolic activity by 60% compared with in vivo
activity at 37o C.
• Deceased in vitro aging
• Slower accumulation of toxic metabolites within platelet
storage container.
• Seven days of storage lead to only three days worth of in vivo
senescent loss.

3.  There is a transient reversible loss of function in vivo that
recovers only after a variable period of time in circulation.
This prevents spontaneous hemorrhage in chronically
thrombocytopenic transfusion recipients as the platelets do not
get immediately activated.

4. Critical variables affecting stored
platelet health
1. Temperature
2. Metabolic fuel availability
3. Respiratory capacity.

5. Platelets are exposed to the cold
Plasma membranes undergo phase transitions
Coalescence of lipid rafts
Shape change
Activation priming
Clustering of neoepitope-expressing glycoprotein (GP)-Ibα domains
with enhanced von Willebrand factor (vWF) binding capacity
Acceleration of apoptosis
Effect of temperature on platelet membrane

6. Ref: Rossi’s Principles of Transfusion Medicine
6th edition
Effect of various temperatures on T1/2 of the platelets
Temp(inoC) Half Life(in
days)
4 1
13 2
20 4
Between 22-30 5

7. Holme et al compared
Temperature of storage of
platelets
Number of hours after which
platelet circulatory capacity
was lost in vivo
18 ° C 24 hours
16 °C 16 hours
12 °C 10 hours
4 °C 6 hours

8.  Stored platelets must generate adenosine triphosphate (ATP), the cell’s
principal “energy currency.”
 This ATP is derived from either aerobic or anerobic metabolism
Metabolic fuel availability

9. 1)Aerobic Metabolism:
•Main source of energy
•85% of platelet energy is derived from aerobic metabolism in
continuously agitated PCs
•Occurs through citric acid cycle and oxidative
phosphorylation
•More efficient energy source as 36 moles of ATP are
produced per mole of glucose consumed.

10. Aerobic Metabolism
Metabolic fuel availability

11. Metabolic fuel for aerobic metabolism
 Free fatty acids derived from plasma triglycerides
 Amino acid glutamine.
 Acetate: also carries an advantage of having a buffering
action by consuming hydrogen ion during its metabolism.

12. 2) Anerobic metabolism:
• Down regulation of pyruvate dehydrogenase in stored
platelets
• Less efficient as number of ATP produced are 2 per mole of
glucose consumed
• Pyruvic Acid Lactic Acid
• Lactic acid + Bicarbonate CO2 + H2O
( buffer)
• CO2,diffuses through the storage container wall

13. Anaerobic
Metabolism

14. Glucose
Lactic Acid
With progressive increase in lactic acid production
Lactic Acid Bicarbonate
Gets accumulated Gets exhausted
Lack of lactic acid buffering
Decrease in pH
Metabolic fuel for anaerobic metabolism

15. Under low oxygen conditions
Glycolysis is upregulated by six to seven fold (Pasteur effect)
Increase in lactic acid production
Exacerbate plasma buffer exhaustion
Deleterious decline in pH
 Platelets begin to suffer:-
• Deleterious effects below a pH of 6.5–6.8 at room temperature (22 °C)
• Irreversibly damage in a pH below 6.2 at 22 °C.

16. Respiratory capacity
 Maintenance of aerobic metabolism requires a PO2 above 5–10
mmHg
 Oxygen consumption is constant per platelet and the presence
of a high total platelet bag content requires a storage
container having a high oxygen permeability.
Oxygen Delivery to the platelets depends upon:
 Storage container: •gas permeability
•wall thickness
•volume-to-surface area
 Continuous agitation: which promotes gas diffusion and
helps maintain normal platelet oxygen consumption.

17. Increase oxygen consumption by platelet concentrate
Exceeds the ability of the container to transport oxygen
PO2 will decline to hypoxic levels
Up regulation of glycolysis
Lactic acid accumulates
Deleterious decrease in pH
Importance of oxygen permeability

18.  O2 permeability is more important than CO2 permeability
 Kilkson et al. demonstrated a little change in pH when carbonic acid was
formed in a least permeable container.
 It is the lactate concentration which was the principal determinant of pH.
 High CO2 permeability Rapid diffusion of CO2 from the container Fall
in H+ ion concentration Increase in pH Cellular damage.
 A pH >7.4 at 22 °C has been associated with cellular damage.
 Hence, high CO2 permeability is not as critical as high oxygen permeability.
Why CO2 permeability is not critical

19.  First generation containers:
 Platelet storage of lower content PCs for 3 days
 Made up of Polyvinyl chloride (PVC) plasticized with di- ( 2- ethylhexyl) phthalate
(DEHP)
 Second- generation containers:
 Provide twice the oxygen permeability as of first generation
 Can store platelets for: 7 days
 Made up of •Polyolefin or
•Ethylene- vinyl acetate (EVA), or
•PVC plasticized with triethyl hexyl trimellitate (TOTM) and
butyryl- tri hexyl citrate (BTHC).

20. Agitation
 Prevent settling and contact mediated switch from oxidative to glycolytic
metabolism.
 Facilitate oxygen utilization by platelet mitochondria
 Supports platelet metabolism by ensuring effective exchange of oxygen,
carbon dioxide, and lactic acid between the platelets and the suspending
media
 Note: Agitation may be safely interrupted for 24 hours without lactate
accumulation and subsequent fall in pH.

21.  Various platelet rotators and agitators are:
• Ferris wheel and elliptical rotators
• Face-over-face tumblers (3–6 rpm)
• Flatbed platform shakers (with 1”-1.5” strokes at 50–70 cpm)
 In a lower content platelet product, PO2 can be maintained without
agitation. Mixing of product even once a day could prevent
injurious pH decrements.

22. Platelet Storage Lesions
Definition:
• The loss of platelet quality during storage is known as
platelet storage lesion.
• Include progressive decline in function accompanied by
characteristic morphologic changes.

23. Reduced oxygen tension(pO2)
Increase in rate of glycolysis in compensatory to decrease in ATP generation
from oxidative meatabolism
Increase in glucose consumption
Increase in lactic acid
Initially buffered with bicarbonate
Bicarbonate gets depleted
pH decreases
Platelets swell and there is disc to sphere transformation
Platelets become irreversibly swollen, aggregate together, lyse and when
infused will not circulate or function

25. Tests to assess platelet storage lesions
Platelet Storage
Lesions
Platelet structure
•Visual inspection
for swirling
phenomena
•Platelet
morphology by
microscopy
Metabolic status
•Supernatant pH,
pO2, pCO2, HCO3
•Glucose
consumption
•Lactate production
Platelet activation
•P-selectin (CD62P)
surface expression
•Soluble P-selectin
in supernatant
•Platelet factor 4
and β-
thromboglobulin
•Annexin V binding
•Lactate
dehydrogenase in
supernatant
•Platelet
microparticle
formation
Functional tests
•Platelet
aggregation,
spontaneous and to
agonists
•Hypotonic shock
response
•Extent of shape
change
•Thrombin-
stimulated ATP
release

26. Visual inspection for swirling
phenomena
Score 0: homogenous turbid and not change with pressure.
Score 1: homogenous swirling in some part of the bag and not clear
Score 2: clear homogenous swirling in all part of the bag
Score 3: very clear homogenous swirling in all part of the bag.
Swirling is lost when the platelets are exposed to cold, bacterial
contamination and low pH.
(Singh et al. quality assessment of platelets. Asian J Transf Sci. 2009;
3:88)

27. Platelet morphology by microscopy
Shape changes from:
discs spheres dendritic ballooned
Assessed by Kunicki morphology scoring system:
• 4 points to discs
• 2 points to spheres
• 1 point to dendritic forms
• none to ballooned platelets

28. Metabolic status
Characteristic Change Observed
pO2 Decreases
Glucose consumption Increases
Lactic Acid Increase
Bicarbonate Decrease
pH Decrease
ATP Decrease
Sequence
of
changes

29. Platelet activation
 Surface expression of P selectin(CD62P)
 Soluble P-selectin in supernatant
 Release of platelet factor 4 and β-thromboglobulin due to
degranulation
 Surface expression of negatively charged phospholipids

30. Functional assay
 Hypotonic shock response:
• To assess platelet viability
• Also known as osmotic reversal reaction
• Platelets are exposed to hypotonic environment.
• This results in initial swelling followed by a gradual decline as the
platelets resume a baseline size.
• This is measured with a spectrophotometer.

31. Aggregometry:
• To measure the decrease in light transmission after ADP stimulation
of stirred PRP.
• Aggregation occurs only by discord form of platelets.
• When agonist is added normal discoid platelets aggregate
absorb less light transmission increases detected by the
photocell.

32. Platelet additive solutions
• These are synthetic storage solutions

33. Advantages of Using Platelet Additive
Solutions
 Saves plasma for other purposes (e.g., transfusion or fractionation)
 Facilitates ABO-incompatible platelet transfusions
 Lower titers of anti-A and B antibodies.
 Reduces plasma-associated transfusion side effects, such as febrile
and allergic reactions, and may reduce risk of transfusion-related
acute lung injury (TRALI)
 Improves effectiveness of photochemical pathogen reduction
technologies
 Potentially improves bacterial detection

34. Constituents of PAS:
Constituent in PAS Function
NaCl( common to all PAS) •Osmotic neutrality
Glucose •Difficult to add because caramelizes
upon heat
sterilization
•Obtained from plasma as 20–40%
plasma
carryover
Acetate •Fuel for TCA cycle;
•Has lactate buffering function
Citrate •Maintains anticoagulation
•Upregulates glycolysis and renders
platelets •more susceptible to activating
stimuli
•Hence lower concentrations used
•Main constituent in PRP-PC and BC-PC

35. Phosphate •Acts as buffer
•Maintains adenine nucleotide levels
•Main constituent in apheresis platelets
•Upregulates glycolysis hence more plasma
carryover is required.
Magnesium and potassium •Reduce platelet activation
•Down regulates glycolysis

36.  Amicus cell separator allows addition of PAS and thereby facilitates
the use of photochemical pathogen reduction technology.
 Trima cell separator has pathogen reduced platelets in plasma.

37. International Council for Commonality in Blood
Banking Automation (ICCBBA) Designations
for Platelet Additive Solution Products
Gives a common nomenclature
To ensure unambiguous labeling PAS
The nomenclature has the format PAS-X, where X is an alpha
character
Sodium Chloride and Bicarbonate are not listed as they are
common in many PAS.

38. New
Name
Citrate Phosphate Acetate Magnesium Potassium Gluconate Glucose Trade name
PAS-A    PlasmaLyte A
(Fenwal/Baxter)
PAS-B   T-Sol,
PASII, or SSP
(Fenwal/Baxter)
PAS-C    PAS III or Intersol
(Fenwal/Baxter)
PAS-D      Composol
(Fresenius)
PAS-E      PAS IIIM or SSP+
(MacoPharma)
PAS-F    
PAS-G       (No Trade Name)
(Pall)

39. Disadvantage of Using Platelet
Additive Solutions
Lower corrected count increments after transfusion.
Enhance bacterial growth by reducing bactericidal properties
of plasma.