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RED BLOOD CELL AND

PLATELET PRESERVATION

RED BLOOD CELL BIOLOGY AND
PRESESERVATION

RBC metabolism .Three areas of RBC biology are crucial for normal erythrocyte survival and function: 1. Hemoglobin structure and function 3. Normal chemical composition and structure of the RBC membrane 2.

is located and limited to the cytoplasmic surface of the membrane forming the RBC cytoskeleton . are arranged in a bilayer structure comprising the framework in which globular proteins traverse and move. the main lipid components of the membrane. called peripheral proteins.RBC MEMBRANE  The RBC membrane represents a semipermeable lipid bilayer supported by a meshlike protein cytoskeleton structure  Phospholipids. a second class of membrane proteins. Proteins that extend from the outer surface and span the entire membrane to the inner cytoplasmic side of the RBC are termed integral membrane proteins. Beneath the lipid bilayer.

and HCO3-) but is relatively impermeable to cations such as sodium (Na+) and potassium (K+).  Accumulation or increase in deposition of membrane calcium also results. a loss of membrane deformability. PERMEABILITY  The RBC membrane is freely permeable to water and anions (Cl.DEFORMABILITY  The loss of adenosine triphosphate (ATP) (energy) levels leads to a decrease in the phosphorylation of spectrin and. . in turn. causing an increase in membrane rigidity and loss of pliability.

Metabolic pathways  The RBCs metabolic pathways that produce ATP are mainly anaerobic. because function of the RBC is to deliver oxygen not to consume it.  RBC metabolism may be divided into the Anaerobic Glycolytic Pathway and three ancillary pathways that serve to maintain the structure and function of hemoglobin: the Pentose Phosphate Pathway. the Methemoglobin Reductase Pathway. and the Luebering-Rapoport shunt. .

EMBDEN-MEYERHOF PATHWAY  Major pathway generating energy for RBCs  90% glycolysis. aerobic pathway  Provides reduced glutathione to prevent denaturation of LUEBERING-RAPOPORT PATHWAY  Generates 2.3-DPG (2.↓ Hgb affinity for Oxygen METHEMOGLOBIN REDUCTASE PATHWAY  Maintains Hgb iron in ferrous state to be functional .↑ Hgb affinity for Oxygen  ↑ 2.3-DPG .3diphosphoglyserate) that regulates the Hgb affinity for oxygen  ↓ 2.3-DPG . anaerobic pathway  2 ATP is produced for every glucose broken down to lactic acid PENTOSE PHOSPHATE PATHWAY  10% glycolysis.

3-DPG  ↑ 2.HEMOGLOBIN OXYGEN DISSOCIATION CURVE SHIFT TO THE LEFT SHIFT TO THE RIGHT  ↑ Hb affinity for Oxygen  ↓ Hb affinity for Oxygen  ↓ delivery of Oxygen to tissues  ↑ Delivery of Oxygen to tissues Factors: Factors:  ↑ pH  ↓ pH  ↓ 2.3-DPG  ↓ CO2  ↑ CO2  ↓ Temperature  ↑ Temperature .

RBC PRESERVATION  The goal of blood preservation is to provide viable and functional blood components for patients requiring blood transfusion. blood is stored in the liquid state between 1°C and 6°C for a specific number of days.  RBC viability is a measure of in vivo RBC survival following transfusion. as determined by the preservative solution(s) used.  The loss of RBC viability has been correlated with the lesion of storage.  To maintain optimum viability. which is associated with various biochemical changes .

RBC STORAGE LESION CHARACTERISTIC CHANGE OBSERVED % viable cells Decreased Glucose Decreased ATP Decreased Lactic Acid Increased pH Decreased 2.3-DPG Decreased Oxygen Dissociation Curve Shift to the Left Plasma K+ Increased Plasma Hemoglobin Increased .

for improved survival of RBCs (extends shelf life from 21 to 35 days) X .ANTICOAGULANT PRESERVATIVE SOLUTIONS CHEMICALS IN ANTICOAGULANT SOLUTIONS CHEMICAL FUNCTION PRESENT IN ACD-A CPD CP2D CPDA-1 Citrate Chelates Ca++.3-DPG X X X X Dextrose Substrate for ATP production (cellular) energy X X X X Adenine Production of ATP. necessary for maintenance of adequate levels of 2. prevents clotting X X X X Monobasic Sodium Phosphate Maintains pH during storage.

APPROVED ANTICOAGULANT PRESERVATIVE SOLUTIONS ABBREVIATIO N STORAGE TIME (DAYS) ACD-A 21 days CPD 21 days Citrate-Phosphate Double Dextrose CP2D 21 days Citrate-Phosphate-Dextrose-Adenine CPDA-1 35 days Citrate-Phosphate-Dextrose-Adenine CPDA-2 42 days NAME Acid Citrate Dextrose (formula A) Citrate-Phosphate Dextrose .

ADDITIVE SOLUTIONS  Additive solutions (AS) are preserving solutions that are added to the RBCs after removal of the plasma with or without platelets.  These solutions reduce hematocrits from around 70% to 85% to around 50% to 60%. . AS-1 and AS-5 also contain mannitol.  One of the reasons for their development is that removal of the plasma component during the preparation of RBC concentrates removed much of the nutrients needed to maintain RBCs during storage.  Additive solutions are approved for 42 days of storage. Adenine and Glucose. while AS-3 contains citrate and phosphate for the same purpose. which protects against storagerelated hemolysis.  Additive solutions consists of: Saline.

 Frozen RBCs can be stored for 10 years from the date of freezing. . frozen RBCs may be stored up to 10 years before thawing and transfusion.  Cryoprotective agents such as glycerol are used to prevent rupture of RBCs during freezing. that is. Autologous transfusion (auto meaning “self”) allows individuals to donate blood for their own use in meeting their needs for blood transfusion.FREEZING AND REJUVENATION  RBC freezing is primarily used for autologous units and the storage of rare blood types.  Two concentrations of glycerol have been used to freeze RBCs: a high-concentration glycerol (40% weight in volume [w/v]) and a low-concentration glycerol (20% w/v) in the final concentration of the cryopreservative.

Advantages of High-Concentration Glycerol Technique Used by Most Blood Banks Over LowConcentration Glycerol Technique ADVANTAGE HIGH GLYCEROL LOW GLYCEROL –80°C –196°C No Yes Mechanical Liquid Nitrogen Maximum storage temperature –65°C –120°C Shipping requirements Dry ice Liquid Nitrogen Can be thawed and frozen Critical Initial freezing temperature Need to control freezing rate Type of freezer Effect of changes in storage temperature .

Advantages and Disadvantages of RBC Freezing ADVANTAGES DISADVANTAGES Long-term storage (10 years) A time-consuming process Maintenance of RBC viability and function Higher cost of equipment and materials Low residual leukocytes and platelets Storage requirements (–65°C) Removal of significant amount of plasma proteins Higher cost of product .

and adenine. inosine. pyruvate.RBC REJUVENATION  Rejuvenation of RBCs is the process by which ATP and 2. depending on RBC preservative solutions used.  Rejuvesol is currently approved for use with CPD. . and CPD/AS-1 RBCs. Rejuvesol (enCyte Systems) is the only FDA approved rejuvenation solution sold in the United States.3-DPG levels are restored or enhanced by metabolic alterations.  It contains phosphate.  RBCs stored in the liquid state can be rejuvenated at outdate or up to 3 days after outdate.  Currently. CPDA1.

Development of methods to produce RBCs through bioengineering (blood pharming) 5. Development of RBC substitutes . Development of procedures to convert A-.Current Trends in RBC Preservation Research Research and development in RBC preparation and preservation is being pursued in five directions: 1. and Abtype RBCs to O-type RBCs 4. Development of procedures to reduce and inactivate the level of pathogens that may be in RBC units 3. B-. Development of improved additive solutions 2.

PLATELET PRESERVATION .

initial arrest of bleeding by platelet plug formation and 2.PLATELETS  Disk-shaped cells derived from the megakaryocytes found within bone marrow.  Platelets are intimately involved in primary hemostasis. . maintenance of vascular integrity.  The role of platelets in hemostasis includes: 1. stabilization of the hemostatic plug by contributing to the process of fibrin formation 3.  2-4 µm in diameter  Life span: 9-12 days  Approximately 1/3 of the platelets released from the bone marrow are sequestered in the spleen and the remaining 2/3 are found in the circulation. which is the interaction of platelets and the vascular endothelium in halting and preventing bleeding following vascular injury.

Adenosine triphosphate (ATP) 3. Fibronectin Dense granules contain nonprotein factors including: 1. von Willebrand’s factor (factor VIII:R) 4. β-Thromboglobulin (BTG) 5. or serotonin) 4. Platelet fibrinogen 2. 5-Hydroxytryptamine (5-HT. Calcium . including: .PLATELETS  Glycogen granules provide energy substrate.  Alpha (α) granules contain contact-promoting factors. 1. Platelet factor 4 (heparin-neutralizing) 6. Platelet-derived growth factor (PDGF) 3. Adenosine diphosphate (ADP) 2.

PLATELET STORAGE LESION CHARACTERISTIC CHANGE OBSERVED Lactate Increased pH Decreased ATP Decreased Morphology scores change from discoid to spherical (loss of swirling effect) Decreased Degranulation Increased Platelet Activation Markers Increased Platelet Aggregation Drop in responses to some agonists .

 Platelets are also utilized in some instances to treat other disorders in which platelets are qualitatively or quantitatively defective because of genetic reasons.CLINICAL USE OF PLATELETS  Platelet components are effectively used to treat bleeding associated with thrombocytopenia. a marked decrease in platelet number. .  Platelets are also transfused prophylactically to increase the circulating platelet count in hematologyoncology thrombocytopenic patients to prevent bleeding secondary to drug and radiation therapy.

 Platelet concentrates can also be stored at 1°C to 6°C for 48 hours without agitation .STORAGE CONDITIONS  Whole blood derived platelet apheresis are stored at 20-24degC with continuous agitation for up to 5 days.

Donor has bacterial infection 3.culture based system 3. Sources of bacterial contamination: 1.culture based system 2.is a laser-based. eBDS (Pall Corp. BacT/ALERT (bioMérieux) . Environmental contamination during storage and processing 3 commercial systems approved for screening platelets for bacterial contamination: 1. Contamination at the phlebotomy site 2. Scansystem (Hemosystem) .PLATELET STORAGE AND BACTERIAL CONTAMINATION  .) . scanning cytometry method .

 Following pretreatment. . the sample is loaded into a disposable plastic cartridge with built-in controls that turn from yellow to blueviolet when the test is ready to be read. A pink-colored bar in either the grampositive or gram-negative test window indicates a positive result.Disadvantages of Culture Methods  Product loss due to sampling  Delay in product release. in approximately 20 minutes. further reducing already short shelf-life  False-negative results  Cost  Logistical problems of culturing WBD platelets Pan Genera Detection (PGD) test (Verax Biomedical)  first rapid test approved to detect bacteria in WBD platelets  An immunoassay that detects lipoteichoic acids on grampositive bacteria and lipopolysaccharides on gram-negative bacteria.

CURRENT TRENDS IN PLATELET PRESERVATION RESEARCH Research and development in platelet preservation is being pursued in many directions. also termed synthetic media 3. Development of platelet substitutes 5. New approaches for storage of platelets at 1°C to 6°C 6. The development of processes to cryopreserve platelets . Development of methods that would allow platelets to be stored for 7 days 2. Development of procedures to reduce and inactivate the level of pathogens that may be in platelet units 4. including the following: 1. Development of additive solutions.