Goals Of Blood Collection Maintain viability and function Prevent physical changes Minimize bacterial contamination

Anticoagulants Preservative Solutions Anticoagulants prevent blood clotting Preservatives provide nutrients for cells Heparin Rarely if ever used anymore Anticoagulant ONLY Transfuse within 48 hours, preferably 8

Anticoagulants CPD CPD-A1 Storage time 21 days 35 days Temperature Slows glycolytic activity Adenine None Substrate for ATP synthesis Volume 450 +/- 10% Dextrose Supports ATP generation by glycolytic pathway Citrate Prevents coagulation by binding calcium

Additive Solution Primary bag with satellite bags attached. One bag has additive solution (AS) Unit drawn into CPD anticoagulant

Additive Solution Add additive solution to RBCs, ADSOL, which consists of: Saline Adenine Glucose Mannitol Extends storage to 42 days Final hematocrit approximately 66%

Changes Occur During Storage Shelf life = expiration date At end of expiration must have 75% recovery At least 75% of transfused cells remain in circulation 24 hours AFTER transfusion

Storage Lesion Biochemical changes which occur at 1-6C Affects oxygen dissociation curve, increased affinity of hemoglobin for oxygen. Low 2,3-DPG, increased O2 affinity, less O2 released. pH drops causes 2,3-DPG levels to fall Once transfused RBCs regenerate ATP and 2,3-DPG Few functional platelets present Viable (living) RBCs decrease

Plasma hemoglobin Plasma K+ Na+ K+ Viable cells pH ATP 2,3-DPG Plasma Na+ pH maintained by the phosphate Helps release oxygen from hemoglobin (once transfused, ATP & 2,3-DPG return to normal)

Storage Lesion Significant for infants and massive transfusion. Other biochemical changes ATP decreases Potassium increases Sodium decreases Plasma hemoglobin increases

Preparation of Components Collect unit within 15 minutes to prevent activation of coagulation system Draw into closed system – primary bag with satellite bags with hermetic seal between. If hermetic seal broken transfuse within 24 hours if stored at 1-4C, 4 hours if stored at 20-24C

Preparation of Components Centrifuge – light spin, platelets suspended Remove platelet rich plasma (PRP) Centrifuge PRP heavy spin Remove platelet poor plasma Freeze plasma solid within 8 hours Thaw plasma at 1-4C – precipitate forms Centrifuge, express plasma leaving cryoprecipitate. Store both at -18C RBCs – CPD – 21 days, ADSOL – 42 days – 1-6C

Preparation of Components Summary – One unit of whole blood can produce: Packed RBCs Fresh frozen plasma (FFP) Cryoprecipitate (CRYO) Single donor plasma (SDP) – cyro removed Platelets – terms PC (platelet concentrate) OR RD PC (random donor platelet concentrate)

Blood Component General Information Transfusion practice Transfusion requires doctor’s prescription All components MUST be administered through a filter Infuse quickly, within 4 hours D (Rh) neg require D neg cellular products ABO identical preferred, ABO compatible OK “Universal donor” – RBCs group O, plasma AB

Blood Component General Information Fresh Whole Blood Candidates Newborns needing exchange transfusion Patients requiring leukoreduced products

Blood Component General Information Summary of storage temperatures: Liquid RBCs 1-6C Platelets, Cryo (thawed) and granulocytes 20-24C (room temperature) ANY frozen plasma product ≤ -18C ANY liquid plasma product EXCEPT Cryo 1-6C

Blood Components Cellular Plasma Red blood cell products Platelets Granulocytes Plasma FFP Cryoprecipitate

Products With Red Cells

Changes in Stored Blood

Red Blood Cells, Packed (PRBC) Used to treat symptomatic anemia and routine blood loss during surgery Hematocrit is approximately 80% for non-additive (CPD), 60% for additive (ADSOL). Allow WB to sediment or centrifuge WB, remove supernatant plasma.

Leukocyte Reduced Red Cells (LR-RBC) Leukocytes can induce adverse affects during transfusion, primarily febrile, non-hemolytic reactions. Reactions to cytokines produced by leukocytes in transfused units. Other explanations to reactions include: immunization of recipient to transfused HLA or granulocyte antigens, micro aggregates and fragmentation of granulocytes. Historically, indicated only for patients who had 2 or more febrile transfusion reactions, now a commonly ordered, popular component. “CMV” safe blood, since CMV lives in WBCs. Most blood centers now leukoreduce blood immediately after collection. Bed side filters are available to leukoreduce products during transfusion.

Leukocyte Reduction

Washed Red Blood Cells (W-RBCs) Washing removes plasma proteins, platelets, WBCs and micro aggregates which may cause febrile or urticarial reactions. Patient requiring this product is the IgA deficient patient with anti-IgA antibodies. Prepared by using a machine which washes the cells 3 times with saline to remove and WBCs. Two types of labels: Washed RBCs - do not need to QC for WBCs. Leukocyte Poor WRBCs, QC must be done to guarantee removal of 85% of WBCs. No longer considered effective method for leukoreduction. e. Expires 24 hours after unit is entered.

Cell Washer to Prepare Washed Cells

Frozen Blood

Red Blood Cells Frozen; Red Blood Cells Deglycerolized (D-RBC) Blood is frozen to preserve: rare types, for autologous transfusion, stock piling blood for military mobilization and/or civilian natural disasters. Blood is drawn into an anticoagulant preservative. Plasma is removed and glycerol is added. After equilibration unit is centrifuged to remove excess glycerol and frozen. Expiration If frozen, 10 years. After deglycerolization, 24 hours. Storage temperature high glycerol -65 C. low glycerol -120 C, liquid nitrogen.

Rejuvenated Red Blood Cells A special solution is added to expired RBCs up to 3 days after expiration to restore 2,3-DPG and ATP levels to prestorage values. Rejuvenated RBCs regain normal characteristics of oxygen transport and delivery and improved post transfusion survival. Expiration is 24 hours or, if frozen, 10 years

Platelet Products

Platelets (PLTS), Platelet Concentrate (PC) or Random Donor Platelet Concentrate (RD-PC) Used to prevent spontaneous bleeding or stop established bleeding in thrombocytopenic patients. Prepared from a single unit of whole blood. Due to storage at RT it is the most likely component to be contaminated with bacteria. Therapeutic dose for adults is 6 to 10 units. Some patients become "refractory" to platelet therapy. Expiration is 5 days as a single unit,. Store at 20-24 C (RT) with constant agitation. D negative patients should be transfused with D negative platelets due to the presence of a small number of RBCs.

Preparation of platelet concentrate Plasma RBCs PRP Platelet concentrate

Platelets (PLTS), Platelet Concentrate (PC) or Random Donor Platelet Concentrate (RD-PC) One bag from ONE donor Need 6-10 for therapeutic dose

Pooling Platelets 6-10 units transferred into one bag Expiration = 4 hours

One pheresed unit is equivalent to 6-8 RD-PC. Platelets Pheresis, Apheresis Platelet Concentrate, Single Donor Platelet Concentrate (SD-PC) Used to decrease donor exposure, obtain HLA matched platelets for patients who are refractory to RD-PC or prevent platelet refractoriness from occurring. Prepared by hemapheresis, stored in two connected bags to maintain viability. One pheresed unit is equivalent to 6-8 RD-PC. Store at 20-24 C (RT) with agitation for 5 days, after combining, 24 hours D negative patients should be transfused with D negative platelets due to the presence of a small number of RBCs

Apheresis

Apheresis

Platelets Pheresis One bag (unit) from one donor One unit is a therapeutic dose Volume approximately 250 ccs

Granulocytes Lymphocyte Monocyte Neutrophils Eosinophils Basophils

Granulocytes Primary use is for patients with neutropenia who have gram negative infections documented by culture, but are unresponsive to antibiotics. Therapeutic efficacy and indications for granulocyte transfusions are not well defined. Better antimicrobial agents and use of granulocyte and macrophage colony stimulating factors best for adults, best success with this component has been with babies Daily transfusions are necessary. Prepared by hemapheresis. Expiration time is 24 hours but best to infuse ASAP. Store at 20-24 C.

Plasma Components

Fresh Frozen Plasma – Volume 200-250cc

Fresh Frozen Plasma (FFP) Used to replace labile and non-labile coagulation factors in massively bleeding patients OR treat bleeding associated with clotting factor deficiencies when factor concentrate is not available. Must be frozen within 8 hours of collection. Expiration frozen - 1 year stored at <-18 C. frozen - 7 years stored at <-65 C.thawed - 24 hours

Fresh Frozen Plasma (FFP) Storage temperature frozen -18 C, preferably -30 C or lower thawed - 1-6 C Thawed in 30-37C water bath or FDA approved microwave Must have mechanism to detect units which have thawed and refrozen due to improper storage. Must be ABO compatible

Pooled Plasma/Solvent Detergent Treated Most recently licensed product. Prepared from pools of no more than 2500 units of ABO specific plasma frozen to preserve labile coagulation factors. Treated with chemicals to inactivate lipid-enveloped viruses. Contains labile and non-labile coagulation factors but lacks largest Von Willebrand’s factor multimers. Used same as FFP.Safety concerns Decreases disease transmission for diseases tested for. Doesn’t inactivate viruses with non-lipid envelopes: parvo virus B19, hepatitis A, and unrecognized pathogens

Cryoprecipitate (CRYO), Factor VIII or Anti-Hemophilic Factor (AHF) Cold insoluble portion of plasma that precipitates when FFP is thawed at 1-6C. Cryoprecipitate contains high levels of Factor VIII and Fibrinogen, used for treatment of hemophiliacs and Von Willebrands when concentrates are not available. Used most commonly for patients with DIC or low fibrinogen levels. A therapeutic dose for an adult is 6 to 10 units. Can be prepared from WB which is then designated as "Whole Blood Cryoprecipitate Removed" or from FFP Plasma is frozen. Plasma is then thawed at 1-6 C, a precipitate forms. Plasma is centrifuged, cryoprecipitate will go to bottom. Remove plasma, freeze within 1 hour of preparation

Cryoprecipitate (VIII, vW) Thaw at 30-37°C Store at RT 4 hrs FFP Plasma cryoprecipitate, reduced (TTP, FII, V, Vii, IX, X, XI) Frozen within 8 hours Thawed FFP Refrozen with 24 hrs of separation Store at ≤18°C 1 yr 5 day expiration at 1-6°C

Cryoprecipitate (CRYO), Factor VIII or Anti-Hemophilic Factor (AHF) Storage Temperature Frozen -18 C or lower Thawed - room temperature Expiration: Frozen 1 year Thawed 6 hours Pooled 4 hours Best to be ABO compatible but not important due to small volume

Cryoprecipitate – volume 15ccs

Irradiation of Blood Components

Irradiation of Blood Components Cellular blood components are irradiated to destroy viable T- lymphocytes which may cause Graft Versus Host Disease (GVHD). GVHD is a disease that results when immunocompetent, viable lymphocytes in donor blood engraft in an immunocompromised host, recognize the patient tissues as foreign and produce antibodies against patient tissues, primarily skin, liver and GI tract. The resulting disease has serious consequences including death. GVHD may be chronic or acute

Irradiation of Blood Components Patients at greatest risk are: severely immunosuppressed, immunocompromised, receive blood donated by relatives, or fetuses receiving intrauterine transfusions Irradiation inactivates lymphocytes, leaving platelets, RBCs and granulocytes relatively undamaged. Must be labeled "irradiated". Expiration date of Red Blood Cell donor unit changes to 28 days. May be transfused to "normal" patients if not used by intended recipient.

Irradiation of Blood Components

Inspection of Donor Blood Segment closest to unit is hemolyzed. May indicate bacterial contamination

Safe-T-Vue Temperature Monitor

Transportation of Blood and Blood Components Frozen components Temperature must be maintained at or below required storage temperature. Use dry ice in well insulated container. Platelets and granulocytes Maintain at 20-24 C. Transport in well insulated containers without ice. Commercial coolers available to maintain at 20-24C.

Cont… Blood processing:- Donor selection Blood collection Component preparation Storage , issue and transportation Contract testing

Donor selection Principle of self-sufficiency from voluntary and non- remunerated donations have been recommended and promote by the Council of the Europe Recommendations No.R(95)14 Main purpose is to determine whether the person in good health, in order to protect the donor against damage to his/her own health, and to protect the recipient against transmission of disease or drugs which could be detrimental to the patient.

Blood collection & processing Aseptic technique Seal closed method Immediate storage at 1-6ºC Components preparation has to be done within 6 hours after collection Labels/records : ABO and Rh grouping Screening, expiratory date and volume of the blood

QC of blood component preparation Whole blood: Frequency of control: 1% of all units with minimum of 4 units per month Storage :- 2ºC to 6 ºC, for CPDA-1 the storage time is 35 days, CPD & CD2D – 22days. On expire date :- measure HCT, pH, total Hb , K+ and perform sterility assays

QA:- Volume : 450ml ± 10 % of body volume excluding anticoagulant HCT : 40±5% pH > 6.5 K < 27mmol/L Hb minimum 45g/unit Sterility : no growth

Red cell concentrates Perform the same assay as for Whole blood on the expiry date Storage : 2-6º C, for 35 days if prepared from WB collected in CPDA-1 QA: Volume : 280ml± 50ml, frequency of control 1% of all units HCT : 0.65-0.75 pH > 6.5 K < 78 mmol/L Hb : minimum 45g/unit Sterility : no growth

Platelet concentrates: Prepared within 6H of blood collection Must evaluate at least 4 platelet preparations monthly for platelet count,pH and plasma volume Platelets should be selected from each centrifuge in use The Tº at which pH is measured should be the same as stored Label the volume, Storage : 20-24ºC Tº should be recorded at least every 4H during storage.

QC Volume > 40ml pH : 6.8-7.4 Plt count : at least 5.5 x 1010 /bag in at least 75% of the units tested at the end of the storage.By apheresis : minimum 3 x 1011/bag platelets in at least 75% units tested WBC contamination: < 2 x 103/bag RBC contamination: < 2 x 109/bag Macroscopic appearance : no visible platelets aggregates Sterility : no growth

Fresh Frozen Plasma Every 10 unit/week estimate the volume Storage: 24 months at below –30ºC 12 months at –25 to –30ºC 3 months at –18 to –25ºC Thawed at Tº between 30-37ºC and transfused within 24H after thawing QC Volume: 220-250ml Factor VIIIc : > 0.7IU/ml- every 2 months No leakage after pressure in plasma extractor, before freezing and after thawing

QC Macroscopic : no abnormal color or visible clots Residual cell: Red cell: < 6.0 x 109/l Leukocyte: < 0.1 x 109/l Platelets : < 50 x 109/l

Cryoprecipitate Assayed on at least 4 bags/ month –for factor VIII Storage: 24 months at below –30ºC 12 months at –25 to –30ºC 3 months at –18 to –25ºC Must be thawed at 37ºC and used within 6H

QC Volume : 10-20 ml Factor VIII : > 70 IU/unit Fibrinogen : > 140 mg per unit Macroscopic : homogenous Sterility: no growth

Granulocytes Prepared by apheresis Storage: 20-24ºC for 24H QC Volume : < 500ml Granulocytes : > 10 x 109 per unit- present in at least 75% of all units tested

Transportation A system must be use to ensure that all blood and blood components shipped by or received into a blood bank or blood transfusion service have been maintained within T required. All liquid RBC components kept at T of 1-10ºC during transport All component routinely stored at 20-24ºC should be maintain T during shipment All frozen components should be transport in frozen state at –18ºC or colder Periodic T check and documented to ensure the transportation adequate to meet the criteria

QC FOR EQUIPMENT

Quality control for reagents Select the reagent with high specifications- reference preparation has been established for ABO, Rh and anti-human globulin (AHG) by FDA Color codes by the FDA: Blue for anti-A Yellow for anti-B Green for AHG Use according to manufacturer's instruction The new reagent has to be assessed & confirmed satisfactory The appearance each reagent has to be checked each day The reactivity and specificity has to checked each new lot