Blood Transfusion: ABO and RhD Systems Explained by Dr. Ahmad Hassaneen

1. Blood transfusion Dr. Ahmad Hassaneen

2. Important Notes

3. Comparison of ABO and RhD systems

4. In ABO system, there are 4main types of blood groups- A, B, AB, and O. Identification of these groups is based on presence or absence of A and/or B antigens on red cells. According to Landsteiner’s law, anti-A and/or anti-B antibodies are always present in plasma of individuals who lack corresponding antigen(s) on their red cells. There are two major subgroups of A: A1 (80%) and A2 (20%). Thus ABO system comprises of six groups. A1: Having anti-B A2: Having anti-B , and in 1-8% cases anti-A1 B: Having anti-A A1B: nil ( No antibodies) A2B: Having anti-A1in 22-35% cases O: Having anti-A and anti-B.

5. Usually, anti-A1 antibodies are weak and are of little clinical significance in routine practice. • N.B. : A2B can be wrongly grouped as B

6. Secretors and non-secretors Secretors are persons who secrete A, B, and H antigens into body fluids (such as plasma, saliva, sweat, tears, semen, milk, etc.). This ability is dependent on presence of a dominant secretor gene (Se). About 80% of individuals are secretors (genotype Seseor SeSe) and the remaining are non-secretors (genotype sese). Both secretors and non-secretors express ABO antigens on red cells. • Antigens secreted by different ABO blood groups are: • Group A: A, H • Group B: B, H • Group AB: A, B, H • Group O: H

7. RH SYSTEM ●According to Fisher and Race, three closely linked genes are inherited together on one chromosome (haplotype) from each parent. Allelic forms of these genes are C and c, D and d, and E and e with eight possible haplotypes- CDe, cde, cDE, cDe, cdE, Cde, CDE, and CdE. ● It has been found that the RH locus is located on chromosome 1 and consists of two closely linked genes- RHD and RHCE. The allelesof RHCE are CE, Ce, ce, and cE. ●The presence of D in either homozygous (D/D) or heterozygous (D/d) state makes that individual Rh positive, while Rh negative persons are homozygous for d (d/d). ● d gene is an amorph→ No d Ag or Anti-d Ab

8. Blood Components

9. ●In the past , whole blood was the only preparation that could be given to replace red cells , platelets , coagulation factors , …. ● Now , whole blood can be separated into different components by centrifugation due to differences in specific gravities of these components. ● This is facilitated by introduction of double and triple bags having closed integral tubing. ● After their separation, various components can be transferred from one bag to another in a closed circuit thus maintaining the sterility. ● Blood should be processed for component separation within 6 hours of collection.

10. Whole blood: • It remains a commonly employed blood product at some places because of lack of facility for component separation. • Total volume of one unit is about 400 ml (350 ml blood+50 ml of anticoagulant-preservative solution) • Commonly used anticoagulant is citrate phosphate dextrose adenine (CPDA) • It consists of the cellular elements and plasma. • Stored in blood bank refrigerator at 2-6 °C • Shelf life is 35 days (for CPDA ). • Hematocriteof whole blood is about 40 %

11. ●Before ordering whole blood transfusion , the following should be noted: ▪ Transfusion of one unit raises the hemoglobin by 1 g/dl or the hematocrite by 3 % (detectable 24 hours after transfusion) ▪ If stored for 48 hours does not contain functionally effective plateletsand labile coagulation factors (V and VIII). ▪ There is risk of volume overload in patients with impaired cardiovascular function. ● Currently , the only indications are : 1-Acute massive blood loss 2-Exchange transfusion of newborn 3-Non-availability of packed red cells.

12. Blood components: ●Red cells: Packed red cells Leucocyte-poor red cells Frozen red cells Irradiated red cells. ●Platelets: Platelets concentrate ●Granulocytes Granulocytes concentrate ●Plasma Fresh frozen plasma Cryoprecipitate

13. ●Red cells components: Packed red cells Prepared by removal of most of the plasma from one unit of whole blood. Method: Whole blood is either allowed to sediment overnight in a refrigerator at 2 °C to 6 °C OR is spun in a refrigerated centrifuge → supernatant fluid (PLASMA)is separated from red cells in a closed system by transferring it to the attached empty satellite bag. Red cells and a small amount of plasma are left behind in the primary blood bag. The main indication: Replacement of red cells in case of anemia.

14. • Volume of 1 unit of packed red cells is about 300 ml •Hematocriteof packed red cells is 55-75 % • Transfusion of one unit raises the hemoglobin by 1 g/dl or the hematocrite by 3 % (detectable 24 hours after transfusion) • Stored in blood bank refrigerator at 2-6 °C • Shelf life is 35 days (for CPDA )

16. Leucocyte-poor red cells • Most of white cells are removed. • Prepared by passing blood through special leucocyte-depletion filter at the time of transfusion. • Also can be prepared in the blood bank and then stored. • Indications: 1-To avoid febrile reactions in persons who need repeated transfusions or who have been sensitized to white cells antigens before. 2-To reduce the risk of transfusion of cytomegalovirus (CMV) if CMV-seronegative blood is not available.

17. Frozen red cells • Cells can be stored frozen for up to 10 years. Indications: • Donor red cells with rare blood groups can be stored frozen for transfusion to other recipients ORfor autologous transfusion.

18. Irradiated red cells • Red cells are irradiated by gamma rays to prevent graft-versus-host disease in susceptible individuals for example: 1-Immunodeficient patients 2-Patients receiving blood from first-degree relatives. (In this case, the donor's T- lymphocytes may not be rejected as the Human Leucocyte Antigen (HLA) type of the recipient has similarity with that of the donor) ▪ Gamma irradiation inhibits the replication of donor lymphocytes → they will not react against tissues of recipient.

19. ● Granulocyte concentrates • These are prepared either from a single donor unit by differential centrifugation or by leucapheresis. • Leucapheresis is preferred because it gives better granulocyte yield . • They are used in patients with severe neutropenia (< 500 neutrophils /ul) with bacterial or fungal infections who are not responding to antibiotic therapy. • Complications: 1-Transmission of cell-associated viruses like CMV . 2-Risk of causing GVHD (Graft versus host disease) , so it must be irradiated (Gamma rays) first.

20. Platelets There are 2 methods for obtaining platelets: 1-Differential centrifugation of a unit of whole blood (platelets concentrate) 2-Plateletpheresis

21. Platelet concentrate (Random donor platelets) - Prepared by differential centrifugation of one unit of whole blood within 6 hours of donation. • one unit of whole blood is centrifuged at low speed to obtain platelet-rich plasma (PRP). • PRP is then transferred to the attached satellite bag and spun at high speed to get platelet aggregates (at the bottom) + platelet-poor plasma (PPP) (at the top). • Most of PPP is returned back to the primary collection bag leaving the platelets with only 50-60 ml of PPP.

23. Platelets are stored at 20°C to 24 °C with continuous shaking (agitated) in a storage device called platelet agitator. • Maximum storage period is 3-5 days. - Transfusion of one unit will raise the platelets count by about 5000/ul - The usual adult dose is 4 – 6 units of platelet concentrate (from different donors) which are pooled into one bag before transfusion.

24. Plateletpheresis (single donor platelets) • A donor is connected to a blood cell separator machine in which whole blood is collected. • Platelets are separated and retained. • The remaining components are returned back to the donor. • By this method , a large number of platelets can be obtained from a single donor (equivalent to 6 units) Notes about platelets transfusion: • Most of adverse reactions associated with platelets are due to presence of contaminating leucocytes or plasma (e.g. febrile reactions , allergic reactions , …)

25. Transfusion of multiple platelet concentrates from random donors can induce alloimmunization to HLA antigens → causing resistance to further platelets transfusions. Those patients should receive HLA-compatible platelets from single donor by plateletpheresis. • Bacterial proliferation can occur in platelet concentrates as they are stored at room temperature , may cause septicemia in the recipient. • As platelet concentrates also contain a small amount of red cells it is better to be from the same or compatible ABO group and same Rh group.

26. Plateletpheresis

27. Preparations from human plasma: • Fresh frozen plasma (FFP) ▪Rapidly frozen plasma separated from fresh blood is stored at less than − 25° C. ▪ Volume of FFP is 200 to 250 ml ▪It contains all the coagulation factors. ▪Can be stored for 1 year if temperature maintained below -25 °C ▪When needed for transfusion , it is thawed between 30 and 37 °C and then stored at 2 to 6 °C and should be given within 2 hours of thawing because labile factors (V & VIII) are lost rapidly. ▪Used for: 1- Deficiency of multiple coagulation factors e.g. liver disease , DIC. 2- Inherited deficiency of a coagulation factor for which no specific concentrate is available.

28. Fresh frozen plasma (FFP)

29. Cryoprecipitate -Prepared by slowly thawing 1 unit of FFP at 4to 6 °C → formation of plasma + white precipitate. -After centrifugation , most of the supernatant plasma is removed leaving sediment of Cryoprecipitate suspended in 10-20 ml of plasma → frozen (− 25° C) for storage , can be kept for 1 year. - When required for transfusion , it is thawed between 30 °C and 37 °C and then stored in refrigerator at 2 to 6 °C and should be given within 6 hours of thawing. -It contains : F VIII , von Willebrand factor , fibrinogen , F XIII and fibronectin.

30. Factor VIII concentrate • Prepared by fractionation from large pools of donated plasma. • Supplied as freeze-dried powder in vials. • Stored in refrigerator at 2 to 6 °C. • Before use , it is reconstituted according to manufacturer’s directions and given I.V. • Now , it can be prepared by recombinant DNA technology without use of plasma and it is safer ; no risk of infection. • Uses: 1-Hemophilia A. 2-Severe von Willebrand disease.

35. Blood transfusion device

37. Important Notes

38. ►To reduce the risk of haemolysis in a case of non-identical but compatible ABO transfusion, packed red cells instead of whole blood should be transfused (i.e. most of the plasma which contains anti-A and/or anti-B should be removed). • ► This is especially important with group O donor blood, which can contain immune anti-A and anti-B antibodies that will cause serious haemolysis in a non-group O recipient. • ► For AB group recipients, if red cells of group AB are not available, group A donor blood is preferred over other alternatives since anti-B in group A is weaker than anti-A in group B.

41. Criteria for selection of blood donors

42. Criteria for selection of blood donors: ●Donor selection process consists of 4 parts: I- Predonationcounselling. II-Medical history III-Physical examination IV-Hemoglobin estimation

43. I-Predonationcounselling: This is an education programme including: 1-Giving information about health condition , self exclusion/self deferral , procedures involved in blood donation. 2-Donor’s questions are answered. 3-Reassurance is given in case of anxiety.

44. II-Medical history: Age: Lower and higher age limit for donation are 18 and 60 years respectively. This is because of : -Age of increased iron needs of adolescents (under 18) -Risk of cardiovascular and cerebrovascular disease in old age following removal of large quantity of blood. Donation interval: -Interval between 2 consecutive blood donations should be at least 3 months to avoid iron depletion. Pregnancy and lactation: Pregnant and lactating women (up to 1 year after labor) should not donate blood.

45. Infectious diseases: -HIV : - Blood should not be collected from donors who give history suggestive of HIV infection (unexplained fever , weight loss , enlarged lymph nodes , uncontrolled diarrhea) - Elimination of the “window period” which is early antibody-negative period in persons infected with HIV : Persons who have been exposed to the risk of HIV infection are excluded (e.g. homosexuals , intravenous drug abusers , or having contact with AIDS persons)

46. Hepatitis An individual with history of jaundice within 1 year should not be accepted. Malaria: -In endemic areas , a donor may be accepted after 3 months of asymptomatic period following malarial attack and after full treatment. -In non-endemic areas , a donor with any history of malaria is excluded.

47. Various illnesses: Exclusion of donors with : Diabetes mellitus , hypertension , heart disease , renal disease , liver disease , lung disease , cancer , epilepsy , bleeding disorders . Dentistry: 72 hours deferral period following tooth extraction or filling due to the possibility of bacteremia. Skin piercing: Exclusion of donors with history of : Tattooing or accidental needle stick during the last 12 months

48. Blood transfusion: A person should not be accepted as blood donor for 6 months after receiving blood transfusion. Vaccination: ♦ Donors who have received killed viral vaccines at any time are acceptable. ♦ Other types of immunization and respective deferral period are as follow: -Attenuated live virus vaccine for measles , mumps , yellow fever , poliomyelitis : 2 weeks -German measles(Rubella) : 4 weeks • Rabies : 1 year

49. ●III-Physical examination: • Weight: should be at least 45 kg • Blood pressure: Systolic: from 100 – 180 mmHg Diastolic: from 50 – 100 mmHg • Pulse: 50-100 /min , and should be regular • Body temperature: normal ● IV-Hemoglobin estimation Hemoglobin should be at least 12.5 g /dl

50. Complications (Adverse effects) of Blood transfusion