Introduction to Immunohematology

Encompasses activities, procedures, and testsdone to ensure that blood for transfusion is properly collected, preserved, stored, and dispensed for later use in blood transfusion

Blood Banking

is a multidisciplanary branch of medicine concerned with the transfusion of blood and blood components, including proper selection and utilization of blood components as well as removal of blood components in the treatment or prevention of disease

Transfusion Medicine

History of Transfusion Medicine

• 1492

• First recorded unsuccessful blood transfusion

• Donors were three young men in the hope of curing the stricken Pope Innocent VII

History of Transfusion Medicine

• 1628

• William Harvey > English physician, discovered the circulation of blood

History of Transfusion Medicine

• 1665

• The first recorded successful blood transfusion occurs in England

• Physician Richard Lower > kept dogs alive by transfusion of blood from other dogs

History of Transfusion Medicine

• 1818

• James Blundell > a british obstetrician performs the first successful transfusion of human blood to a patient for the treatment of postpartum hemorrhage

History of Transfusion Medicine

• 1840

• Samuel Armstrong Lane> At St. George's School in London, aided by consultant Dr. Blundelll performs the first successful whole blood transfusion to treat hemophilia

History of Transfusion Medicine

• 1867

• Joseph Lister > English surgeon, used antiseptics to control infection during transfusions

History of Transfusion Medicine

• 1869

• Braxton Hicks > recommended sodium phosphate in an attempt to find a non-toxic anticoagulant as clotting was the principal obstacle to overcame

History of Transfusion Medicine

• 1873 to 1880

• US physicians transfuse milk from cows, goats, and humans

History of Transfusion Medicine

• 1884

• Saline infusion replaces milk as a blood substitute due to the increased frequency of adverse reactions to milk

History of Transfusion Medicine

• 1901

• Karl Landsteiner > an Austrian physician, discovered the ABO blood groups

• He received the Nobel Prize for Medicine for this discovery in 1930

History of Transfusion Medicine

• 1902

• Alfred Decastello and Adriano Sturli, Landsteiner's colleagues, add AB blood group

History of Transfusion Medicine

• 1907 - Hektoen

• suggests that the safety of transfusion might be improved by crossmatching blood between donors and patients to exclude incompatible mixtures

History of Transfusion Medicine

• 1907 - Reuben Ottenberg

• performs the first blood transfusion with performed blood typing and crossmatching in Newyork

• also observed the mendelian inheritance of blood groups and recognized the universal utility of group ) donors

History of Transfusion Medicine

• 1908 - Alexis Carrel

• Alexid Carrel > French surgeon, devised a way to prevent clotting by sewing the vein of the recipient directly to the vein of the donot

• This vein-to-vein or direct method known as ANASTOMOSIS was practiced by a number of physicians

• Among them, J.B. Murphy in Chicago and George Crile in Clevelans

• The procedure was proved unfeasible for blood transfusion but paved the way for successful organ transplantation for which Carrel received the Nobel Prize in 1912

History of Transfusion Medicine

• 1908 - Moreschi

• Describes the antiglobulin reaction

• The antiglobulin is a direct way of visualizing an antigen-antibody reaction that has taken place but is not directly visible

• The antigen and antibody react with each other, them after washing to remove any unbound antibody, the antiglobulin reagent is added and binds between the antibody molecules that are stuck onto the antigen

• This makes the complex big enough to see

History of Transfusion Medicine

• 1912

• Roger Lee > a visiting physician at the Massachusetts General Hospital, along with Paul Dudley White develops the Lee-White clotting time

• Lee demonstrates that is is safe to give group O blood to patients of any blood group and that blood from all groups can be given to group AB patients

• The terms universal donor and universal recipient are coined

History of Transfusion Medicine

• 1913

• Edward Lindemann > Vein-to-vein blood transfusion was carried out using multiple syringes and a special cannula for puncturing the vein through the skin

History of Transfusion Medicine

• 1914

• Hustin > reported the use of sodium citrate as anticoagulant allowing longer preservation of blood

History of Transfusion Medicine

• 1915

• Richard Lewisohn > used sodium citrate as an anticoagulant to transform the transfusion procedure from direct to indirect

• Richard Weil > demonstrated the feasibility of refrigerated storage of such anticoagulated blood

History of Transfusion Medicine

• 1916

• Francis Rous and J.R. Turner > introduced a citrate-glucose solution that permits storage of blood for several days after collection

• Oswald Robertson > an American Army Officer, was credited with creating the blood depots, he received the AABB LAndsteiner Award in 1958 as developer of the first blood bank

History of Transfusion Medicine

• 1927 to 1947

• The MNSs and P systems are discovered

History of Transfusion Medicine

• 1939/1940

• The Rh blood group system was discovered by Karl Landsteiner, Alex Wiener, Philip Levine, and R.E. Stetson and was soon recognized as the cause of majority of transfusion reactions

History of Transfusion Medicine

• 1940

• The United States government established a nationwide program for the collection of blood

• Charles R. Drew > developed the “Plasma for Britain” Program which is a pilot project to collect blood for shipment to the British Isles

• The American Red Cross participated, collecting 13 million units of blood by the end of World War II

History of Transfusion Medicine

• 1941

• Dr. Drew > appointed director of the first american Red Cross Blood Bank at Presbyterian Hospital

History of Transfusion Medicine

• 1943

• J.F. Loutit and Patrick L. Mollison > introduce acid citrate dextrose (ACD) solution

• P. Beeson > published the classic description of transfusion-transmitted hepatitis

History of Transfusion Medicine

• 1945

Coombs, Mourant, and Race > described the use of antihuman globulin (later known as Coombs Test) to identify “incomplete” antibodies

History of Transfusion Medicine

• 1947

• The American Association of Blood Banks (AABB) was formed to promote common goals among blood banking practitioners and the blood donating public

History of Transfusion Medicine

• 1949 to 1950

• The US blood collection system include 1,500 hospital blood banks, 46 community blood centers, and 31 American Red Cross regional blood centers

History of Transfusion Medicine

• 1950

• Audrey Smith > reported the use of glycerol cryoprotectant for freezing red blood cells

• Carl Walter and W.P. Murphy Jr. > introduced the plastic bag for blood collection

History of Transfusion Medicine

• 1953

• Development of refrigerated centrifuge further expedited blood component therapy

History of Transfusion Medicine

• 1957

• The AABB formed its committee on Inspection and Accreditation to monitor the implementation of standards for blood banking

History of Transfusion Medicine

• 1958

• The AABB published its first edition of Standards for a Blood Transfusion Service (now titled as Standards for Blood Banks and Transfusion Services)

History of Transfusion Medicine

• 1960

• The AABB begins publication of TRANSFUSION, the first American journal wholly devoted to the science of blood banking and transfusion technology

History of Transfusion Medicine

• 1957

• Gibson > introduced an improved preservative solution called CPD (a less acidic anticoagulant)

History of Transfusion Medicine

• 1959

• Max Perutz > from Cambridge University deciphered the molecular structure of hemoglobin, the molecule that transports oxygen and gives red blood cells their color

History of Transfusion Medicine

• 1960

• A. Solomon and J.L. Fahey > reported the first therapeutic plasmapheresis procedure (a procedure that separates whole blood into plasma and red blood cells

History of Transfusion Medicine

• 1979

• A new anticoagulant preservative, CPDA-1, extends the shelf life of whole blood and red blood cellls to 35 days, increasing the blood supply and facilitating resource sharing among blood banks

History of Transfusion Medicine

• 1983

• Additive solutions extend the shelf life of red blood cells to 42 days

• Traditionally, the amount of whole blood in a unit has been _ of blood

• More recently, _ of blood are being collected

• 450 mL ± 10%

• 500 mL ± 10%

Units of whole blood collected can be separated into three components: (3)

packed RBCs, platelets, and plasma

• In recent years, less whole blood has been used to prepare platelets with the increased utilization of _

apheresis platelets

• Whole blood-prepared RBCs may be stored for _ to _ days

• 21 to 42 days

• Donated blood is free but a fee is still charged for each unit to cover the cost associated with (4)

collection, storage, testing, and transfusion

The Donation Process

Current Donor Screening Test for Infectious Diseases

Three areas of RBC biology are crucial for normal erythrocyte survival and function:

• Normal chemical composition and structure of the RBC membrane

• Hemoglobin structure and function

• RBC metabolism

• RBC membrane is a semi-permeable lipid bilayer supported by a mesh-like protein cytoskeleton structure

• Biochemical composition:

• 52% protein

• 40% lipid

• 8% carbohydrates

RBC membrane

• _ extend from the outer surface and span the entire membrane of the inner cytoplasmic side of RBC

Integral Membrane Proteins

RBC membrane

• _ > located and limited to the cytoplasmic surface of the membrane; form the RBC cytoskeleton

Peripheral Proteins

Integral and Peripheral Proteins

• The normal chemical composition, structural arrangement and molecular interactions of the erythrocyte membrane are crucial to the normal length of RBC survival in the circulation and in RBC's two important characteristics:

• Deformability

• Permeability

DEFORMABILITY

• To remain viable, nrmal RBCs must also remain flexible, deformable, and permeable

• The loss of ATP levels leas to decrease in _ and in turn a loss of membrane deformability

• An accumulation or increase in deposition of membrane _ also results, causing an increase in membrane rigidity and loss of pliability

• phosporylation of spectrin

• calcium

PERMEABILITY

• The permeability properties of the RBC membrane and the active _ prevent colloid hemolysis and control the volume of RBC

• RBC IC to EC ratios of Na+ and K+: _

• When RBCs are ATP-depleted, Ca2+ and Na+ accumulate intracellularly and and are lost > dehydrated and rigid cell subsequently sequested by the spleen

• RBC cation transport

• 1:12 (Na+) and 25:1 (K+)

• K+ and H2O

• Primary function is as transport: O2 delivery to tissues and CO2 excretion

Hemoglobin

Hemoglobin molecule is composed of four subunits, each containing heme and globin:

1 heme = 1 mole of O2

1 Hb molecule = 4 moles of O2

One of the most important controls of hemoglobin's affinity for oxygen is _

2,3-DPG (diphosphoglycerate or biphosphoglycerate)

• Graphically describes the relationship between oxygen content of hemoglobin (% of saturation) and the partial pressure of O2 (PO2)

Oxygen Dissociation Curve - As 𝑃𝑂2 increases, hemoglobin's affinity for oxygen increases, causing a rapid rise in saturation, which then plateaus at higher pressures. 

Normal shape of the curve is _

sigmoid

Oxygen Dissociation Curve

• Shift to the LEFT

• Increased Hb affinity for O2, decreased delivery of O2 to tissues

• Increased pH

• Decreased 2,3-DPG, CO2, and temperature

Oxygen Dissociation Curve

• Shift to the RIGHT

• Decreased Hb affinity for O2, increased delivery of O2 to tissues

• Decreased pH

• Increased 2,3-DPG, CO2, and temperature

RBC Metabolic Pathways

• Anaerobic Glycolytic Pathway

→Embden-Meyerhof Pathway

• Ancillary Pathways

→Pentose Phosphate Pathway / HMP - prevents denaturation of hemoglobin by oxidation

→Methemoglobin Reductase Pathway - Heme iron is constantly exposed to oxygen and peroxides. Peroxide oxidizes heme iron from the ferrous (2+) to the ferric (3+) state (methemoglobin); maintains iron in heme in its reduced form (Fe+2). This pathway Converts methemoglobin back to normal hemoglobin using the methemoglobin reductase enzyme which makes it a corrective mechanism

→Leubering-Rapoport Shunt - generates 2,3 - DPG

• Blood collection is a _ system consisting of main bag with needle, tubing, and up to four satellite bags attached

• The entire system is sterile

• closed system

How many mL of blood is taken in blood collection

• 450 mL ± 45 mL (with 63 mL anticoagulant) or 500 mL ± 50 mL (with 70 mL anticoagulant)

Blood Preservation

• Component of anticoagulant used in blood donation (4)

• adenine - used in ATP synthesis

• citrate - binds calcium to prevent coagulation

• dextrose - food for cells

• phosphate - source of 2,3-DPG which promotes oxygen release to tissues

4 anticoagulant sued in blood banking

• ACD: Acid-Citrate-Dextrose (21 days)

• CPD: Citrate-Phosphate-Dextrose (21 days)

• CP2D: Citrate-Phosphate-Double Dextrose (21 days)

• CPDA-1: Citrate-Phosphate-Dextrose-Adenine (35 days)

Biochemical changes in stored blood that can lead to decreased RBC viability

lesions of storage

Lesions of Storage

• (5) decrease as RBCs are stored

• After cells are transfused, (2) are restored in about 24 hours

• Glucose, ATP, 2,3-DPG, pH, plasma sodium

• ATP and 2,3-DPG

Lesions of Storage

• Substances that increase during storage include (4)

• plasma hemoglobin

• plasma potassium

• ammonium

• lactic acid

These are added to RBCs after removal of plasma with or without platelets

Additive Solutions

Additive solutions contain _

MAGS

• Mannitol

• Adenine

• Glucose

• Saline

Additive solutions must be added within _

72 hours

Additives extend their shelf life to _ and reduce RBC viscosity during transfusion

42 days

Freezing of RBCs is primarily used for (2)

• autologous units

• storage of rare blood types

Freezing of RBCs

• Cryoprotective agent is added to RBCs that are less than _ old

6 days

• allows individual to donate blood for their own use in meeting their needs for blood transfusion

Autologous transfusion

• auto meaning self

Advantages of Freezing RBCs (4)

• Long term storage (10 years)

• Maintenace of RBC viability and function

• Low residual leukocytes and platelets

• Removal of significant amounts of plasma proteins

Disadvantages of Freeezing RBCs

• Time-consuming process

• Higher cost of equipment and materials

• Storage requirements (-65 °C)

• Higher cost of product

Advantages of HIgh-Concentration Glycerol Technique Used by Most Blood Banks Over Low-Concentration Glycerol Techniques

• These are used to restore ATP and 2,3-DPG levels before freezing or transfusing a unit, and may be necessary for autologous or rare units

Rejuvenation Solutions

Rejuvenation Solutions contain _

PIGPA

• Phosphate

• Inosine

• Glucose

• Pyruvate

• Adenine

Current Trends in RBC Preservation Research

• Development of improved additive solutions

• Development of procedures to reduce and inactivate the
  level of pathogens that may be in RBC units

• Development of procedures to convert A-, B-, and ABtype RBCs to O-type RBCs

• Development of methods to produce RBCs through
  bioengineering (blood pharming)

• Development of RBC substitutes

Current Trends in Platelet Preservation Research

• Development of methods that would allow platelets to
  be stored for 7 days

• Development of additive solutions, also termed synthetic
  media

• Development of procedures to reduce and inactivate the
  level of pathogens that may be in platelet units

• Development of platelet substitutes

• New approaches for storage of platelets at 1°C to 6°C

• The development of processes to cryopreserve platelets