Blood component therapy and blood substitutes

1. Blood component
therapy and blood
substitutes
DR. HIMANSHU JANGID

2. INTRODUCTION
Blood products are any therapeutic substances that are prepared from human blood.
These can be classified into
1.Whole blood
2.blood components :red cell concentrates, platelet concentrates, fresh plasma and
cryoprecipitate
3.plasma derivatives :albumin, coagulation factors and immunoglobulins.

3. Whole blood
Although in many countries it still accounts for most transfusions, whole blood is rarely used
in many developed countries.
Whole blood has a shelf life of 35 days.
Citrate phosphate dextrose adenine (CPDA-1) is an anticoagulant preservative in which
blood is stored at 1°C to 6°C.
The storage at 1°C to 6°C assists preservation by slowing the rate of glycolysis approximately
40 times the rate at body temperature.

4. Storage of Blood
1. Citrate is an anticoagulant (prevents clotting by binding calcium).
2. Phosphate serves as a buffer.
3. Dextrose is a red cell energy source (allows the RBCs to continue
glycolysis).
4. adenine allows RBCs to resynthesize adenosine triphosphate
(ATP).
which extends the storage time from 21 to 35 days.

5. Storage of Blood
1. The shelf life is extended to 42 days with AS-1 (Adsol), AS-3 (Nutricel), or AS-5 (Optisol).
2. Adsol contains adenine, glucose, mannitol, and sodium chloride.
3. Nutricel contains glucose, adenine, citrate, phosphate, and sodium chloride.
4. Optisol contains only dextrose, adenine, sodium chloride, and mannitol.

6. COMPONENT PRODUCTION
Blood components may be prepared from
whole blood donations or
by apheresis.
Apheresis
is the process by which only the component that is to be used is separated out and collected and
the rest is returned to the donor.

7. Key Elements
Donors are chosen to exclude anyone whose blood may harm the recipient.
Whole blood is centrifuged to separate plasma from red cells.
After collection, red blood cells are re-suspended and stored in a preservative solution to
keep them in good condition.
Each donation is tested to establish the ABO and RhD group of the donor’s red cells.

8. Blood screening
Blood screening varies across the globe.
hepatitis B (surface antigen),
human immunodeficiency virus (HIV) (antibody),
human T-lymphotrophic virus (HTLV)(antibody),
hepatitis C (antibody and RNA),
and syphilis (antibody).
Tests for malaria antibodies,
Trypanosoma cruzi (Chaga’s disease) antibodies or West Nile virus RNA

9. Key Elements
No test yet exists for the reliable detection of variant Creutzfeldt-Jakob disease (vCJD).
Leucodepleted (removal of white blood cells by filtration) in an attempt to reduce the
potential risk from transfusion-transmitted vCJD.

10. Key Elements
Plasma derivatives are partially purified preparations of human plasma proteins
Thawing, the addition of ethanol and exposure to varying temperatures, pH and ionic
strengths are combined with filtration, chromatography and centrifugation to separate the
different groups of proteins.
Further purification and virus inactivation steps are also performed.
This pooling of huge numbers of donations potentially increases the risk of infective disease
transmission.

11. Key Elements
The USA supplier only provides plasma from male donors to reduce the risk of Transfusion-
Related Acute Lung Injury (TRALI).
Women are excluded, as they may be a source of antibodies that increase the risk of TRALI.
These antibodies are thought to occur during pregnancy.

12. Blood components

13. Centrifugation

15. Packed RBC
Packed red cells are produced by
removing between 150-200ml of citrated plasma from a unit of whole blood.
 Haematocrit = 60-70%.
Storing red cells just above freezing allows survival for up to 42 days, however during this
time, the cells slowly deteriorate.

16. Comparison of Whole Blood and
Packed Red Blood Cells
Value Whole Blood Packed Red Blood Cells
Volume (mL) 517 300
Erythrocyte mass (mL) 200 200
Hematocrit (%) 40 70
Albumin (g) 12.5 4
Globulin (g) 6.25 2
Total protein (g) 48.8 36
Plasma sodium (mEq) 45 15
Plasma potassium (mEq) 15 4
Plasma acid (citric-lactic) 80 25 (mEq)
Donor-to-recipient ratio 1 unit per patient 1 unit per 4-6 patients

17. Packed RBC
Gradual loss of 2,3-diphosphoglycerate (2,3-DPG)
leading to increased haemoglobin affinity for oxygen
reduced oxygen delivery to the tissues.
 Gradual loss of membrane integrity .
Potassium leaks out of the cells into the suspension fluid and there is a gradual build up of
lactate.
Some cells may eventually die and become haemolysed.

18. Packed RBC
Despite these storage problems
 It is the fastest way to increase the oxygen delivering capacity of the blood.
1 unit of packed red cells = haematocrit by 3% and haemoglobin by 1 g/dl.

19. Packed RBC
The administration of packed RBCs is facilitated by reconstituting them with a crystalloid or
colloid; however, not all crystalloids are suitable.
If the solution contains calcium, clotting occurs.
Lactated Ringer's solution is not recommended.
Solutions recommended for reconstituted packed erythrocytes are 5% dextrose in 0.4%
saline, 5% dextrose in 0.9% saline, 0.9% saline, and Normosol-R with a pH of 7.4.

20. Microaggregate-free blood
Microaggregate-free blood is used to prevent reactions to leucocyte and platelet antigens.
Specially designed machines are used to wash the red blood cells (RBCs), which are then
suspended in sterile saline.
Washed red cells usually have haematocrits of 70-80% and a volume of about 180ml.

21. Irradiated red cells
Gamma radiation is used to destroy the lymphocytes in a unit of packed red cells that are
responsible for transfusion related graft versus host disease.
This is used for
Severely immuno compromised patients
Lymphoma patients,
Stem cell and marrow transplants
Unborn children undergoing intrauterine transfusion.

22. Leucocyte depleted red cells
Leucocyte depleted red cells have had 99.9% of the white cells removed by freezing or
microfiltration.
This reduces, but does not eliminate the risk of cytomegalovirus (CMV), Epstein-Barr, HTLV
infections and febrile reactions.
Since October 1999, all blood components in the UK and Ireland have been leucodepleted in
an attempt to decrease the potential risk from transfusion-transmitted C-J Disease.

23. Granulocytes
Granulocytes are indicated for
Life-threatening infections in neutropenic cancer patients who are unresponsive to
antibiotics.
They are prepared by
Separating white blood cells from blood donated by volunteers whose leucocyte count has
been increased by pre-treatment with corticosteroids,
Or those with chronic granulocytic leukaemia.

24. Granulocytes
The donations must be cross-matched because they contain large numbers of red blood
cells.
 They should also be irradiated to remove the lymphocytes.
They are collected by
Apheresis
Derived from whole blood
Can only be stored for 24 hours at 20-24 °C.

25. Platelet Concentrates
Platelets for transfusion are collected in two ways:
Pooled platelets:
This is a two-step procedure.
Firstly, one unit of platelets is produced from a unit of whole blood.
Then, 4-6 of these units (from different donors) are ‘pooled’ together in a single pack.

26. Platelet Concentrates
Apheresis platelets:
Being collected from a single donor (to reduce the risk of disease transmission).
As blood cycles through the apheresis machine, platelets are removed and all other blood
constituents are returned to the donor.
The amount of platelets collected with this procedure represents the equivalent of 4-6 units
of random donor platelets.

27. Platelet Concentrates
Platelets last for 3-5 days if stored on an agitator at 22°C and at a pH of between 6.2 and 7.8.
Each bag has a volume of 250-350ml.
Platelets should be inspected prior to infusion and packs should be rejected, or referred for
further opinion, if there is any unexpected appearance such as discolouration.
Platelets are not usually cross-matched with the recipient, but where possible ABO specific
platelets should be used.

28. Platelet Concentrates
The risk of transmission of bacterial infection is higher with platelet transfusions.
Bacterial contamination at the time of collection
The storage bags are made of special plastic, which allows gas exchange (oxygen and carbon
dioxide) to occur across its walls at 22°C.
This helps preserve platelet function but promotes bacterial growth.
The longer the platelets are kept prior to transfusion, the higher the risk of bacteraemia.

29. History of Platelet Concentrates
Shelf-Life
Year Shelf-Life Practical Shelf-Life *
1984-1986 7 days 6-7 days †
1986-1999 5 days 3 days ‡
1999-2004 5 days 3 days §
2004-present 5 days 2.5 to 3 days
*
Days that platelet concentrates are actually available to clinicians.
† Reports of bacterial contamination.
‡ Nucleic acid technology testing, centralized blood donor testing.
§ Bacterial detection implemented.

30. ASA Recommendations for platelet
transfusion
Prophylactic platelet transfusion is ineffective and rarely indicated when thrombocytopenia
is due to increased platelet destruction (e.g., idiopathic thrombocytic purpura).
Prophylactic platelet transfusion is rarely indicated
when the platelet count is greater than 100000/ml and
usually indicated when the platelet count is less than 50000/ml.

31. ASA Recommendations
With intermediate platelet counts (50 to 100 × 109/L), the determination should be based
on the patient's risk for more significant bleeding.
Vaginal deliveries or operative procedures ordinarily associated with insignificant blood loss
may be undertaken in patients with platelet counts less than 50 × 109/L.
Platelet transfusion may be indicated despite an apparently adequate platelet count if there
is known platelet dysfunction and microvascular bleeding.

32. ASA Recommendations
Patients with severe thrombocytopenia (<20,000 cells/mm3
) and clinical signs of bleeding
usually require platelet transfusion.
However, patients may have very low platelet counts (much less than 20,000 cells/mm3
) and
not have any clinical bleeding.
Patients such as these probably do not need platelet transfusions.
Individuals who have undergone trauma or require surgery need higher platelet counts,
probably 100,000 cells/mm3
, to maintain adequate hemostasis.
Laboratory determinations and clinical evaluations must be taken into account before a
decision to transfuse platelets is made.

33. Response to platelets
Under ideal circumstances,
one platelet concentrate = 7000 to 10,000 platelets/mm3
at 1 hour after transfusion to the
70-kg adult.
Ten units of platelet concentrates = 100,000 cells/mm3
.
Splenomegaly.
Previous sensitization. decreased survivals of platelets.
Fever, sepsis.
active bleeding.

34. Fresh Frozen Plasma
FFP is collected as the supernatant after centrifuging a donation of whole blood.
It contains all the plasma proteins, Particularly factors V and VIII,
which gradually decline during the storage of blood.
The use of FFP carries with it certain inherent risks
that are observed with the use of essentially any blood product.

35. Collection and storage of FFP
It is frozen within 8 hours and may be stored for up to 1 year at -30 °C.
Under these conditions, the loss of Factors V and VIII is kept to a minimum.
Frozen packs are brittle and should be handled with care.
The frozen plasma can be thawed using a dry oven (10 minutes), microwave (2-3 minutes) or
a water bath (20 minutes).
Thawed FFP is best used immediately but may be stored at 4°C and infused within 24 hours.

36. ASA Guidelines
 For urgent reversal of warfarin therapy
 For correction of known coagulation factor deficiencies for which specific correlates are
unavailable
 For correction of microvascular bleeding in the presence of increased (>1.5 times normal)
prothrombin time or partial thromboplastin time
 For correction of microvascular bleeding secondary to coagulation factor deficiency in
patients transfused with more than 1 blood volume and when prothrombin time and partial
thromboplastin time cannot be obtained in a timely fashion.

37.  FFP should be given in doses calculated to achieve a minimum of 30% of plasma factor
concentration (usually achieved with administration of 10 to 15 mL/kg of FFP), except for
urgent reversal of warfarin anticoagulation, for which 5 to 8 mL/kg of FFP usually suffice.
 Four to five platelet concentrates,
1 unit of single-donor apheresis platelets, 1 unit of FFP (Coag. factors)
 or 1 unit of whole blood provides
 FFP is contraindicated for augmentation of plasma volume or albumin concentration.

38. In the UK, all children born after 1st January 1996 (following which the risk from
contaminated foods was vastly reduced) who require plasma, receive FFP sourced from the
USA to minimise vCJD transmission.
This FFP is methylene blue treated to minimise the risk of viral infection.

39. Cryoprecipitate
If FFP is
Thawed to refrigerator temperature (4°C)
a precipitate forms that is very rich in high molecular weight proteins,
including factor VIII, von Willebrand factor and fibrinogen
Known as cryoprecipitate.
Viral infection risk is as for whole blood, and it is administered through a standard blood
filter.

40. Collection and storage
1 unit of whole blood provides about 15ml of cryoprecipitate.
One pack contains
150-300mg of fibrinogen
70iu of factor VIII.
von Willebrand factor
fibronectin.
trace amounts of other plasma proteins

41. Cryoprecipitate
The use of cryoprecipitate
Factor VIII deficiency or hemophilia A
Treatment of fibrinogen deficiencies
Is preferable to commercially prepared fibrinogen preparations, which have a very high
incidence of hepatitis.

42. Recommendations and
indications
Rarely indicated if fibrinogen concentration is greater than 150mg/dl.
The indications :
1. When the fibrinogen concentration is less than 100mg/dl in the presence of excessive
microvascular bleeding.
2. To correct excessive microvascular bleeding in massively transfused patients when
fibrinogen concentrations cannot be measured in a timely fashion.
3. For patients with von Willebrand’s disease and congenital fibrinogen deficiencies.

43. Cryoprecipitate should be administered through a filter and as rapidly as possible.
The rate of administration should be at least 200 mL/hr, and infusion should be completed
within 6 hours of thawing.
Fibrin glue:
 Is used occasionally by surgeons to create local hemostasis.
It is prepared in a manner similar to that of cryoprecipitate.
When FFP is thawed, the precipitate contains large amounts of fibrinogen.
When centrifuged, about 4 mL of concentrated precipitate results.
With added thrombin, it is applied locally, the efficacy of which is difficult to determine.

44. Prothrombin Complex
Factor IX can be recovered from plasma or plasma fractions by absorption with ion
exchanges or inorganic chemicals.
These products are all complexes of factors II, VII, IX, and X.
The main indication for these products is treatment of factor IX deficiency, or hemophilia B
(i.e., Christmas disease).

45. Single-Donor Plasma
Single-donor plasma is plasma that has been removed from stored blood without any effort
being made to preserve coagulation factors.
Single-donor plasma is very effective as a volume expander.
All the precautions outlined for the administration of FFP should be followed when single-
donor plasma is administered.
It obviously cannot be used to correct deficiencies in coagulation factors.

46. Albumin
Albumin is available as a 5% or a 25% solution in isotonic saline.
Plasma protein fractions containing albumin and α and β globulins are available.
These solutions are prepared commercially from albumin fractions from large pools of
plasma reconstituted in isotonic electrolyte solutions.
Albumin is heat-treated to kill viruses.

47. Albumin
Shelf life of 2 years and should be stored at room temperature.
Such solutions can be given without regard to ABO blood type and without crossmatch and
should be used primarily as volume expanders.
Bacterial sepsis has been associated with albumin administration.
Should be administered within 4 hours of initiation of the infusion because of the potential
for contamination after opening the bottle.

48. Use of Albumin
Administration of the plasma protein fraction of 5% serum albumin solutions should be
restricted for
the treatment of documented hypoproteinemia
conditions such as burns and peritonitis, in which hypoproteinemia is likely.
Albumin's osmotic ability draws fluid into the vascular space from other extracellular fluid
compartments.
Fluids such as 0.9% saline or lactated Ringer's solution, should be given along with.

49. Immunoglobulin
They may be administered to humans and are obtained from:
Pooled human plasma
1. Human normal immunoglobulins
long-term replacement therapy in antibody deficiency disorders
in short, high dose courses, as immunomodulatory agents in a range of autoimmune or
inflammatory diseases.
Intramuscularly for prophylaxis of certain infections, e.g. measles, hepatitis and rubella in
pregnant women.

50. Hyperimmune immunoglobulins
These are obtained from donors with high titres of specific antibodies and are used for
passive immunization against particular infections.
They are given intramuscularly .
Available against hepatitis B, tetanus, rabies, Rhesus D antigen and herpes viruses.
They are screened for hepatitis and HIV infection.

51. Conclusion
Whole blood is a precious resource from which blood components and plasma
derivatives are derived.
 Blood components include red cell concentrates, platelet concentrates, fresh
plasma and cryoprecipitate. Plasma derivatives include albumin, coagulation
factors and immunoglobulins.

52. Conclusion
The collection and storage of blood components and plasma derivatives has
many phases that are specific to type. These are in place to ensure that the
blood product is safe and effective.
 Transfusion of blood components and plasma derivatives is not without risk
and requires careful consideration.

53. Blood Substitutes

54. A blood substitute (also called artificial blood or blood surrogates) is a substance used to
mimic and fulfill some functions of biological blood.
It aims to provide an alternative to blood transfusion, which is transferring blood or
blood-based products from one person into another.
Thus far, there are no well accepted oxygen-carrying blood substitutes, which is the typical
objective of a blood (RBC) transfusion;

55. The main categories of 'oxygen-carrying' blood substitutes being pursued are
Hemoglobin-based oxygen carriers (HBOC)
Perfluorocarbon-based oxygen carriers (PFBOC)

56. Oxygen-carrying substitutes
An oxygen-carrying blood substitute, sometimes called artificial haemoglobin, is an artificially
made red blood cell substitute whose main function is to carry oxygen, as does natural
haemoglobin.
The use of oxygen-carrying blood substitutes is often called oxygen therapeutics to
differentiate from true blood substitutes.
The initial goal of oxygen carrying blood substitutes is merely to mimic blood's oxygen
transport capacity.
There is additional longer range research on true artificial red and white blood cells which
could theoretically compose a blood substitute with higher fidelity to human blood.

57. History
After William Harvey discovered blood pathways in 1616..
many people tried to use fluids such as beer, urine, milk, and animal blood as blood substitute
The demand for more blood substitutes began during the Vietnam Wars
 The first approved oxygen-carrying blood substitute was a perfluorocarbon-based product
called Fluosol-DA-20, manufactured by Green Cross of Japan in 1989.
 Because of limited success, complexity of use and side effects, it was withdrawn in 1994.

58. History
a haemoglobin-based oxygen therapeutic called Hemopure was approved for Phase III trial (in
elective orthopedic surgery) in the U.S
In December 2003, a new haemoglobin-based oxygen therapeutic, PolyHeme, began field tests
in a Phase III trial on emergency patients (in trauma settings) in the U.S.
In 2013, IIT Madras was approved to mass-produce artificial blood

59. Advantages over human blood
Increasing demand over donation of blood.
A disease-free source of blood substitutes specially in endemic areas
In battlefield scenarios, a safe, easy way to manage blood supply
Rapid treatment of patients in trauma situations(no need of crossmatch)
Protection from prion-transmitted diseases
More shelf life, that too at room temperature !!
Blood substitutes allow for immediate full capacity oxygen transport, as opposed to transfused
blood which can require about 24 hours to reach full oxygen transport capacity
Alternative for those patients that refuse blood transfusions for religious or cultural reasons,
such as Jehovah's Witnesses.

60. Risks
Haemoglobin-based blood substitutes may increase the odds of deaths and heart attacks
30% increase in the risk of death and about a three fold increase in the chance of having a
heart attack for the recipients.

61. Haemoglobin-based oxygen
carriers
It is made of chemically stabilized, cross-linked bovine (cow) haemoglobin in a salt solution.
Many safety measures are taken to render free of pathogens, including herd control and
monitoring.
Hemopure molecules can be up to 1/1,000 the size of RBCs, facilitating oxygen transport and
off-loading to the tissues.
Hemopure is currently in Phase III clinical trials in the US, and is approved for use in South
Africa in surgical patients who are anemic, thereby reducing or eliminating the need for blood
transfusions for these patients.

62. Oxyglobin
approved for veterinary use.
 It consists of chemically stabilized bovine haemoglobin in a balanced salt solution and
contains no red blood cells.
The cross-linked haemoglobin, several tetramers bound together, works by circulation in the
plasma and supplying oxygen to tissues.
 Introduced to veterinary clinics and hospitals in March 1998 and nationally distributed by
October 1998,

63. Polyheme:
The only blood substitute to have completed a Phase III trial
PolyHeme uses human haemoglobin as the oxygen-carrying molecule in solution, and the
extraction and filtration of this haemoglobin from red blood cells is the first step in production.
Then, using a multi-step polymerization process, the purified haemoglobin is associated into
tetramers.
as the final step, is incorporated into an electrolyte solution.

64. Hemospan:
This technology relies on coupling with polyethylene glycol (PEG) to eliminate the toxicity
associated with free haemoglobin.
Yielding positive results in both Phase Ib/II and Phase II clinical trials.

65. Dextran-Haemoglobin
A conjugate of the polymer dextran with human haemoglobin molecules.
Conjugation of haemoglobin to dextran increases its half-life inside the body
Dextran prevents tissue damage that occurs with free haemoglobin from processing by the
kidneys

66. Hyperbranched polymer-
protected porphyrins
University of Sheffield created artificial blood "from plastic“
The "plastic" blood consists of an iron-containing porphyrin which is permanently bonded to a
hyperbranched polymer (HBP or dendrimer) "shell" which protects the fragile porphyrin.
This is similar to hemoglobin in blood, which consists of an iron containing porphyrin reversibly
bonded to a protein "shell".
This mimics natural heme-proteins more accurately, as well as providing the ability to recycle
the HBP when the contained porphyrin becomes damaged

67. Per fluorocarbon based
Perfluorochemicals will not mix with blood, therefore emulsions must be made by dispersing
small drops of PFC in water.
This liquid is then mixed with antibiotics, vitamins, nutrients and salts, producing a mixture
that contains about 80 different components
PFC particles are about 1/40 the size of the diameter of a red blood cell (RBC). This small size
can enable PFC particles to traversecapillaries through which no RBCs are flowing.
PFC solutions can carry oxygen so well that mammals, including humans, can survive breathing
liquid PFC solution, called liquid breathing.

68. Perfluorocarbon based
PFCs in solution act as an intravascular oxygen carrier to temporarily augment oxygen delivery
to tissues.
PFCs are removed from the bloodstream within 48 hours by the body's normal clearance
procedure for particles in the blood – exhalation.
PFC particles in solution can carry several times more oxygen per cubic centimeter (cc) than
blood, while being 40 to 50 times smaller than haemoglobin.

69. Per fluorocarbon based
OXYGENT:
PFCs are surrounded by a lecithin surfactant in a water-based solution.
The lecithin is digested intracellularly.
 Oxygent has done well overall in most clinical trials, but recently, a cardiac surgery study
found participants to be slightly more likely to suffer a stroke if treated with Oxygent

70. Per fluorocarbon based
Oxycyte
Successful small-scale open label human trials treating traumatic brain injury at Virginia
PHER-O2
In research
Perftoran
Approved an authorized blood substitute in Mexico in 2005.

71. THANK YOU