Blood component preparation

Kaushik Sen
MLT, ESIC MED COLLEGE
 Need for separation
• Optimal survival of each constituents.
– In whole blood stored at 2-6°C, platelets stay
viable for 1 day and factor V & VIII decrease.
– While after separation, platelets stay upto 5 days
and factor V and VIII can be stored as FFP for 1 yr
at -30°C
• Transfusion of only required component thus
avoids the use of unnecessary component
which could be contraindicated in a patient.
• One donor can save several patients.
 Blood
components

Cellular Plasma Plasma derivatives

Albumin 5%, 25%,

PC, Platelets, FFP, Factor VIII,
others Cryoprecipitate coagulation
factors
 Blood Component

Packed Fresh Frozen Plasma Platelet Concentrate Cryoprecipitate

RBC (FFP)

pH should be
>6.0

RDP SDP (Apheresis)

Granulocyte
Platelet Yield 5.5 x 𝟏𝟎𝟏𝟎 Platelet Yield 3 x 𝟏𝟎𝟏𝟏
Concentrate

WBC Yield 1 x 𝟏𝟎𝟗 Plt from buffy coat

Platelet Yield >2 x 𝟏𝟎𝟏𝟏

 Whole Blood

PRBC FFP Plt. Concentrate Cryoppt.

 Precautions
A. In collection of blood:
1. Proper selection of donor
2. Clean aseptic site of venipuncture
3. Uninterrupted flow of blood
4. Collect correct amount of blood
5. Automatic mixing of blood during collection
6. Separate platelets within 6-8 hours
7. triple pack and quad pack system
 Precautions
B. In centrifugation
1. Balancing & use of rubber discs
2. Broad side of the bag faces the outside wall of
the cup
3. Correct speed and time to be maintained
4. Look for any abnormal vibration
 Precautions
• During blood collection:
– Flow should be continuous
– 450ml bag should not take more than 8 mins to fill
• During processing:
– Freezing the plasma as soon as it is separated
– Maintaining the storage temperature
• During thawing:
– The entry ports of the bag should remain above the
water
– Should be administered within 12 hours if kept at 2-
6°C.
 BLOOD COMPONENTS
INDICATIONS,PREPARATION OF
BLOOD COMPONENTS

1
 INTRODUCTION
• These days effective blood transfusion
depends upon the availability of different
blood components.
• These components are used in separately or in
combinations, can meet most patients
transfusions need and keeping the risk of
transfusion to a minimum.

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 WHY THERE IS NEED FOR
COMPONENT SEPARATION
• 1. Separation of blood in to components allows
optimal survival of each constituents.

• 2. component preparation allows transfusing only

specific blood component that the patient requires.

• 3. Transfusion of only specific constituent of blood

needed avoids the use of unnecessary component,
which could be contraindicated in a patient

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• 4.By using blood components, several patients
can be treated with the blood from one donor,
giving the optimal use of every unit of
donated blood.
• 5. use of blood components, supplements
blood supply- adds to the blood inventory

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 BLOOD COMPONENTS

Cellular components :
• Red cell concentrate
• Leucocytes-Reduced Red cells
• Platelet Concentrate
• Leukocytes-Reduced Platelet Concentrate
• Platelet Apheresis
• Granulocytes Apheresis

5
 Plasma components

• Fresh frozen plasma

• Single donor plasma
• Cryoprecipitate
• Cryo-poor plasma
• Plasma derivatives
Albumin 5% & 25%

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 Plasma Protein Fractions

• Factor VIII concentrate Immunoglobulins

• Fibrinogen Other coagulation factors

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 PREPARATION OF BLOOD
COMPONENTS
• Equipment:
• Freezer - 40°C and - 70°C
• Blood bank refrigerator 2-6°C
• Refrigerated centrifuge with swing out head and oval
cups
• Laminar flow
• Weighing scale
• Dielectric sealer or aluminium clips & sealer
• Stripper
• Water bath 37°C or Plasma Defroster (MicroWave)
• Platelet reciprocator (platelet incubator with
agitator)
• Cryoprecipitate bath 4°C

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 Equipments
• Freezer: – 40°C and -70°C
• Blood bank Refrigerator 2-6°C
• Refrigerated centrifuge with swing out head and oval
cups
• Laminar flow
• Weighing scale
• Dielectric sealer or aluminium clips & sealer
• Stripper
• Water bath 37°C or plasma defroster (Microwave)
• Platelet agitator
• Sterile docking device
Primary bag with satellite bags
COLOR CODING FOR BLOOD BAG
COLOR CODING FOR BLOOD BAG

The printed labels are colour coded for all

component as per Blood Group A- yellow,
group B- pink , group O- blue and group AB-
white . Negative labels also have the same
color code.
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Platelet agitator

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CRYOCENTROFUGE

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CRYOCENTROFUGE

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Loading the blood collection pack into a centrifuge bucket
(a) and centrifuge (b). Note that the heaviest items are
placed towards the outside of the centrifuge.
Folding of satellite bag. Extra tubing is placed
inside bag folds.
Weighing machine to balance the buckets
DONOR BED
SUNIL KUMAR.P 14
EXPRESSOR
a. Set up for plasma expression in plasma press. Note the use
of the bag eyelets for vertical stable positioning of the bag in
the plasma press (arrows). b. Spring plate is unlatched to
apply pressure to blood donation. When connector seal (black
arrow) is broken, plasma is allowed to flow into satellite bag. c.
Spring plate is latched to remove pressure from blood
donation at the end of the plasma separation process.
BLOOD BAG TUBE SEALER
a–c. Dielectric (heat)
sealing.
 a. Hand sealer. b–e. Correct use and end result of line
sealing with rectangular metal clips. f, g. Incomplete line
sealing due to hand sealer exerting uneven pressure on
the clip.
Tools for temporary clamping include plastic haemostat
clamps (top) or plastic clips (bottom) for use during
centrifugation.
 Preparation of blood components is possible due
to :
• Multiple Plastic packs system
• Refrigerated centrifuge
• Different specific gravity of cellular components
• Red cells specific gravity 1.08 - 1.09
• Platelet specific gravity 1.03 - 1.04
• Plasma specific gravity 1.02 - 1.03

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16
 Methods of preparation

• There are various methods to separate these

components and the yield and quality of
component depends upon the methods
applied.
Various methods used are:
• a)Gravity separation
• b)Low and high speed centrifugation
• c)Apheresis by cell separator

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 WHOLE BLOOD

• 350-450 ml of blood
• 49-63ml of anticoagulant solution
• Hct- 36-44%
• No components removed
• Stored at 1- 6 degree celsius
• Shelf life :
• Citrate –phosphate –Dextrose(CPD)- 21days
• CPDA (adenine) – 35 days
• AS-1, AS-3, AS-5 – 42 days

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• Increase Hb by 1 g/dl
• Rate of Blood Transfusion -3ml/kg/hr

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 WHOLE BLOOD Indications
• Acute active blood loss with hypovolemia
• Exchange transfusion Contradictions
Risk of volume overload
• chronic anemia
• Incipent cardiac failure

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 WHOLE BLOOD Drawbacks

• After storage for 24 hours platelets and wbc

are non functional
• Factor V and Viii decrease with storage
• Fluid overload

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 Preparation of Packed RBCs Principle

• RBCs are obtained by removal of supernatant plasma

from centrifuged whole blood
Preparation
• 1. Collect appropriate volume of donor blood in CPDA
double or triple bag.
• 2. Store at 2-6C till processed.
• 3. Place bags in the buckets of refrigerated centrifuge
and balance the opposite bags accurately.
• 4. Centrifuge at heavy spin (5000 x g) for 5 minutes at
2-6C.

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• 5. Express approximately 3/4 of the plasma
into the satellite bag.
• 6. Double seal the tube between primary and
satellite bags with plasma Separate the
satellite bag with plasma and keep at -30°C or
below.
• 7. Keep the red cells at 2-6°C.

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 Indications

• Patient need urgent operation and has Hb < 10g/dl

• Anticipated surgical blood loss > 1000 ml
• Other acute Loss of blood
• Replacement Fluid blood loss < 20% of blood vol.
• Anemia associated with incipient/established cardiac
failure
• Hb value < 6 g/dl
• Patients approaching delivery and has Hb value < 7g/dl
• In hereditary hemolytic anemias and betathalassaemia
major.

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 Packed RBCs
• Shelf life – 35 days
• Storage temp- 2 – 6 Degree celsius
• QC Requirements: PCV 80% (Range 65-80%)
• Volume- 250- 300 ml
Contents :
• red cells- 65-80%,
• Plasma 20- 35%,
• Some platelets,
• white blood cell and anticoagulant preservative
solutions Effect – 1 unit RBC raise Hct by 3% and hb by
1g/dl

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27
 Guidelines for Paediatric red cell
transfusions
Anaemia
Infants within first 4 months of life
• Hb< 13.0 g/dL and severe pulmonary disease
• Hb < 10.0 g/dL and moderate pulmonary disease
• Hb< 13.0 g/dL and severe cardiac disease
• Hb<10.0 g/dL and major surgery
• Hb < 8.0 g/dL and symptomatic anemia
• Acute blood loss >10% of total blood volume.

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 Guidelines for Paediatric red cell
transfusions
Infants >4 months & children
• Acute loss > 25% circulating blood volume
• Hb < 8.0 g/dL in perioperative period
• Hb < 13.0 g/dL and severe cardiopulmonary
disease
• Hb < 8.0 g/dL and marrow failure
• Hb < 8.0 g/dL and symptomatic chronic
anemia
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 IRRADIATED RED BLOOD CELLS
• Gamma radiated to kill the lymphocytes
• The lack of T cells prevents graft versus host
reaction
Indications:
• Severely imunocompromised patients
• Lymphoma patients
• Stem cell/marrow transplants
• Intrauterine transfusion
• Neonates undergoing exchange transfusion
• Hodgkin Disease
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 LEUCOCYTE REDUCED RBCS
Methods of the preparation of Leucocyte-
Reduced Red cells
• Centrifugation and removing of buffy coat
• Filtration
• Washing of red cells with saline
• Freezing and thawing of red cells

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 LEUCOCYTE REDUCED RBCS
INDICATIONS
• Multi transfused patients like thalasemia
• Leukemia
• Aplastic anemia
• Immunosupressed
• Immunodeficient
• Prevention of recurrent FNHTRs
• Prevention or delay of primary alloimmunization
to HLA antigen
• Prevention of CMV transmission in at risk
individual
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 PLATELET RICH PLASMA (PRP)
Procedure:
• 1. Collect 450 ml blood by a clean, single venipunctuer
into 450 ml CPDA or Adsol / SAGM triple bags system.
• 2. Keep the blood bag at room temperature (20-22°C)
before preparing platelets, for not more than 6 hours.
Do not chill at any time before or during platelet
separation.
• 3. Centrifuge the blood bags at 20-24°C at light spin for
appropriate time (2000 x g for 3 minutes).
• 4. Separate 4/5 of the platelet rich plasma (PRP) into
one satellite bag

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 Procedure
• Collect in triple pack system and store at room temperature at 20-22°C for
1st less than 6 hours.

• Place bag in buckets of refrigerated centrifuge and balance the opposite bags
2nd • Centrifuge at 2000 x g for 3 mins at 20-24°C

• Express approx 4/5th of the platelet rich plasma in the satellite bag
3rd

• Centrifuge the bag with PRP and another satellite bag at 20-24°C at 5000 xg
4th for 5 mins.

• Express supernatant platelet poor plasma into another empty bag and leave
5th approx 50 ml of plasma.

• Store platelet at 20-22°C under constant agitation.

6th
 PLATELET CONCENTRATE
• Shelf life:
1 day if no storage cabinet
5 days in platelet incubator and agitator
• Storage temp :20-24 degree celsius
• Q.C Requirements: to be prepared within 6hr
of collection and ph should be >6.2
• volume: 30 to 50 ml
• Contents: 5.5x1010 unit/bag

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 Precautions
• Agitation during storage helps in:
– Exchange of gases
– Maintenance of pH
– Reduce formation of platelet aggregates
• pH should never fall below 6 because:
– Change in shape of platelets from disc to sphere
– Pseudopod formation
– Release of platelet granules
 Indications
➢Platelet count is < 5000 / µl regardless of
clinical condition
➢Effect: increases platelet count by 10000/ µl
per unit

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43
 PROCEDURE
1.Centrifuge the bag with PRP and another
satellite bag at 20-40°C at ‘heavy spin’ for
appropriate time e.g. 5000xg for 5 minutes.
2.Express supernatant platelet - poor plasma
into another empty satellite bag.

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 SINGLE DONOR PLATELETS
• Obtained by plasmapheresis technique
• 6 to 8 times more platelets as in random
donor unit
• Larger volumes and HLA compatibility results
in increase of 30 to 60 k
• Leucoreduced because of apheresis technique
• ABO Matched platelets preferred

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 FRESH FROZEN PLASMA
• FFP is plasma obtained from
a) a single donor either by normal donation or
b) by plasmapheresis
It contains all coagulation factors and
a) plasma proteins Factor viii – 0.6 IU/ml
b) Factor ix – 0.9 IU/ml
c) Fibrinogen- 250-300 mg/bag Proteins – albumin,
globulin etc 1 IU/kg of factor viii or factor ix raises
factor viii in plasma by 2% and factor ix by 1%
d) Volume: 200 – 250ml
e) Shelf life- 1 year Storage temp(-20 to -70) degree or
below
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 Procedure
• 1. Collect appropriate volume of blood in 350-
450 ml CPDA double bags systems or 450 ml
SAGM/Adsol triple bags system.
• 2. Centrifuge at heavy spin (5000 x g for 5
minutes) at 4°C .
• 3. Express approximately four fifth of the
plasma into a satellite bag.

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• Double seal the tube between primary bag
and the satellite bag having plasma with metal
clips or dielectrical sealer.
• Separate the satellite bag having plasma

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Indications of Fresh Frozen Plasma
• Actively bleeding and multiple coagulation
factors deficiencies
• Liver diseases
• Disseminated intravascular coagulation (D1C)
• Coagulopathy in massive transfusion
• TTP

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• Antithrombin III deficiency
• Congenital or acquired coagulation factor
deficiency
• Use of FFP in conjunction with red cells has
largely replaced the transfusion of fresh
blood.
• Deficiency of Factors II, VII, IX,, and X

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 SINGLE DONOR PLASMA
• Prepared from stored blood
• Poor in coagulation factors
• Cannot be used in correction of coagulation
factor deficiencies
• Effective as a volume expander

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 CRYOPRECIPITATE
• Shelf life: frozen – 1 year
• Thawed : 6 hours
• Storage temp : frozen -20 degree or less
• thawed : 37 degree
• Volume- 10 to 20 ml

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• Contents –
Factor viii- 80 to 150 units/bag
Fibrinogen- 150 to 250 mg/ bag
Factor xiii- 20 to 30 % of whole blood
Von wilebrand factor- 40 to 70 % of whole blood
• Transfusion criteria :
➢ ABO compatibility not required
➢ Must be infused within 6 hours of thawing

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 Procedure
▪ Prepare fresh frozen plasma (FFP), as described under FFP, for processing into
1st cryoprecipitate.

▪ Freeze the plasma at -70° C in freezer

▪ Thaw frozen plasma at 4°C in circulating water bath.
2nd

▪ Centrifuge the bag when the plasma is slushy at 5000 x g for 5 minutes at
3rd 4°C.

▪ Supernatant cryo-poor plasma is siphoned out in the satellite bag, leaving 10-15 ml
4th plasma with cryoprecipitate. Seal the tubing and separate the bags.

▪ Storage and shelf life of cryoprecipitate : One year at -30°C or below.

5th

6th
 INDICATIONS
• Quantitative and qualitative fibrinogen
deficiency : DIC
• Correction of factor viii deficiency
Haemophilia A
• Von wilebrand disease
• Factor XIII deficiency
• Uremic coagulopathy

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 CRYOPRECIPITATE-POOR-PLASMA
• It is a by-product of cryoprecipitate
preparation.
• It lacks labile clotting factors V and Viii and
fibrinogen.
• It contains adequate levels of the stable
clotting factors II, VII, IX and X.
• It is frozen and stored at -20°C or lower
temperature for 5 years Indications
• Burn
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 Component Temperature Shelf Life
Red Cells 2 -60 C 42 days

Platelets 20-240 C 5 days

Fresh Frozen Plasma -300 C 1 year

Cryoprecipitates -400 C 1 year

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SUNIL KUMAR.P 60
 VIDEO LINK FOR PRACTICAL

https://youtu.be/tCJBdWOA3Ok