Blood Component and its storage

Surendra
 Utility of Blood Component
• economy of blood, as one unti of blood can be seperated into
different component
• Minimise the hazards of whole blood transfusion
 Blood Component
• Whole Blood
• Irridiated whole blood
• Washed whole blood
• PRBC
• Leukocyte reduced PRBC
• FFP
• PRP
• PC
• Cryoprecipitate
 Defination of component
• Whole blood: It contains all the elements of blood plus
anticoagulant –preservative commonly used as a source for
component production. Min hematocrit should be 33%.

• Irradiated Blood: Irradiated blood & components shall be

prepared by a method known to ensure that irradiation has
occurred using Cesium137 or Cobalt 60 as a radiation source.
• Red Blood Cells : RBCs are units of Whole Blood with most
of the plasma removed. If prepared from whole blood
collected into CPD,CP2D, or CPDA-1, the final hematocrit
must be ≤80%.

• RBC Leucocytes Reduced: It is prepared by in-line

filtration of WB or red cells. RBC leucocyte reduced should
contain <5x106 leucocytes and retain 85% of the original red
cells.

• Washed Red Cells: Washing of red cells with normal saline

removes 70-95% leucocytes. Plasma proteins and
microaggregates are also removed. There is loss of 15-20 ml
of RBC
• Platelet Concentrate: Platelet collected from one unit of
whole blood and resuspended in an appropriate volume of
normal plasma. Source may be PRP(5.5x1010 )or by
plateletpheresis (3.0x 1011 )

• Granulocyte Concentrate: A unit of granulocytes having

leucocytes of 1x 1010 prepared by cells separator. Shelf life is
24 hours at 20-24 0 C.

• Fresh Frozen Plasma: Plasma frozen within 6 hours of

collection and stored at less than -30 0 C for a period of not
more than one year.
• Thawed Plasma: If thawed FFP is not transfused within 24
hours, it must redesignated as thawed plasma and can be
stored upto 5 days at 1-6 0 C

• Plasma And Liquid Plasma: Plasma separated from a WB at

any time of storage up to 5 days after expiration and stored at -
180 C. If not frozen but stored at 1-60 C and transfused within
5 days after the expiration that is called liquid plasma. If FFP
is not used within 12 months, it can be redesignated as plasma
and stored up to 4 years.
• Cryoprecipitate: They are precipitated proteins of plasma
rich in factor VIII & fibrinogen, VWF & fibronectin obtained
from a single unit of fresh frozen plasma. It is stored at -30 0 C
or below for 1 year.

• Plateletpheresis: A process by which blood drwan from a

donor after platelet concentrate has been prepared, is
retransfused simultaneously into the said donor. Platelet count
≥3.0xx1011 per unit

• Plasmapheresis : A process by which blood drwan from a

donor after plasma has been prepared, is retransfused
simultaneously into the said donor.
Method of component preparation
• Gravity separation: old time, crude but cheaper method to
separate plasma from whole blood

• Low speed refrigerated centrifugation: used for preparing

Platelet Rich Plasma (PRP)

• High speed refrigerated centrifugation : used for preparing

Platelet Concentrate (PC), Fresh Frozen Plasma (FFP) and
Cryoprecipitate (CP).

• Apheresis by cell separator:

 Principle of Component Preparation
The basic principle of component preparation lies in the fact that
red cells plasma and platelets have different specific gravity. They are
separated using differential centrifugation i.e. applying different relative
centrifugal force (g) for different time period (revolution /minute).

what is “g”.
A centrifuge uses centrifugal force to isolate suspended particles
from surrounding medium. When a suspension is rotated at a certain
speed(revolution/minute), centrifugal force causes the particle to move
away from the axis of the rotation. The force applied on the
particles(compared to gravity) is called relative centrifugal force( “g”).
Specific gravity of various blood components are:
RBC : 1.08
Whole blood : 1.053
Platelet : 1.035
Plasma : 1.025
Refrigerated centrifuge , rotor speed and duration of spin( time period ) are
critical in component preparation.
 Relationship Between RCF(g) and RPM

rpm 2
rcf (in g) = 28.38 × R × 1000 where “R” is the radius of the rotor of the
centrifuge, “ rpm “ is the revolution per minute and “ 28.38 ” is a constant

Types of Centrifuge Rotors:

Centrifuge Rotors can be broadly classified into :
1. Vertical Rotor: Sample tubes are held in vertical position during rotation. It is
used for isopycnic (Same density) separation due to short path length.
2. Fixed Angle rotor: Sample tubes are held at fixed angle of the rotor cavity.It is
used for pelleting application.
3. Swinging Bucket Rotor: Buckets are held in vertical position during rest. Buckets
swings out to horizontal position during rotation providing longer path length
which permits better separation of particles . It allows easy separation of
supernatant without disturbing pellet. Blood component separation centrifuge
uses swinging bucket rotor
Centrifuge Rotors
Centrifugation: Principle Of Separation

Platelets 1.035

Lymphocytes
1.050-1.061

Monocytes
1.065-
1.069
Granulocytes
1.087-
1.092
RBC
1.10
Blood Components Separation Centrifuge
Components From Whole Blood
Specific Gravity of Blood Components
 Component Preparation Procedures
Step 1
Step 2

Step 3
 Methods Of Component Preparation
1. PRP-PC Collect WB in CPDA-1 ,450ml
triple bag

SOFT SPIN AT 1750 rpm for 10

min at22⁰C

PRBC PRP

HARD SPIN AT 3940 rpm

for 5 min at 22⁰C

FFP PC in 50 – 70 ml plasma
Buffy Coat Components
 Pooling Of Buffy Coats

Plasma
Semi-automated Blood Components Separator
 Important Factors Affecting Blood Component
I. Venipuncture:
Blood should be collected from a healthy donor by a clean ,aseptic single
venipuncture technique with minimal trauma to tissue. Blood should flow rapidly
and the collection time should be 4 -10 minutes. There should be frequent and
gentle mixing of blood with anticoagulant.
II. Collection Container:
To simplify the the separation of whole blood into its component parts,
blood is collected in PVC plastic bags attached with one or more satellite bags. To
remove the brittleness of PVC bags, plasticizers are incorporated into a plastic to
increase its flexibility.
Plastic bags intended for platelet storage should be sufficiently
permeable to gases to guarantee availability of oxygen to platelets and diffusion
of carbon dioxide . The amount of oxygen required is dependent on the number of.
Lack of oxygen increases anaerobic glycolysis and lactic acid production.
Types of Blood Containers

Fig:Triple bag system

Quadruple Bag System- Top and Top Bag (TAT)

Quadruple bag system(TAB)

Fig. A Top and Bottom Bag
 Some Common Types of Plasticizers and Their Uses

1. DEHP(Di-[2-Ethyihexyl] Pthallate 1. Maintains membrane flexibility of RBC

for improved survival. Use for RBC
storage and also for Platelet storage for
three days.

2. BTHC(Butryl-Tri-n Hexyl)Citrate
2. Less toxic than DEHP. Use for RBC
storage.

3. TEHTM(Tri-[2EthylHexyl]Trimellitate 3. Has sufficient gas permeability for

Oxygen and Carbon Dioxoide.
Can store platelets for seven days if
bacterial contamination is prevented.
4. Polyolefin without Plasticizer
4. For Seven days storage of platelets if
bacterial contamination is prevented.

5. DEHA (Diethylhexyladipate)
5. Less toxic than DEHP

6. DINCH, (Diisononylester of
6. Tradename is Hexamoll. Less toxic and
Cyclohexanedicarboxylic Acid) odorless unlike DEHP
III. Transportation (Shipping) of blood from collection site to the
component preparation facility:

 Blood for preparing platelet concentrate should be transported at

20-24⁰ C.
 Below 20 °C platelets undergo membrane phase transition and cold
activation and the discoid platelet structure gradually converts to a
sphere.
 Blood for preparing FFP and PRBC should be transported at 2- 10⁰C. At
higher temperatures, labile factors are considerably lost.
Therefore, cold chain must be maintained to preserve the
morphological and functional properties of the blood constituents.
 Cryofuge Programs

Use
RPM/ Temp Time
No Ac/Dc
RCF (oC) (min) Top & bottom Double
Top & Top Quad Triple Bag
Quad Bag
1st spin
3650 /
1 22 10 8/3 (SPRBC + FFP +
4423
Buffycoat)
1750 / 1st spin PRBC
2 22 11 9/4
1017 (PRBC + PRP) PRP
2nd spin buffycoat
1050 /
3 22 5 8/3 (BCPC +
366
Buffycoat)
1st spin
3950 / 2nd spin
4 22 5 9/4 (SPRBC + FFP +
5181 (FFP + PRPPC)
Buffycoat)
1st and 2nd spin
3950 / PRBC
5 4 5 9/4 Triple Bag
5181 FFP
Cryo/CPP

1100 / 2nd spin buffycoat

7 22 6 9/4
402 (BCPC + Buffycoat)
 Whole Blood
450 ml of blood
63 ml of anticoagulant solution.
Hct-36-44%
No components have been removed.
Store at 1-6 oC
Shelf life-
Citrate-Phophate-Dextrose (CPD) - 28 days
CPDA-1 (adenine) - 35 days
SAG-M – 42 days
Administer through standard blood filter (150-280 micron)
Infuse within 4 hours of issue
 Whole Blood

• Drawbacks:
– After storage for >24 hours, platelets and WBC are non-
functional
– Factor V and VIII decrease with storage
– Fluid overload
• Indications:
– Acute blood loss > 25% TBV
– Exchange transfusion
Contraindication
- Risk of volume overload : Chronic anemia
Incipient cardiac failure
 WHOLE BLOOD

Precaution in whole blood transfusion

- Must be ABO and RhD compatible
- Never add medication to a unit of blood
- Use blood administration set

Dosage
1 unit  Hct 3 % or Hb 1 g / dL
 Packed Red Cell

• units with red blood cells and some plasma

• with Anticoagulant ACD / CPD / CPDA – 1
• Hct ~ 75 – 80 %
 Preparation of Packed RBCs
Principle:
RBCs are obtained by removal of supernatant plasma from centrifuged whole
blood.
Preparation:
Centrifuge whole blood unit in refrigerated centrifuge containing the
parameters RPM-3850,Time-5 Min, Temp- 40C
Express the supernatant plasma with the help of plasma expressor.
Double seal the tubing between the primary and satellite bag.
Check that the satellite bag has the same donor number as that on the primary
bag and cut the tubing between the two seal.
Advantages:
Oxygen carrying capacity equal to that of whole blood in half the volume.
Significantly decrease levels of isoagglutinins, metabolites and electrolytes.
 Preparation of Packed RBCs
Shelf life: (If CPDA1 anticoagulant) -35 days
Storage temp. : 40C (Range is 2-60C)
QC Requirements: PCV 80% (Range is 65-80%)
Volume: 250- 300 ml.
CONTENTS: Red cells- 65-80%
Plasma – 20-35%
Some platelets, white cells storage lesion by products
and anticoagulant preservative solutions.
Transfusion Criteria's: ABO/Rh specific and compatible
Indications: Restore oxygen carrying capacity symptomatic anemia and
surgical blood loss.
Effect: 1 unit RBCs should raise HCT -3%, Hb 1 g/dl
 Packed Red Cells

Indication
- Replacement of red cells in anemic patients
- Use with crystalloid or colloid solution in acute blood loss

Dosage 10 - 15 ml / kg
PRC 1 unit  Increase Hct 3 % or Hb 1 g/dL
 Guidelines for
Transfusion of Packed Red Cells in Adults
Transfusion for patients on cardiopulmonary bypass with
hemoglobin level ≤6.0 g/dL is indicated.
Hemoglobin level ≤7.0 g/dL in patients >65 years and
patients with chronic cardiovascular or respiratory diseases
justifies transfusion.
For stable patients with hemoglobin level between 7 and 10
g/dL, the benefit of transfusion is unclear.
Transfusion is recommended for patients with acute blood
loss more than 1,500 mL or 30% of blood volume.
Evidence of rapid blood loss without immediate control
warrants blood transfusion.
 Red cell aliquots

• For babies
• 10-25 mL units.
• 5 mL/kg will raise Hb by ~1 gm/dL.
 Irradiated Red Cells
• Gamma-radiated to kill the lymphocytes.
• The lack of T-cells prevents graft-vs-host disease.
• Use for
• Severely immunocompromised patients
• Lymphoma patients
• Stem-cell / marrow transplants
• Intrauterine transfusion
• Units from close “blood relatives”
• Neonates undergoing exchange transfusion or ECMO
• Hodgkin’s Disease
Leukocyte poor red blood cell

Unit of red blood cells with some plasma,

anticoagulant and additive solution
Hct ~ 55 – 65 %
Leukocyte poor red blood cells

Centrifugation method
- easiest and least cost
- least efficient
- reduce WBC only 70 - 80%
Leukocyte depleted red blood cells
Filtration method
easy, quick, but more expensive
high efficient
remove WBC more than 99.9%
( third generation )
little loss of RBC volume

42
 Leukocyte reduced red cells
Indication
- Minimizes white cell immunization in patients
- Prevention of FNHTR (Febrile Non-Hemolytic Transfusion Reaction )
- Reduces risk of CMV transfusion
Contraindication
- Not prevent graft –vs- host disease
Dosage
- same as Packed Red Cell
Administration
- same as Whole Blood
 Granulocyte Concentrate
• Obtained by apheresis from family members
for administration to cancer patients.
• Contain 1.0 x 1010 granulocytes
• Pre-treatment with recombinant G-CSF and
dexamethasone can yield 4-8 x 1010
granulocytes
• Stored at 24o C
• Infuse within 24 hours of collection
 Criteria

• ANC <500
• Fever
• Documented infection (bacterial or fungal) for
24-48 hours
• Unresponsive to appropriate antibiotics
• Reasonable hope of marrow recovery
 PRP
• It is prepared from the whole blood within six-hours of
collection, preferably stored at room temperature of 20-
24°C.
• 1750 rpm for 11 min
• PRP for storage can be kept at 22-24°C in a, platelet incubator
with constant agitation for a maximum of 48-72.hours
• must contain 5.5 x 1010/ bag
• one bag of PRP generally raises the platelet count in the
recipient by 5000-10000 / µl.
• Aspirin and related analgesics affect the platelet function, so
the donor for platelets is accepted after 3 days of ingestion of
these drugs.
 Platelets Concentrate (PC)
This supplies the same amount of platelets as PRP, but in lesser
volume (40 - 50ml).

Principle:
Platelets are harvested from whole blood following ‘light spin‘ centrifugation. The
platelets are concentrated by 'heavy spin' centrifugation with subsequent removal of
supernatant plasma.
Steps
1. Blood is collected in the triple bag system only
2. PRP is prepared by following the above mentioned steps
3. After detaching the satellite bags from the primary bag, the PRP in again spun at
4000 - 5000 rpm (high spin) for 4 to 5 minutes.
4. A platelet button is formed at bottom and platelet poor plasma is expressed from the
1st satellite bag to the 2nd satellite bags, leaving behind 40 - 50 ml plasma for
platelet button suspension.
5. The platelet concentrate in stored at 20°C-24°C for a maximum of 3 days (at PGI) with
constant agitation.
 Platelets Concentrate (PC)
Shelf life: 3 days in platelet incubator & agitator.
24 hrs if no storage cabinet
Storage temp.: 20°C - 24°C
Q.C. Requirements: To be prepared within 8 hrs
aftercollection, pH should be 6.2 or more
at the end of storage time. Platelet count
> 5.5 x 1010 /unit.
Volume: 30 to 50 ml
Contents: Platelet - 5.5 x 1010 /bag
Plasma - 30 to 50 ml and some white cells
Transfusion Criteria: ABO / Rh specific and compatible
Indications: Severe thrombocytopenia, qualitative platelet
defects
Effect: Increases in platelet count 10,000 / ul per unit
 PLATELET CONCENTRATE

• Dosage
• 1 unit of PC / 10 kg B.W.
• Increment will be less in
- Hypersplenism
- DIC
- Septicemia

1 unit of PC  Platelet 5000-10,000 / ul

 PLATELET CONCENTRATE

• Administration should be ABO & Rh

compatible
• After pooling, should be infused as soon as
possible
• Use blood administration or platelet infusion
set
• Must not be refrigerated before infusion
Random Donor Platelet

Volume 45 – 65 ml
Single Donor Platelet

Volume ~ 300 ml
 Single-donor platelets
– Obtained by plateletpheresis technique.
– 6 - 8 times as many platelets as in a random-donor
unit.
– Larger volumes and HLA-compatibility results in an
increase of 30k-60k.
– Leukoreduced because of apheresis collection
– ABO matched platelets preferable
– Rh negative receive Rh negative platelets
 Single Donor Platelet

• Indication
– same as random PC
– special requirement  obtain from selected donor

• Dosage
• Usually 1pack of SDP = 1 therapeutic dose
 Plasma Components

– Fresh Frozen Plasma

– Frozen Plasma :- Aged plasma
– Cryoremoved plasma
– Cryoprecipitate
 FFP
• It is prepared from the whole blood collected
in a CPDA-1 double or triple bag system within
6-8 hours of its collection.
• contains coagulation factors and other plasma
protein (per unit or bag)
Shelf life: One year
Storage temp.: -20°C or below
 Indications for FFP Transfusion
Clinically significant deficiency of Factors II, V, X, XI
DIC
Plasma exchange
Immunodeficiencies
Massive transfusion of stored blood.
Liver disease
Urgent reversal of warfarin therapy
Correction of known coagulation factor deficiencies for which specific
concentrates are unavailable
Correction of microvascular bleeding in the presence of elevated (> 1.5 times
normal) PT or PTT
Correction of microvascular bleeding secondary to coagulation factor
deficiency in patients transfused with more than one blood volume and when
PT and PTT cannot be obtained in a timely fashion
 FFP
• Dose
– 10-15 ml/kg B.W
– For warfarin reversal, 5-8 ml/kg of FFP

• Contraindication
– Volume expansion
– Immunoglobulin replacement
– Nutritional support
– Wound healing
 FRESH FROZEN PLASMA

• Precaution
– Acute allergic reaction are common
– Anaphylactic reaction may occur
– Hypovolemia alone is not an indication for use
• Dosage
Initial dose of 15 - 20 ml / kg B.W
 PRINCIPLES OF APHERESIS

Remaining blood
components recombine &
ACD return
Plasma

Platelets
WB in vein Lymphocytes WB in vein
Granulocytes

Erythrocytes

Blood components separated by centrifugation &

selectively removed