coagulation 1
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M.R.T. 4DMT
COAGULATIONHemostasis
- cellular and biochemical events which function in harmony to keep blood with the veins andarteries.
- prevent blood loss from injuries.
- re-establish blood flow.
1. Primary Hemostasis
- involves platelets and endothelium.- activated by small injuries.
- rapid, short-lived reponse.
- positive feedback: formation of platelet plug.
A. Platelets
Production:Hemocytoblast Megakaryoblast Promegakaryocyte
Megakaryocyte Metamegakaryocyte Platelets
Distribution:
a. Peripheral blood 70%
b. Spleen 30%
RR: 150 400 x 10^9 / L
Life Span: 8 11 days
Size: 2.5 m in diameter
Zones:
a. Peripheral zone adhesion and aggregation.b. Sol Gel zone shape and contraction.c. Organelle zone metabolic activities.
Function:
a. Adhesion -platelets roll and cling to non - platelet surfaces.
b. Activation - occurs when vWF and collagen binds to glycoprotein Ib receptoron the surface of the platelet.
c. Aggregation -platelets adhere to each other.
d. Secretion -platelets discharge the contents of their granules.e. Clot Retraction creates more bulk in the clot making it more resistant to
stress.
f. Cytokine signalling -platelets secrete platelet derived growth factor.
B. Endothelial cells
- assist in platelet activation.- limits coagulation mechanism.
- clot dissolution.
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2. Secondary Hemostasis
- involve coagulation factors.- activated by large wounds from trauma, surgery, dental procedures.
- delayed, long-term response.
- positive feedback: formation of fibrin clot.
A. Coagulation
- mechanism whereby after injury to a blood vessel, plasma coagulation factors, tissue factors,and calcium work together on the surface of platelets to form a fibrin clot.
B. Coagulation Factors
- proteins engaged in formation of a fibrin clot from fibrinogen.
Factor Customary name FunctionI Fibrinogen Thrombin substrate, polymerizes to form
fibrin
II Prothrombin Serine protease
III Tissue factor Cofactor
IV Ionic Calcium Mineral
V Labile factor Cofactor
VII Stable Factor Serine protease
VIII Antihemophilic Factor
(vWF)
Cofactor
Factor VIII carrier and platelet adhesion
IX Christmas factor Serine protease
X Stuart-Prower factor Serine protease
XI Plasma thromboplastin antecedent
(PTA)
Serine protease
XII Hageman factor Serine protease
Prekallikrein Fletcher factor, PK Serine protease
High Molecular Weight
Kininogen
Fitzgerald factor, HMWK Cofactor
XIII Fibrin-stabilizing factor (FSF) Transamidase
Platelet factor 3 Phospholipids, phosphatidyl serine
PF3
Assembly molecule
C. Coagulation Cascade
- series of biochemical reactions and feedback mechanisms by means of intrinsic and extrinsic
pathway, or both, leading to a common pathway forming a fibrin clot.
C.1. Intrinsic pathway
- utilization of plasma contact factors to initiate coagulation, beginning with theactivation of factor XII; all necessary factors required are contained in the circulating
blood.
- activated partial thromboplastin time (aPTT) test monitors this pathway.
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- aPTT measures factors XII, XI, X, IX, VIII, V, II and I.
C.2. Extrinsic pathway
- coagulation pathway that is activated by tissue thromboplastin; necessary components
are factor VII and calcium.- prothrombin time (PT) test monitors this pathway.
- PT measures factors VII, X, V, II, I.
C.3. Common pathway
- final stage of the coagulation cascade, beginning with the convergence of the extrinsic
and intrinsic pathways (factor X) ending with the formation of fibrin clot.
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3. Specimen Collection and Handling
A. Anticoagulants
- 3. 8 % or 3.2 % sodium citrate- normal ratio of blood to anticoagulant is 9:1
B. Holding sample
- siliconized glassware (glass activates factor XII and platelets will adhere to glass).
C. Collection of Blood Sample
- venipuncture should be clean and quick with minimal stasis.
- if drawing tubes for other tests, draw blue top last, except that if an EDTA tube is needed for a
CBC, the purple top tube should be drawn after the blue top coagulation tube. If only drawingblood for coagulation testing, allow some blood to drip into vacutainer before collection. (The first
blood drawn in a venipuncture is most likely to be contaminated with small amounts of tissue
thromboplastin. This can activate the extrinsic pathway and lead to variable results).- prompt and gentle inversion must be done to bind all calcium immediately.
D. Specimen Processing
- transportation to the laboratory should be done as quickly because some changes can begin in vitro.
- specimens should be centrifuged for 10 minutes to get the cell - free plasma - anticoagulant
mixture.
- always check for micro-clot formation, if present, specimen is unacceptable for testing.- hemolyzed specimens are also unacceptable for testing because of possible clotting factor
activation.
4. Routine Test of Hemostatic Function
A. Primary Hemostasis
A.1. Bleeding Time
- measures the time required for the cessation of bleeding after a standardized capillarypuncture to a capillary bed.
- the time required will depend on the capillary integrity, number of platelets and the
platelet function.
- types:a. Duke method: earlobe; RR: 0-6 minutes.
b. Ivy method: forearm; RR: 1-6 minutes.
c. Template method: forearm; RR: 2-9.5 minutes.
- prolonged with aspirin.
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A.2. Clot Retraction
- evaluates platelet function, fibrinogen, red cell volume and fibrinolytic activity.
- whole blood is allowed to clot and observed for retraction.
- platelets of adequate number and viability are required for clot retraction to occur.
- normal value: evidence of retraction within 2 hours (varies with methodology).
B. Secondary Hemostasis
B.1. Coagulation time/ Clotting Time
- measures the period required for the free flowing blood to clot or solidify after it has
been removed to the body.- types:
a. Drop or Slide method: capillary blood; NV: 2-4 minutes
b. Lee and White method (tube method): venous blood: NV: 7-15 minutes
B.2. Prothrombin Time (PT)
- test for extrinsic and common pathway.
- measures factors I, II, V, VII and X.- monitors oral anticoagulants (warfarin, coumarin, dicoumarol).
- reagent: tissue thromboplastin and CaCl2.- sensitive to vitamin K factors.- International normalized ratio (INR)
INR = (patient result)ISI
(mean of the reference range)
*ISI = International Sensitivity Index from manufacturer
- reference range: < 14 secondsa. Therapeutic goal: INR 2.0 3.5
B.3. Activated Partial Thromboplastin Time
- test for intrinsic and common pathway.
- measures all factors except factor VII and XII.
- monitors heparin therapy.
- reagents: activator (kaolin, celite or ellagic acid), platelet phospholipid (PF3) and CaCl2.
- reference range: 20 40 seconds
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OTHER TESTS:
Laboratory Tests for Primary
Hemostasis
Laboratory Tests for Secondary
Hemostasis
Laboratory Tests for
Fibrinolysis
1. Capillary Resistance /Fragility/ Tourniquet
/Rumpel Leedes or HessTest
2. Platelet Adhesiveness Test3. Platelet Aggregation Test
4. Platelet Count
5. Platelet Morphology andMPV
1. Plasma Recalcification Time2. Activated Clotting Time
3. Stypven Time4. Thrombin Time / Thrombin
Clotting Time
5. Reptilase Time6. Substitution Test (Mixing
Studies)
7. ProthrombinConsumption/Serum
Prothrombin Test
8. Thromboplastin GenerationTest
9. Specific Factor Assay
10. Assay of vWR:Ag and
vWR:Reo Rockett/Laurel11. Duckerts or Clot Solubility
Test
12. Tests for Circulating Inhibitorsof Coagulation
1. Determination ofFibrinolytic Products
2. Lysis Time3. Proteins involved in
Fibrinolysis
5. Bleeding Disorders
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Disorder of Primar Hemostasis
Vascular Disorde
Acquire
Anaphylact
purpura,Henoch
Schonlein,Senile purp
Scurvy, Pur
simplex,Infectious
purpura, Dr
induced
purpurasassociated w
paraprotene
s, Amyloid
Idiopathic
purpuras
Hereditary
Hereditary
Hemorrhagic
Telangiectasia/
Rendu-Weber-
Osler,Hemangioma-Thrombocytopenia
Kasabach-Merit,
Ehler-Danlos,Marfan,
Osteogenesis
imperfect,
Pseudoxanthoma
elasticum
Platelet Disorders
Qualitative Quantitative
Thrombocytosis
primary
reactive
Thrombocytopenia
Disorders relatedto distribution or
dilution /big
spleen
Increased platelet
destruction/ utilization
TTP, drug induced,
non immunemechanisms, DIC
Impaired/
decreased
platelet
production BM failure
Congenital hypoplasia, MHA,
WAS, BS, Fanconi, TAR
Neonatal hypoplasia
- drugs, infections
Acquired hypoplasia -ionizing radiation, drugs
Disorders of Platelet Secretion
Disorders
of Platelet
AdhesionDisorders of
PlateletAggregation
Glanzmanns
thrombasthenia
Acquired von
WillebrandDisease
Bernard
Soullier/
Giant
PlateletSyndrome
vonWillebrand
Disease
Thromboxane
Pathway Disorders
AcquiredHereditary
aspirin like
defects due to inhibitors of
prostaglandin pathway(chronic aspirin intake or
inhibitors of
thromboxane or cyclo-
oxegenase pathway
Storage Pool
Diseases
Primary granuledeficiency
Hemmeler
anomalyElectron dense/delta granules
deficiency
Pudlak,
WiskottAldrich,
Chediak
Higashi,
TAR Alpha granules deficiency
Gray Platelet Syndrome,
Quebec Platelet Disorder
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References:
McPherson, R. & Pincus, M. (2006). Henrys Clinical Diagnosis and Management by Laboratory Methods. US: SaundersElsevier
Linne, J. & Ringsrud, K. (1999). Clinical Laboratory Science The Basics and Routine Techniques
Harmening DM: Clinical Hematology and Fundamentals of Hemostasis, ed 3. Philadelphia, FA Davis Co, 1997.
Turgeon ML: Clinical Hematology: Theory and Procedures, ed 3. Philadelphia, Lippincott-Raven Publishers, 1998.
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