1. Dr. Agtuca - Bt, Ct, Pt & Ptt

7/28/2019 1. Dr. Agtuca - Bt, Ct, Pt & Ptt

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Bleeding Time, Clotting Time,

Prothrombin Time/PTT


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Platelets (Thrombocytes)

Function interact with injured vascular wall

structures, plasma proteins, other circulating

blood cells

a. Adhesion: adhere to basement membrane of

injured blood vessel

b. Release action: release of alpha & dense

granules, & lysosomesc. Aggregation: formation of secondary, irreversible

platelet aggregation


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Coagulation:

Starts with activation ofThromboplastin, from

injured tissue

Fibrinolytic System: Endothelial cells

Heparin

Antithrombin


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Evaluation of Coagulation Factor Disorders

Medical History:

1. Symptoms

2. Onset of Bleeding

3. Location of Bleeding (?)

4. Family History

5. Drugs & Medications (?)

6. Other Systemic diseases

Physical Exam:


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Laboratory Screening Tests

1. Prothrombin Time (Protime/PT)

2. Partial Thromboplastin Time (PTT)

3. Platelet count

*If abnormal add:

1. Bleeding Time2. Clotting Time


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I. Prothrombin time /Protime/PT

Screen for deficiencies in: Extrinsic Pathway

FI, FII, V, VII, X

Test of choice for monitoring anticoagulant

therapy by Vit K antagonists (II, VII, IX, X)

Principle: Thromboplastin + Platelet-poorplasma & Ca++ react w/ FVII to activate FX ---

convert Prothrombin to Thrombin


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Specimen source:

Citrated platelet-poor plasma (PPP) Reference Range: Values differ

10 12 seconds or 12 14 seconds

Reporting:

1. Patient time in sec w/ Ref range

2. Patient time in sec w/ control time

3. Prothrombin ratio (PT divided by mean of Ref

range multiplied by 100)

4. Percent Activity (outdated?)


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5. International Normalized Ratio (INR):

- Widely used nowadays

- Comparison of Patient value with the normal value =

result is normalized to an international referencepreparation (IRP) of thromboplastin

- Values usually provided by manufacturers of

reagents


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Interpretation of results:

Sensitivity: detect deficiencies of


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II. Partial Thromboplastin Time (PTT)

Test of choice for factor deficiencies of the

Intrinsic & Common Pathway

Test of choice for monitoring Heparin therapy

PTT Reagent: 2 ocmponents

1. Platelet substitute (phospholipid) prepared from

brain & plant phospholipids

2.Activator- Kaolin, Celite, micronized silica, or

ellagic acid


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Specimen source:

Citrated platelet-poor plasma (PPP)

Reagents:

Phospholipid w/ Activators (PTT reagent)

0.025 M CaCl2

Procedure: PPP (0.1mL) + 0.1mL PTT reagent,incubate at 37C for 3-5min, + 0.1mL warm CaCl2.


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Reporting:

Reported in seconds, to the nearest tenth, along

with reference range.

Reference range: vary according to reagent,

method, instrument used

Lower limit of 20 sec to an upper limit of 45 sec


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Interpretation:

Prolonged PTT in the absence of Heparin use

indicate:

Factor deficiency

Acquired circulating anticoagulant (e.g. Lupus inhibitor)

Antibody to a specific Factor (e.g. anti-FVIII)

Prolonged PTT if factor measured is


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Sources of Error:

1. Sample collection & preparation

a. Improper collection & specimen processing

b. Incorrect anticoagulant-to-plasma ratio

c. Hemolysis

d. Platelets in plasma sample

e. Unexpected heparin contamination


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2. Reagent preparation

a. Improper storage

b. Water impurities

c. Incorrect dilution

3. Instrumentation

a. Failing light source

b. Fluctuations in temperaturec. Loss of calibration of tubing

d. Contamination


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III. Platelet Count

Formed in the BM from Megakaryocytes

Difficult to count:

a. Small & difficult to discern

b. Attached to surfaces/particles in diluting fluid

c. Disintegrate easily

d. Form clumps with other platelets


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Specimen:

Capillary blood from finger puncture

Values are lower because of clumping at puncture site

Venous blood

EDTA (Lavender tube) as anticoagulant of choice


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Method of Determination:

1. Manual Count

a. Unopette system

b. Hemacytometer

2. Automated Method

a. Optical methods

b. Impedance methods


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Calculation: Manual Platelet Count

Ave # of Plt in 4 squares X 1mm2 x 100 X 106 = Platelets X 109/L

0.1 mm

Platelet Reference Range: 150 450 x 109/L


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Clinical Significance:

Thrombocytopenia:

Platelet values lower than normal

Thrombocytosis:

Platelet values higher than normal


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TESTS FOR PLATELET FUNCTION

1. Platelet Closure time

2. Platelet Aggregation Studies

3. Automated Platelet Function Analysis


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Platelet closure time:

Assess platelet-related primary hemostasis; greateraccuracy & reliability than Bleeding Time (BT).

Instrument and test cartridge system which simulates theprocess of platelet adhesion and aggregation following avascular injury.

Rapid evaluation of platelet function on samples ofanticoagulated whole blood.

The time required to obtain full occlusion of the apertureis reported as the closure time (CT) in seconds.

Sensitive to platelet adherence and aggregation

abnormalities ; allows discrimination of aspirin-like defectsand intrinsic platelet disorder.


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Platelet Aggregation Studies:

Measure response of platelet during hemostatic

process

Change in shape

Increase in surface adhesiveness Platelet plug formation

Use of Aggregating Agent

Measurement: Turbidimetry Method


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CLOTTING TIME

Time required for a blood sample to coagulate/clot in

vitro under standard conditions.

Most common method: Capillary Tube Method.

Affected by calcium ion levels and many diseases.

Normal value: 5 to 8 minutes.

Time taken for blood to clot reflects time required for

the generation of thrombin


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BLEEDING TIME

Principle: BT is the time it takes for a standard wound

to stop bleeding

Comprehensive test of platelet action in vivo & is

sensitive to the following abnormalities:1. Platelet numbers & function

2. Plasma VIII:vWF deficiencies

3. Vessel wall composition Standardization of Wound size is a problem

Employs BP cuff inflated at 40 mmHg


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Reference range:

2 to 9 minutes

Considerations for the Test:

Volar skin must be dry Adequate incision/proper depth

Site should not touch filter paper

Aspirin& aspirin-containing meds must be discontinued 1

week prior to testing

NSAIDS must be discontinued 24 hrs prior to testing


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1. Dr. Agtuca - Bt, Ct, Pt & Ptt

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