HEM Coag Testing Flashcards
PT (Prothrombin Time): Purpose
identifies deficiency in EXTRINSIC + COMMON pathway (I/ fibrinogen, II/ prothrombin, V, VII, X)
The PT reagent contains (3)
thromboplastin, phospholipid, and calcium
PTT (Partial Thromboplastin Time): Purpose
- identifies deficiency in INTRINSIC + COMMON pathway (all factors except VII and XIII)
- monitors unfractioned heparin treatment + some thrombin inhibitors
Mixing Studies: Purpose
Determines if abnormal PT/PTT is due to factor deficiency OR an inhibitor
Mixing Studies: Principle
- mix patient plasma + normal control (has all coag factors)
- test PT or PTT
Corrected = factor deficiency
Uncorrected = perform inhibitor screen
Inhibitor Screen: Purpose
Determines if inhibitor is immediate OR delayed
Inhibitor Screen: Principle
- (In 1:1 AND 4:1 patient ratios) = mix patient plasma + Normal control (has all coag factors)
- test PT/ PTT 3 times:
1. Mixed: Immediately
2. Mixed: after 1 HOUR of incubation at 37 degrees
3. Separately: after 1 HOUR of incubation at 37 degrees
What does it mean if PT/PTT of an inhibitor screen is prolonged immediately after patient plasma + Normal control are mixed ?
- Inhibitor screen = POS
- Inhibitor reacts IMMEDIATELY
What does it mean if PT/PTT of an inhibitor screen is prolonged after 1 HOUR of (patient plasma + Normal control) being mixed when it was normal after being immediately mixed ?
- Inhibitor screen = POS
- Inhibitor is delayed reacting
How can heparin be removed before performing an inhibitor screen ? Why is this done ?
- remove Heparin using Hepzyme
- Heparin causes FALSE POS inhibitor screen
- prolongs PT/ PTT
Why is patient plasma and Normal control incubated SEPARATELY for 1 hour at 37 degrees as part of the Inhibitor Screen ? What should the results be ?
- separate incubation ensures clotting factors did not degrade during incubation
- Normal control = POS
- patient plasma = neg
Why does the inhibtor screen use 1:1 AND 4:1 patient to Normal control ratios ?
4:1 ratio increases sensitivity to inhibitors
Single Factor Assay: Purpose
Determines which factor(s) patient is deficient in
Single Factor Assay: Principle
- mix patient plasma + plasma deficient in a single factor
- test PT or PTT
- compare results to standard graph to determine factor concentration
If patient plasma corrects PT/ PTT in a Single Factor Assay, what does this mean ?
Recall: patient plasma + plasma deficient in a known factor are mixed
Corrected = patient corrects deficient plasma; patient is not deficient in particular factor
Uncorrected = patient unable to correct deficient plasma; patient is deficient in the same known factor
Fibrinogen Testing: Purpose
Determines levels of fibrinogen in plasma
Fibrinogen Testing: Principle
- EXCESS thrombin + patient sample
- level of plasma fibrinogen is INVERSELY proportional to clotting time
NOTE: excess thrombin neutralizes anti-thrombin activity (FDPs)
Why is thrombin added in EXCESS in fibrinogen testing ?
To neutralize anti-thrombin activity (ie. FDPs)
TT (Thrombin Time): Purpose
- Detect DECREASED levels of fibrinogen, DYSFIBRINOGENEMIA, or THROMBIN INHIBITORS (ie. FDPs, D-dimers)
- Monitor/ evaluate DIC, heparin therapy, fibrinogen diseases
TT: Principle
Thrombin Time:
- dilute thrombin + patient sample to bypass previous steps of coag cascade
- measures time to convert fibrinogen to insoluble fibrin crosslinks
D-Dimer: Purpose
To rule out DIC
D-Dimer: Principle
- turbidimetric IMMUNOASSAY (DIRECTLY proportional)
- MONOCLONAL Abs specific to D-dimers coat latex particles
- D-dimers DIRECTLY bind to Abs = AGGLUTINATION = increased turbidity = INCREASED ABSORBANCE measured
Reptilase Time: Purpose
To detect HEPARIN, DYSFIBRINOGENEMIA, or FIBRINOGEN DEFICIENCY
Reptilase Time: Principle
reptilase (enzyme) = converts fibrinogen to fibrin + NOT INHIBITED by heparin = clot forms
- compares results to PROLONGED thrombin time (TT)
What does it mean if TT = prolonged and Reptilase Time = equally prolonged ?
Fibrinogen Deficiency
- there is no fibrinogen in the first place; reptilase cannot form fibrin
NOTE = prolonged TT cannot be due to heparin because reptilase is not affected by heparin, which is also prolonged in this case
What does it mean if TT = prolonged and Reptilase Time = MORE PROLONGED ?
DYSFIBRINOGENEMIA
What does it mean if TT = prolonged and Reptilase Time = NORMAL ?
HEPARIN is present
- but reptilase is unaffected by the anticoagulant
Which coagulation tests use Turbidimetric end-point clot detection ?
- PT
- PTT
- TT
- Fibrinogen
Platelet Aggregometry: Purpose
To assess PLT function (secretion, aggregation, adhesion w/ ristocetin)
Platelet Aggregometry: Principle
- DO NOT DO if PLT <50 x10^9/L
- PLT-RICH plasma is stirred in AGGREGOMETER with diff agonists:
1. ADP + collagen + EPO + arachidonic acid = tests GPIIb/ IIIa and fibrinogen function (aggregation)
2. Ristocetin = tests GPIb and vWF function (adhesion)
NOTE: agonists initiate PLT aggregometry
Interferences in PLT Aggregometry
- gross lipemia
- aspirin
- PLT <50 x10^9/L
Ristocetin Co-factor Assay: Purpose
- To ensure vWF is present for coagulation
- Differentiate VonWillebrand Disease from Bernard Soulier
Ristocetin Co-factor Assay: Principle
- mix patient plasma with Normal PLTs + Ristocetin
- Ristocetin induces vWF to attach to GPIb receptors on Normal PLTs = aggregation
How to differentiate Unfractioned Heparin (UFH) from Low Molecular Weight Heparin (LMWH) ?
Both:
- indirect anticoagulants
UFH:
- inhibits all serine proteases (plasmin, IIa, IXa, Xa, XIa, XIIa) EXCEPT VII and Kallikrein
- monitored by aPTT or anti-Xa assay
LMWH:
- inhibits Xa
- monitored by anti-Xa assay ONLY
Heparin is a cofactor of __.
Heparin is a cofactor of ANTI-THROMBIN.
Tube/ anticoagulant requirements for coagulation testing ?
- Sodium citrate (light blue top)
- Filled to capacity
- 1:9 anticoagulant to blood