H. Testing of Primary Hemostasis (L6, L7, L9 & Lab 3) Flashcards
Broadly speaking what are the 2 main aspects of primary hemostasis that are assessed?
Platelet number
Platelet & VWF function
What is the quantitative assessment of platelets and when is it performed?
- Platelet count
* Routinely performed as part of a CBC
How commonly are platelet function tests ordered?
- Relatively infrequently due to high cost and the cumbersome nature of the tests
- Generally only do functional analysis if quantitative tests don’t explain symptoms
Platelet functional analysis can be performed on what 2 types of samples? Which is easier to use?
- Platelet rich plasma
- Whole blood
- Whole blood is easier to use because it does not have to be prepared
How is platelet aggregation tested on platelet rich blood?
- Optical density for the sample is measured –> relatively little light passes through as most is scattered by the platelets that are even dispersed through the sample –> high adsorption –> high OD
- Add platelet agonist –> continuing measuring OD –> platelets aggregate leaving more “open spaces” in the sample –> more light passes through –> plot OD on graph as it lowers overtime
What is the shape of the OD vs. time graph for the platelet aggregation test? How does it look for somebody deficient in agonist response?
- OD decreases over time after agonist is added
- The decrease is in 2 distinct phases. The first phase is due to the agonist which is added and the second phase is due to the agonist being released by platelets themselves
- If there is a deficient response to the agonist the curve begins to decrease and then “flat lines”
There are 2 groups of agonist used in platelet aggregation studies. Name the agonists in both groups and the mechanism they are testing:
- ADP, Epinepherine, Thrombin, Collagen and TxA2 –> test GPIIb/GPIIIa & fibrinogen
- Ristocetin –> GpIb & VWF
How is a whole blood aggregometer different than aggregation studies performed on platelet rich plasma?
• Instead of using spectrophotometry, it measures electrical impedance
What agonists are used in whole blood aggregometry?
- ADP, Collagen, Arachidonic acid and Ristocetin
* Epinepherine is not used
What disorders cause impaired clotting in response to ristocetin?
- Von Willebrand Deficiency
* Bernard Soulier Disease (GPIb deficiency)
What is Benard Soulier Disease?
• Impaired clotting due to a deficiency of GPIb (receptor for VWF) –> impaired coagulation
How is it determined whether impaired clotting to Ristocetin is the result of VWF deficiency or a deficiency of the platelet receptor for WVF (GPIb)?
• Ristocetin cofactor test
What is the Ristocetin Cofactor Test?
• Repeat Ristocentin test but use platelets from another healthy individual –> only the patients WVF is being used
What is suspected with an abnormal Ristocetin Induced Platelet Aggregation (RIPA) and normal Ristocetin Cofactor test?
- Bernard Soulier syndrome
* Normal cofactor test –> VWF from patient is fine –> must be problem with platelet –> def of GPIb
What is suspected with an abnormal Ristocetin Induced Platelet Aggregation (RIPA) and abnormal Ristocetin Cofactor test?
- VWF deficiency
* Abnormal cofactor test –> VWF from patient is not functioning
What 3 supplemental laboratory procedures can test VWF?
- Elisa tests the amount of VWF in the plasma
- Western Blot –> probe with anti-VWF –> VWF size distribution (Larger multimers are more active)
- Can test VWF ability to adhere to collagen to tests its ability to initiation adhesion
What is type 1 Von Willebrand Disease? What is it’s pattern of inheritance? Is it heterozygous or homozygous?
- Normal ratios of different sized VWF but absolute deficiency of all sizes
- Autosomal dominant
- Heterozygous
What is type 2 Von Willebrand Disease?
• Improper ratio of different sized VWF –> deficiency of larger more active VWF
What is type 3 Von Willebrand Disease? Is it heterozygous or homozygous?
- Production of VWF is virtually absent
* Homozygous for the mutation that causes type 1
What is the most common type of Von Willebrand Disease?
Type 1
What is necessary for a normal PFA-100 reading?
- Adhesion due to VWF
* Aggregation due to response to agonist
How is the PFA-100 (Platelet Function analyzer) performed?
- Have a membrane with a 150 micrometer whole –> suck blood through it and measure how long it takes to clot = closure time
- Collagen is present on the membrane (adhesion) and either epinephrine or ADP is given as an agonist (aggregation
What type of blood sample is the PFA-100 test performed on?
Whole blood
Platelet aggregation tests are not performed on patients with what bleeding disorder?
• Thrombocytopenia, already know that coagulation will be delayed by CBC & the aggregation tests are more expensive
Approximate size of a platelet?
2 micrometers
How is the PFA-100 closure time affected for patients taking aspirin?
- Delayed if epinephrine is used as the agonist
* Normal if ADP is used as the agonist
Why is PFA-100 the preferred test for platelet function?
• It is cheaper and easier to perform (can use whole blood)
If there is impaired platelet adhension there will be a deficient response to what agonist(s) during coagulation testing?
Ristocetin
If there is impaired platelet aggregation there will be a deficient to what agonist(s) during coagulation testing?
• ADP, epinephrine, thrombin, collagen and TxA2
What impairs platelet aggregation?
- Acquired = NSAIDs, Plavix, dialysis and cardiopulmonary bypass
- Inherited = afibrinogenemia, Glanzman thrombasthenia, gray platelet syndrome
In patients taking NSAIDs like aspirin, how is aggregation impaired?
• NSAIDs –> COX inhibitor –> prevent creation of TxA2 which is a platelet activator
In patients taking Plavix/Clopidrogel, how is aggregation impaired?
Plavix blocks ADP receptor
What is Glanzman Thrombasthenia?
Deficiency in GBIIb/IIIa
What is gray platelet syndrome?
• Storage disorder of granules in platelet prevents platelets own release of agonist
What substance is added to almost all blood samples to reversibly inhibit coagulation?
- Sodium citrate
* Removes Ca ions
What must be added to a blood sample before you can begin testing on it (after it has been stored in a refrigerator)?
- Calcium Chloride (to reverse effect of Sodium citrate)
- Phopholipid (to give factors a surface to work on. Can’t work in suspension)
- Warm the sample to 37 degrees so the enzymes can function
Factor VII circulates bound to what? Why?
- VWF
* Prolongs VII half life
What is an alternative to the OD vs. Time graph for platelet function testing?
- Aggregation vs time
* Graph is basically inverted
How soon after injury do symptoms present if there is a primary hemostasis disorder? How about for secondary?
Immediately
Delayed
Can injury to patients with primary hemostasis disorder be controlled with pressure? What about for patients with disorders of secondary hemostasis?
Yes
No
What are the general set of labs done when trying to diagnose a bleeding disorder?
Platelet count (primary) PFA (primary) PT (secondary) PTT (secondary) Thrombin Time (secondary)
What is the inherited disorder with platelet adhesion? What about aggregation?
Bernard Soulier
Glanzman Thrombastenia
If all aggregations studies are normal except when ristocetin is used, what is the likely diagnosis?
Bernard Soulier
If platelet aggregation studies are always abnormal except to ristocetin, what is the likely diagnosis?
Glanzman Thrombastenia
