Lab Testing in Coagulation

Question 1

Coagulation testing

Answer 1

• tests performed at 37C
• PLASMA is specimen of choice
• Sodium citrate is anticoagulant (9:1)
• most tests require addition of Calcium

Question 2

Platelet Poor Plasma (PPP)

Answer 2

• plasma containing < 10 x 10^9/L platelets
• prepared by centrifuging plasma for 15 mins at high speed
• need because platelets can neutralize heparin and vWF

Question 3

Platelet Rich Plasma (PRP)

Answer 3

• plasma containing 200-300 x 10^9/L platelets

- prepared by centrifuging for 10 mins at low speed

Question 4

Specimens for Coagulation testing

Answer 4

• sodium citrate (3.2%) binds Ca to prevent clotting
• requires plasma (contains coagulation factors)
• blue tops should be collected first
• proper ratio (9:1) is important - no short draw tubes

Question 5

Platelet tests

Answer 5

Peripheral smear
Platelet count
Platelet aggregation
Bleeding time and platelet function analyzer
Platelet secretion studies
Flow cytometry

Question 6

Platelet counts and estimates (specimen problems)

Answer 6

• Clumping due to antibodies
• Satellitism as a artifact of EDTA
• Giant platelets not counted as platelets
• Fragment RBC’s counted as platelets

Question 7

Platelet aggregation studies

Answer 7

• determines whether plts will clump with other plts when stimulated by agonists
• activating agents: ADP, collagen, ristocetin, epinephrine, arachidonic acid
• addition of activators results in shape change
• causes an increase in % light transmission
• aggregation occurs in two waves
• Specimen = platelet rich plasma
• aggregation inhibited by aspirin

Question 8

Primary wave (platelet aggregation)

Answer 8

• direct response of platelets to aggregating reagent

- platelet shape change and formation of small aggregates

Question 9

Secondary wave (platelet aggregation)

Answer 9

• represents complete aggregation

- result of endogenous ADP being released from activated platelet dense bodies

Question 10

Bleeding Time (BT)

Answer 10

• measure of platelet function
• 3 Types: Ivy, Duke and Template
• Prolonged by aspirin
• NORMAL: 1 - 9 minutes

Question 11

Duke Bleeding Time

Answer 11

one incision, usually earlobe

Question 12

Ivy Bleeding Time

Answer 12

• standardized back pressure (blood pressure cuff)
• below inner aspect of elbow
• 1-3 incisions
• depth not standardized

Question 13

Template Bleeding Time

Answer 13

• standardized back pressure
• standardized depth
• below inner aspect of elbow
• blot filter paper every 30 seconds until bleeding stops

Question 14

BT Controversies

Answer 14

• prolonged if low plt count, decreased fibrinogen, vascular disease
• not very sensitive
• platelet function can be impacted by drugs, food, etc.
• SHOULD NOT BE PERFORMED IF PLT < 100,000

Question 15

BT Compromises

Answer 15

• poor circulation at puncture site
• disturbance of platelet plug
• excessive movement by patients (kids)
• scaring, bruising

Question 16

Platelet Function Analyzer (PFA)

Answer 16

• replaced BT test
• specimen at room temp
• blood pumped through 2 apertures (collagen/epi (CEPI) and collagen/ADP)
• measures time needed to occlude aperture
• time need is function of plt count, plt activity, vWF activity and hematocrit

Question 17

abnormal CEPI, normal CADP

Answer 17

probably due to aspirin

Question 18

abnormal CEPI, abnormal CADP

Answer 18

probably a platelet function abnormality

Question 19

Platelet secretion studies

Answer 19

• not done in most labs

- secretion of granules monitored by chemiluminescence

Question 20

Flow Cytometry

Answer 20

• antibodies reactive with platelet surface available

- useful for GP1b/IX and GP1b/IIIa deficiency

Question 21

Prothrombin time (PT)

Answer 21

• tests extrinsic pathway
• patient plasma + tissue thromboplastin (with Ca)
• screen for inherited or acquired deficiencies in extrinsic and common pathways
• measures factors I, II, V, VII, X
• monitors oral anticoagulants (coumadin, warfarin)
• INR used to standardize results

Question 22

INR (International normalized ratio)

Answer 22

• used to correct for differences in coagulation instruments, reagents, etc.
• calculation is reported with all PT results
• 0 < INR < 6

Question 23

PT Sources of error

Answer 23

• traumatic venipuncture can introduce tissue factor and premature activation of factor VII (falsely shorten PT)
• delay between collection and testing decreases factor V
• increased Hct = decreased plasma, improper ratio of anticoagulant

Question 24

Activated Partial Thromboplastin time (APTT)

Answer 24

• add plt PL substitute and activator (APTT reagent) and Ca to activate factor XII
• tests intrinsic pathway
• measures all factors except VII and XIII
• monitors IV anticoagulant therapy (heparin)
• same sources of error as PT