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Introduction to Haematology > Lab Investigation of Coagulation > Flashcards

Lab Investigation of Coagulation Flashcards

1
Q

primary haemostatic testing test

A

platelet function assays (PFA)

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2
Q

secondary haemostatic testing test

A

prothrombin time (PT)
activated partial thromboplastin time (APTT)
fibrinogen - derived and claus
thrombin time (TT)
D-dimers

1st 3 are not FXIII

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3
Q

investigations of prolonged clotting times?

A

mixing studies
factor assays
inhibitor assays
vWF antigen and activity

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4
Q

when should sodium citrate samples be collected?

A

second order of draw

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5
Q

how does sodium citrate work?

A

removes calcium ions preventing coagulation without compromising any clotting proteins

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6
Q

what type of anticoagulant is sodium citrate?

A

a reversible anticoagulant

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7
Q

addition of what will initiate the clotting process?

A

calcium

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8
Q

what speed and time are citrate samples centrifuged for and why?

A

3500 RPM for 5 mins
separate plasma from red cells
provide platelet poor plasms

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9
Q

what happens when a poor venepuncture technique is used?

A

additional tissue factor in the sample (TF) = activated sample = short clotting time (artificial)

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10
Q

what are the 2 main analytical principles for measuring clotting times

analysers may have either or both

A

light absorbance / transmission (optical)
mechanical / viscosity

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11
Q

what else can coagulation analysers measure?

A

light absorbance / transmission using filters of multiple wavelengths

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12
Q

in what situation may you see no coagulation curve?

A

disseminated intravascular coagulation - coagulation consumption

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13
Q

what do platelet functions do

A

assesses process of platelet adhesion and aggregation following stimulation of vascular injury in vitro

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14
Q

what are platelet function assays used for

A

investigation of pathological bleeding

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15
Q

sample requirements for platelet function assay

A

delivered by hand
test samples between 30mins - 4hrs of collection
mix by gentle inversion by hand, not roller
800ul whole blood, sodium citrate
ADP.EPI- antagonists

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16
Q

what is the platelet count/morphology and PF results for type 1, 2a and 3 vWD disorders

A

normal count and morphology
PF results - CADP/CEPI both equally prolonged. very prolonged in 2a and 3.

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17
Q

what is the platelet count/morphology and PF results for platelet type 2B or vWD

A

normal count and morphology
PF results both abnormal

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18
Q

what is the platelet count/morphology and PF results for glanzmann thromboasthenia?

A

platelet count and morphology normal
PF results - CADP/CEPI both very prolonged

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19
Q

what is the platelet count/morphology and PF results for Bernard-Soulier Syndrome

A

platelet count/morphology - mild/moderate. macrocytopenia.
CADP/CEPI both very prolonged

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20
Q

what does prothrombin time evaluate?

A

the extrinsic and common pathways

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21
Q

what is prothrombin time testing sensitive to deficiencies in

A

FII, FV, FVII and FX

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22
Q

what is PT (prothrombin time) affected by?

A

hereditary disorders
liver disease
vit k deficiency
anticoagulant therapy e.g. warfarin therapy

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23
Q

what is PT tested

A

added calcium and thromboplastin to the blood sample then measuring the time (in seconds) required for fibrin clot formation
uses % detection method to calculate the formation of a clot in vitro
does not assess fibrinogen or FDP levels

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24
Q

how is the international normalised ratio (INR) derived?

A

from PT testing

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25
Q

what is the calculation for INR

A

INR = patient PT / normal PT x ISI

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26
Q

What is ISI?

A

the international standardised ratio
it is specific to the batch of thromboplastin reaction used

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27
Q

what is INR generally used for?

A

to monitor anticoagulant therapy e.g. warfarin

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28
Q

what INR result needs urgent assessment due to bleeding risK?

A

INR > 5.0

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29
Q

what to remember with warfarin

A

warfarin patient general age = ? accurate compliance

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30
Q

what type of testing do new/direct anticoagulants require for evaluation?

A

FXa testing

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31
Q

what is APTT?

A

activated partial thromboplastin time

32
Q

what pathways does PTT evaluate?

A

the intrinsic and common pathways

33
Q

APTT cannot detect deficiency of?

A

FVII or FXIII

34
Q

what is APTT commonly affected by

A

heparin therapy

35
Q

what could prolonged APTT clotting times be due to?

A

anticoagulant therapies e.g. heparin
factor deficiency
lupus anticoagulant

36
Q

what is APTT particularly sensitive to deficiencies in?

A

FVIII, FIX and FXI

37
Q

if both the PT and APTT are prolonged where is the defect likely to be in?

A

the common clotting pathway i.e. deficiency of factor I, II, V or X is suggested

38
Q

where defect likely to be in normal PT with abnormal APTT?

A

intrinsic pathway
deficiency of factor VIII, IX, X, or XIII is suggested

39
Q

what does a normal PT with an abnormal APTT mean?

A

that the defect lies within the extrinsic pathway and suggests a possible factor VII deficiency

40
Q

when do PT and APTT detect a factor deficiency?

A

when the factor decreases to 25-40% of normal

41
Q

what is the clauss fibrinogen test

A

thrombin is added to various known concentrations of fibrinogen
the test requires a reference plasma with a known level of fibrinogen calibrated against a known internal standard
a calibration curve is constructed using this reference plasma by preparing a series of dilutions (!:5-1:40) in buffer to give a range of fibrinogen concentrations
the clotting time of each of these dilutions is established and the clotting time(s)/fibrinogen concentration (g/L) are plotted on log-log graph paper.
the 1:10 concentration is considered to be 100% i.e. normal
there should be a linear correlation between clotting times in the region of 10-50s
the test diluted plasma (diluted 1:10 in buffer) is incubated @37degrees C, phospholipid and thrombin are added followed by calcium (all pre-warmed to 37 degrees C)
on the addition of calcium timing begins
the time taken for the clot to form is compared to a calibration curve and the fibrinogen concentration is determined
automated method in which clot formation is determined to have occurred when the optical density of the mixture has exceed a certain threshold

42
Q

what does thrombin time (TT) measure?

A

the conversion of fibrinogen to fibrin

43
Q

what is TT dependent on

A

clotting time dependent on quantity and quality of fibrinogen

44
Q

what is TT used for

A

monitoring of fibrinolytic therapy
evaluation of fibrin formation

45
Q

how is TT performed

A

addition of an excess of thrombin to undiluted plasma

46
Q

when is TT increased

A

in heparin therapy

47
Q

what is the normal range for TT time, the time taken for a fibrin clot to form

A

normal range 15-22 seconds

48
Q

coag test results for liver disease

A

low platelet count
prolonged PT
Prolonged APTT
TT normal (rarely prolonged)

49
Q

coag test results for DIC

A

plt count low
PT prolonged
APTT prolonged
TT grossly prolonged

50
Q

coag results from massive transfusion

A

plt count low
prolonged PT
Prolonged APTT
Normal TT

51
Q

coag results coumarin anticoagulants

A

plt count normal
PT grossly prolonged
APTT prolonged
TT normal

52
Q

coag results for heparin

A

Plt normal (rarely low)
PT midly prolonged
APTT prolonged
TT prolonged

53
Q

coag results for circulating anticoagulatant

A

plt count normal
PT normal / prolonged
APTT prolonged
TT normal

54
Q

what do DDIMERS measure

A

amount of FDPs after clot lysis

55
Q

what is ddimers an indicator of?

A

coagulationa ctivity

56
Q

where are raised DDIM results seen

A

DIC
thromboembolic diseases
myocardial infarction
3rd trimester of pregnancy

57
Q

what is the major diagnostic application of ddimers?

A

exclusion of thromboembolic events e.g. DVT

58
Q

how is bleeding time measured

A

standardised incision on pt forearm- blade
a pressure cuff put on pt @ 40mmHg to standardise pressure of the blood flow
blood is mopped up taking care not to disturb the platelet plug. the time for the bleeding to stop is noted.

59
Q

disadvantages of bleeding time

A

operator dependent, poorly reproducible, neither objective or sensitive

60
Q

mixing studies

A

50/50 mix of normal / patient plasma - retest APTT
80/20 mix of normal / patient plasma
correction = factor deficiency
no correction ? lupus anticoagulant

61
Q

what are factor assays used for?

A

diagnose congenital or acquired factor deficiency states
distinguishing between dysproteinaemias and protein synthesis disorders
monitoring factor replacement therapy

62
Q

what does an extrinsic factor assay look for?

A

FII, FVII and FX

63
Q

What factors are tested for using an intrinsic assay

A

FVIII, FIX, FXI and FXII

64
Q

what is used to confirm a factor deficiency in patient plasma

A

factor deficient used to confirm a factor deficiency in patient plasma and PT/APTT reassessed

65
Q

how is a factor assay result interpreted?

A

using reference curve of standard human plasma or pooled normal plasma (PNP). patient deficient in that particular factor cannot compensate for the absence in factor deficient plasma

66
Q

what are inhibitors linked to coagulation?

A

inhibitors are antibodies that in coagulation are usually targeted against either
- specific clotting factors e.g. Factor VIII
- phospholipids e.g. Lupus

67
Q

an example of a time-dependent circulating inhibitor

A

FVIII inhibitors are lost quicker than FIX inhibitors

68
Q

where else can inhibitors develop?

A

inhibitors can develop for those receiving recombinant factor therapy e.g. haemophilia patients

69
Q

what is the assay used to test for

A

Bethesda Assay

70
Q

what is a bethesda unit (Bu)?

A

it is defined as the amount of an inhibitor that will neutralise 50% of 1 unit of FVIII:C in normal plasma after 120 mins incubation @ 37 degrees C

71
Q

vwf antigen…

A

determines the levek of von willebrand factor vwf in your blood by measuring the vwf protein (antigen) using immunoloical assaysh

72
Q

how is vwf antigen measured

A

typically by ELISA or automated latex immunoassays

73
Q

how do latex immunoassays work for vwf

A

uses latex micro-particles coated with an antibody to human vwf. in the presence of vwf the latex particles agglutinate and in proportion to the concentration of vwf:Ag in the plasma sample (turbidity)

74
Q

how do we measure vwf activity?

A

risocetin cofactor [vWF:RCo] assay measures the ability of a patient’s plasma to agglutinate platelets in the presence of the antibiotic Ristocetin. the rate of ristocetin induced agglutination is related to the concentration and functional activity of the plasma vwf

75
Q

how are factor assay results interpreted?

A

using reference curve of standard human plasma or pooled normal plasma (PNP)

76
Q

what happens with factor deficient plasma

A

plasma deficient in that particular factor cannot compensate for absence in factor deficient plasma

77
Q
A

Introduction to Haematology (10 decks)

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