This document provides guidance on evaluating a patient presenting with a potential bleeding disorder. It describes the terminology used to describe different types of bleeding and bruising. It emphasizes taking a thorough bleeding history including details of past bleeding events, family history, medication use and menstrual/pregnancy history. The physical exam focuses on signs of bruising, cutaneous abnormalities and other systemic findings. Laboratory tests evaluate primary hemostasis (platelets), secondary hemostasis (clotting factors) and fibrinolysis. Specific bleeding patterns may suggest deficiencies in certain coagulation factors.
3. TERMINOLOGY
• PETECHIAE
• Small, flat, red, discrete areas of skin bleeding ,<2-3 mm in diameter
• Nonblanching and nonpalpable
• Occur in dependent areas of the body
• This finding is typical in severe thrombocytopenia as well as in areas of skin
fragility or with glucocorticoid use
• PURPURA
• Results from coalesced petechiae, >3mm in diameter
• Wet purpura
• Hemorrhagic blisters in areas of mucous membranes
• Most predictive of serious bleeding in individuals with thrombocytopenia
• Dry purpura
• Purpura in the skin
• Purpura due to thrombocytopenia is flat and non blanching and is usually
seen in dependent areas of the body
• Purpura due to vasculitis is usually palpable and may be pruritic, and the
distribution does not follow dependent areas
5. TERMINOLOGY
• Bruise/ecchymosis
• Caused by subcutaneous accumulation of extravasated blood
• The skin is flat
• The color evolves over time from purplish blue to reddish
brown to greenish yellow, reflecting metabolism (breakdown)
of hemoglobin to biliverdin and bilirubin
• Significant when five or more are seen
• Bruises of the same age are consistent with a single traumatic
event, whereas bruises of different ages are consistent with
an ongoing process
• Hematoma
• Collection of blood in the extravascular space
• A subcutaneous hematoma may raise the skin profile
• Hematomas in deep tissues (eg, muscle, retroperitoneal) may
be suspected due to pain, a drop in hemoglobin level, or a
fluid collection on imaging studies
• Hematomas and hemarthroses (joint bleeding) are typical of
coagulation factor deficiencies
7. TERMINOLOGY
• Primary hemostasis
• Refers to the initial steps in clot formation, which mostly rely on the
integrity and elasticity of the vessel wall and interactions between
the vessel wall, platelets, and von Willebrand factor (VWF)
8. TERMINOLOGY
• SECONDARY HEMOSTASIS
• Vascular injury exposes tissue factor at the site of injury.
• Tissue factor binds and activates factor VII , setting in motion a cascade
of reactions that culiminates in thrombin generation.
• Thrombin cleaves circulating fibrinogen into insoluble fibrin, creating a
fibrin meshwork,and also is a potent activator of platelets, leading to
additional platelet aggregation at the site of injury
• THROMBUS AND ANTITHROMBOTIC EVENTS
• Clot stabilization and resorption
• Polymerized fibrin and platelet aggregates undergo contraction to form
a solid, permanent plug that prevents further hemorrhage.
• At this stage, counterregulatory mechanisms (eg.t-PA) are set into
motion that limit clotting to the site of injury and eventually lead to clot
resorption and tissue repair
11. TERMINOLOGY
• Major bleeding
• Defined as bleeding that results in death, affects a
critical organ such as the brain or a large joint,
causes a fall in hemoglobin level of 2 g/dL or more,
or requires transfusion of 2 or more units of red blood
cells
• Clinically relevant non-major bleeding (CRNMB)
includes bleeding that is more than expected for the
circumstances and that requires some intervention
(hospitalization, medication, face-to-face discussion)
but is not severe enough to constitute major bleeding
• Bleeding assessment tool (BAT)
• A BAT, also called a bleeding score, is used to
quantify bleeding by site and severity as a means for
predicting the likelihood of a bleeding disorder and/or
to codify bleeding in clinical studies
12.
13. PATIENT HISTORY
• Underlying medical conditions
• Several medical conditions predispose to bleeding
• Cancer
• Excess alcohol use
• Liver disease
• Kidney disease
• Connective tissue disorders
• Hypothyroidism
• Over-the-counter medications, is essential to identify agents
that impair hemostasis (nonsteroidal antiinflammatory
agents, anticoagulants, long-term glucocorticoids)
14. Elements of the bleeding history
• Detailed personal bleeding history and family
history
• Prior bleeding events over time:
• Bleeding during infancy (eg, umbilical stump
bleeding) and childhood(eg, bleeding with loss of
deciduous teeth)
• Bleeding during adolescence and adulthood
(additional questions related to menstruation and
pregnancy)
• Any bleeds severe enough to require surgical
intervention, nasal packing or cautery, a visit to the
emergency department, or transfusion
• History of iron deficiency or iron-responsive anemia
15. Elements of the bleeding history
• Outcome of bleeding challenges:
• Trauma, including severity of bleeding, need for surgical
intervention,and need for transfusions
• Surgical procedures, including timing and location,
comments of the surgeon regarding the character of the
bleeding (eg, diffuse oozing versus bleeding from a
specific source), and need for transfusions
• Dental procedures (eg, extractions), including timing,
duration, and need for packing or additional interventions
• If a woman has bled with some procedures but not others,
she should be asked if she was taking oral contraceptives
or hormone therapy during procedures in which she had
good hemostasis, since these hormones can increase
levels of von Willebrand factor (VWF) and normalize
hemostasis
• History of poor wound healing
16. Elements of the bleeding history
• Menstrual history
• Especially the presence of nighttime "flooding,“ passage of
clots larger than approximately 2.5 cm (size of a quarter
[United States coin]), duration of menses longer than eight
days, development of iron deficiency, and/or heavy
menstrual bleeding (previously called menorrhagia) leading
to hysterectomy at a young age
• Measures such as number of pads or tampons used can be
imprecise because they depend on the fastidiousness of the
individual, and the volume of blood lost is nearly impossible
to determine if pads or tampons are used
• Scoring systems have been devised that use pictures of
pads or tampons with varying degrees of saturation; a high
score correlated well with the presence of heavy menstrual
bleeding in some studies
• Endometriosis and hemorrhagic ovarian cysts are seen with
increased frequency in women with von Willebrand disease
17. Elements of the bleeding history
• Pregnancy history
• Including bleeding with delivery or postpartum and early
pregnancy loss (seen with some rare coagulation factor
disorders)
• In VWD, postpartum bleeding (24 to 28 hours after delivery)
is more common than bleeding during pregnancy because
higher estrogen levels in the second and third trimesters
promote increased VWF production
• Pro-hemostatic changes during pregnancy may also reduce
bleeding due to other disorders
• Antibiotic use or malabsorption syndromes that could
cause vitamin K deficiency
18.
19.
20. Normal range is <4 in adult males, <6 in adult females and <3 in children
bleedingscore.certe.nl
21.
22. Interpretation of the bleeding history
• A history of bleeding severe enough to require surgical
interventions, transfusions, or iron replacement
supports the suspicion of an underlying bleeding
disorder
• Conversely, a history of surgical procedures, dental
extractions, and/or significant trauma that did not
require transfusions or other nonstandard
interventions is evidence against the presence of an
inherited bleeding disorder
• In contrast, a history of easy bruising, which is
commonly reported, lacks sensitivity and specificity
and may be a sign of trauma or skin fragility rather
than a true bleeding disorder
• Symptoms such as hematuria, melena, and heavy
menstrual bleeding are also less helpful since these
are more likely to be due to structural/anatomic lesions
rather than to a bleeding diathesis
23. Interpretation of the bleeding history
• The sites and character of bleeding may help distinguish disorders of
primary hemostasis (platelet function) from secondary hemostasis
(coagulation factor function)
• Excessive menstrual bleeding has many causes (eg, an anatomic
lesion, hormonal factors, or impaired hemostasis)
• A bleeding disorder is found in as many as 10 to 30 percent of women
with excessive menstrual bleeding
• Platelet and vascular disorders are generally characterized by mucosal
bleeding (eg, mouth, nose, urinary tract, gastrointestinal tract) and
immediate bleeding upon a bleeding challenge
• Epistaxis may be a presenting finding in VWD or hereditary
hemorrhagic telangiectasia (HHT)
• New-onset bruising may herald a new thrombocytopenic disorder (eg,
immune thrombocytopenia [ITP], acute leukemia) or an acquired
coagulation disorder such as acquired hemophilia or disseminated
intravascular coagulation (DIC)
24. Interpretation of the bleeding history
• Coagulation factor disorders are generally characterized by
bleeding into muscles and joints and delayed bleeding after a
bleeding challenge
• Bleeding with circumcision is characteristic of severe hemophilia
A (factor VIII deficiency) or hemophilia B (factor IX deficiency)
• Exceptions are factor XI deficiency, which tends to have more
mucosal bleeding, and factor XIII deficiency, which tends to show
delayed bleeding after trauma or surgery
• Umbilical stump bleeding is characteristic of factor XIII deficiency
• Disorders of fibrinolysis tend to show delayed bleeding after
trauma or surgery
• Inherited disorders tend to present in childhood if severe but
frequently present in adulthood or only after a bleeding challenge
if mild.Some inherited disorders have additional syndromic
features
• Some acquired disorders are associated with an underlying
condition such as pregnancy, cancer, or connective tissue
diseases
25. • Family history
• A family history of bleeding should be obtained to
help determine the likelihood of an inherited
condition
• Some individuals may be aware of a specific
diagnosis in a first- or second-degree relative; others
may report a family history of severe bleeding
requiring medical interventions (surgery,
transfusions)
• While a positive family history increases the
likelihood of an inherited bleeding disorder, lack of a
family history does not eliminate the possibility
• Medication use
• Bleeding risk is increased with the use of certain
medications, herbal preparations, and dietary
supplements that interfere with primary or secondary
hemostasis
26. Drug class or
substance
Mechanism
Anticoagulants Interfere with clot formation (secondary hemostasis)
Antiplatelet agents,
including NSAIDs
Interfere with platelet function (primary hemostasis)
Glucocorticoids Interfere with vascular integrity
Antibiotics Cause vitamin K deficiency, especially with longer use
Some interfere with platelet function
SSRIs Interfere with platelet function (primary hemostasis)
Alcohol complications of liver disease may affect clot formation
and may cause thrombocytopenia
Due to direct marrow toxicity
Vitamin E Interferes with vitamin K metabolism in some
individuals
Garlic Interferes with platelet function in some individuals
Gingko biloba Unknown
27. TARGETED PHYSICAL EXAMINATION
• Cutaneous findings
• Petechiae in dependent areas (legs, sacrum),may indicate
Thrombocytopenia
• Petechiae, purpura, ecchymosis, and scarring on the arms may indicate
solar purpura, which is a common finding related to aging and is
generally not considered a bleeding disorder
• Telangiectasias around the lips or on the fingertips may indicate
hereditary hemorrhagic telangiectasia (HHT)
• Bruises
• New bruises -deeply purple, older bruises -yellow or green
• Bruises that occur only on areas reachable by the patient's hands (arms,
thighs, chest) may suggest self-inflicted injury, whereas bruises in
locations the patient cannot reach may suggest injury by another person
or a fall, depending on the site and pattern
• Hair follicle changes
• Perifollicular hemorrhages and "corkscrew hairs" are seen in scurvy
• Albinism Oculocutaneous albinism is associated with certain platelet
disorders (Hermansky-Pudlak and Chediak-Higashi syndromes)
28. TARGETED PHYSICAL EXAMINATION
• Other findings
• Splenomegaly
• May indicate underlying liver disease, lymphoma, or a
myeloproliferative neoplasm
• Joint laxity
• Joint hypermobility and skin hyperelasticity may be seen in some
Ehlers-Danlos syndromes (EDS) .Vascular EDS may lead to bleeding
due to arterial rupture
• Cardiac findings
• A harsh systolic murmur may indicate aortic stenosis, a cause of
acquired von Willebrand syndrome (AVWS), which may be associated
with bleeding from gastrointestinal arteriovenous malformations (AVMs)
• Macroglossia and organ infiltration
• Macroglossia and other signs of organ infiltration (carpal tunnel
syndrome and periorbital purpura may indicate amyloidosis, which can
cause a number of acquired clotting factor deficiencies (factors X and V,
von Willebrand factor [VWF], alpha-2-antiplasmin, and plasminogen
activator inhibitor- 1 [PAI-1])
• Syndromic findings
• Certain disorders have skeletal findings
(eg, thrombocytopenia-absent radius syndrome)
29. Clinical features of bleeding disorders
Bleeding
characteristics
Thrombocytopenia or
platelet function disorders
Clotting factor
deficiencies or
inhibitors
Major sites of
bleeding
Mucocutaneous (mouth,
nose, gastrointestinal tract,
urinary tract, menorrhagia)
Deep tissue (joints,
muscles) or soft tissue
hematomas
Petechiae Common Uncommon
Ecchymoses Generally small and
superficial. May be
significant, depending upon
the degree of
thrombocytopenia
May develop large
ecchymoses
Excessive bleeding
after minor cuts
Yes Not usually
Excessive bleeding
with surgery or
invasive procedures
Often immediate; severity is
variable(no excess bleeding
with mild thrombocytopenia,
severe bleeding with certain
platelet function disorders
such as GT)
Often during the
procedureIndividuals
with factor XIII
deficiency may
experience delayed
bleeding
31. LABORATORY EVALUATION
• Primary hemostasis
• Tests for primary hemostasis (platelet and vascular function defects) include:
• Testing for platelet function defects
• Platelet count and platelet morphology
• Additional testing can be done using platelet aggregation studies or the PFA-100
• Testing for VWD
• Von Willebrand factor (VWF) antigen, tests for VWF function, and factor VIII activity
• Secondary hemostasis
• Prothrombin time [PT]
• Activated partial thromboplastin time [aPTT]
• Thrombin time
• Reptilase time
• Specific clotting factor assay
• Mixing studies
• Prolongation usually indicates a deficiency or an inhibitor (usually an autoantibody) of
one or more clotting factors
• Mixing studies are PT or aPTT assays in which patient plasma is mixed 1:1 with
normal plasma
• This can distinguish between a deficiency (which is corrected by mixing because 50
percent of normal factor activity is sufficient for a normal clotting time) and an inhibitor
(which typically cannot be overcome by mixing)
32. • Tests for defects in fibrin crosslinking or fibrinolysis
• Factor XIII cross links fibrin to stabilize clots
• Factor XIII deficiency is a rare autosomal disorder that presents with
severe bleeding (eg, umbilical stump, intracranial hemorrhage)
• Rare cases of factor XIII inhibitors have also been reported
• Diagnostic testing is with qualitative or quantitative factor XIII levels
• Fibrinolysis (clot breakdown) is a normal hemostatic process
• Disorders of fibrinolysis often show delayed bleeding
• The cause may be inherited (eg, congenital deficiencies of
fibrinolytic inhibitors such as alpha-2-antiplasmin or plasminogen
activator inhibitor-1) or acquired (eg, hyperfibrinolysis associated
with envenomations, acute promyelocytic leukemia, overdoses of
fibrinolytic agents, prostate cancer, or DIC)
• Laboratory testing involves measurement of fibrinogen levels and D-
dimer (a fibrin degradation product) or related tests
• The euglobulin clot lysis test (ECLT) may also be used
• Tests for vascular and other disorders
• Vitamin C levels
• Genetic testing
39. Classification of functional platelet disorders
1.Disorders of adhesion
a)Inherited
Bernard-Soulier syndrome, von Willebrand’s disease
b)Acquired
Uremia, Acquired vWD
2.Disorders of aggregation
a)Inherited
Glanzmann’s thrombasthenia, Afibrinogenemia
b)Acquired
Fibrin degradation product inhibition, Dysproteinemias
Drugs-eg.ticlopidine, Gp IIb/IIIa inhibitors
3.Disorders of granule release
a)Inherited
Occulocutaneous albinism(Hermansky-Pudlak syndrome)
Chediak-Higashi syndrome
Isolated dense granule deficiency
Gray-platelet syndrome
b)Acquired
Cardiopulmonary bypass
Myeloproliferative disorders
Drugs –aspirin and other NSAID’S
40. • Platelet function testing
• Platelet aggregation studies – Platelet aggregation studies
measure platelet-platelet cohesion as induced by various
known platelet agonists (eg, thrombin, collagen, epinephrine,
ADP)
• Aggregation causes platelets to come out of solution, which
increases light transmission in a spectrophotometer-based
assay
• In Glanzmann thrombasthenia (GT; defect in platelet GPIIb-
IIIa complex), platelets will not aggregate to any agonists but
will agglutinate in response to ristocetin
• In Bernard-Soulier syndrome (BSS; defect in platelet GPIb-IX
complex), platelets will aggregate in response to thrombin,
collagen, epinephrine, and ADP but not to ristocetin
• In some granule disorders, platelets may show an abnormal
aggregation tracing (loss of the second wave of aggregation
in response to weaker platelet agonists)
• Aggregometry is considered the gold standard for platelet
function testing
41. • PFA-100
• The platelet function analyzer (PFA-100)
measures the time it takes for blood flow to stop
under shear stress in a capillary tube containing a
membrane impregnated with collagen and
epinephrine (Col/Epi) or collagen and ADP
(Col/ADP) and then exposed to shear stress
• Assesses both platelet adhesion and aggregation
• Normal closure time is 1-3 minutes
• Genetic testing
• Genetic testing for known platelet function
disorders (eg, using a gene panel) is becoming
more widely available and may be reasonable in
certain individuals with a suspected platelet
disorder of genetic origin
42.
43. • Electron microscopy
• Electron microscopy (EM) to examine the
ultrastructure of platelets is typically ordered by
specialized referral centers
• Transmission EM may be used to image the
ultrastructure of platelet granules in individuals with
suspected platelet storage pool diseases (SPDs; eg,
Chediak-Higashi syndrome, Hermansky-Pudlak
syndrome) who have a normal platelet
aggregometry and PFA-100
• EM can also be used to diagnose the nonmuscle
myosin heavy chain 9 (MYH9)-related disorders
(previously referred to a May-Hegglin
anomaly,Fechtner syndrome, Epstein syndrome,
and Sebastian syndrome)
44. • VWD testing
• Plasma VWF antigen (VWF:Ag)
• Testing for VWF function typically includes
the following Plasma VWF activity (ristocetin
cofactor [VWF:RCo] and collagen binding
[VWF:CB])
• Plasma factor VIII activity Analysis of
multimers should be done if there is low VWF
antigen or activity or if the ratio of VWF:RCo
to VWF:Ag is <0.7, suggesting the presence
of a dysfunctional VWF
• Levels of VWF fluctuate in response to
estrogens, stress, exercise, inflammation,
and bleeding; repeated assays may be
required to make the diagnosis
45. • The bleeding time (BT) is no longer
commonly used (and has largely been
replaced by PFA-100)
• The BT is performed at the bedside by
inflating a blood pressure cuff on the upper
arm to 40 mmHg, creating a cut of standard
size and depth on the forearm using an
automated device, and measuring the time
until bleeding stops
• The BT is typically prolonged in platelet
disorders, VWD, thrombocytopenia, and
disorders of vascular contractility.
• However, the test is insensitive and operator-
dependent
47. • Prothrombin time (PT) and INR
• The prothrombin time (PT) measures the time it takes plasma
to clot when exposed to tissue factor, which assesses the
extrinsic and common pathways of coagulation
• The PT test is performed by recalcifying citrated patient
plasma in the presence of tissue factor and phospholipid and
determining the time it takes to form a fibrin clot
• The formation of a fibrin clot is detected by visual, optical, or
electromechanical methods
• The result is measured in seconds and reported along with a
control value and/or an INR
• Normal range - 11 to 13 seconds
• The INR is dimensionless.
• Calculated as a ratio of the patient’s PT to a control PT
obtained using an international reference thromboplastin
reagent developed by the World Health Organization (WHO),
using the following formula:
• INR = [Patient PT ÷ Control PT]ISI
48. PT prolonged, aPTT normal
Inherited
Factor VII deficiency
Acquired
Mild vitamin K deficiency
Liver disease
Warfarin
Acute DIC
49. • Activated partial thromboplastin time (aPTT)
• The activated partial thromboplastin time (aPTT,)
measures the time it takes plasma to clot when exposed
to substances that activate the contact factors, which
assesses the intrinsic and common pathways of
coagulation
• The aPTT test is performed by recalcifying citrated
plasma in the presence of a thromboplastic material that
does not have tissue factor activity (hence the term
partial thromboplastin) and a negatively charged
substance (eg, celite, kaolin [aluminum silicate], silica),
which results in contact factor activation, thereby
initiating coagulation via the intrinsic clotting pathway
• The thromboplastic material provides a source of
phospholipids
• Normal range is approximately 25 to 35 seconds
50. PT normal, aPTT prolonged
inherited
Deficiency of factor VIII,IX,or XI
Deficiency of factor XII,prekallikrein or HMW Kininogen
Von willebrand disease
Acquired
Heparin,Dabigatran,Argatroban,Direct factor Xa inhibitors
Acquired inhibitor of factor VIII,IX,XI or XII
Acquired von willebrand syndrome
Lupus anticoagulant (more likely to be associated with thrombosis than bleeding)
51. Causes of prolonged PT and aPTT
Inherited
Deficiency of prothrombin,fibrinogen,factor V, or factor X
Combined factor deficiencies
Acquired
Liver disease
Acute DIC
Severe vitamin K deficiency
Anticoagulants (warfarin,Direct thrombin inhibitors)
Acquired inhibitor of prothrombin, fibrinogen, factor V or factor X
Amyloidosis-associated factor X deficiency
Anticoagulant rodenticide poisoning
52. • Thrombin time (TT)
• The thrombin time (TT) measures the final step of
coagulation, the conversion of fibrinogen to fibrin
• The test is performed by incubating citrated plasma in
the presence of dilute thrombin (bovine [cow] or
human) and measuring the time to clot formation
• Normal range approximately 14 to 19 seconds
• The thrombin time is prolonged if fibrinogen levels are
low or if an anticoagulant that inhibits thrombin is
present in the sample
• The TT may be used in the following clinical settings:
• Evaluation of a patient with a prolonged PT and aPTT
• Evaluation of an inherited fibrinogen disorder
• Detection of heparin in a sample
• If heparin is present, the TT will be significantly
prolonged and the reptilase time will be normal
53. • Causes of prolonged TT
• Anticoagulants
• Heparin, LMW heparin, and direct thrombin inhibitors
(eg, bivalirudin or argatroban) will prolong the TT
• Acquired fibrinogen disorders
• Prolonged in hypofibrinogenemia if the plasma fibrinogen
level is <100 mg/dL
• TT can also be prolonged in dysfibrinogenemias
• DIC
• Liver disease
• Hypoalbuminemia
• Paraproteinemias
• High concentrations of serum proteins, as occurs in
multiple myeloma or amyloidosis, can prolong the TT via
interference with fibrin polymerization
• Bovine thrombin exposure
54. • Reptilase time (RT)
• The reptilase time (RT) is similar to the TT in measuring the
conversion of fibrinogen to fibrin
• However, unlike the TT and the aPTT, the RT is insensitive to
the effects of heparin because reptilase, an enzyme derived
from the venom of Bothrops snakes, is not inhibited by
antithrombin or the antithrombin-heparin complex.
• The test is performed similarly to the TT (by incubating
citrated plasma in the presence of the diluted enzyme), with
the exception that reptilase is used instead of thrombin
• Reptilase differs from thrombin by generating fibrinopeptide
A, but not fibrinopeptide B, from fibrinogen and by resisting
inhibition by heparin via antithrombin (AT)
• The RT is useful for detecting abnormalities in fibrinogen (in
which case the TT is also prolonged) and in detecting the
presence of heparin; heparin will cause prolongation of the TT
but not RT. Similar to heparin, direct thrombin inhibitors
prolong the TT but not the RT.
• Inadvertent presence of a direct thrombin inhibitor is less
likely to be clinically relevant, but RT could be used to test for
this possibility.
55. • Lupus anticoagulant tests
• Indications and principles of LA testing
• Lupus anticoagulant (LA) assays can detect antiphospholipid
antibodies (aPL) that interfere with in vitro clotting tests that rely on
phospholipids in the assay.
• They are done for two main reasons:
• Evaluating for an aPL as the cause of unexplained prolongation of
the aPTT (or less commonly, PT).
• Testing for aPL in the diagnosis of antiphospholipid syndrome (APS)
• Sources of interference with LA testing
• Common interferences that may result in a false positive LA test
include elevated C-reactive protein (CRP), heparin, and DOACs.
• CRP is a phospholipid-binding protein that can mimic LA in test
systems and should be considered in any patient with a systemic
inflammatory state (eg, COVID-19).
• Heparin can be removed by adsorption or enzymatic degradation.
• DOACs can be effectively removed from plasma using activated
charcoal
56. • dRVVT
• The dilute Russell viper venom time (dRVVT, also
called Russel viper venom time [RVVT]) is a clotting
time test that takes advantage of the ability of the
venom from the Russell's viper (Daboia russelii) to
activate factor X directly
• The major use of the dRVVT is in testing for the LA
phenomenon caused by aPL. The dRVVT is
particularly sensitive to anti-beta-2-glycoprotein I
antibodies, which are most closely correlated with
thrombotic events and APS
• LA due to an aPL can be confirmed by adding
additional phospholipid to the assay
• The dRVVT is also sensitive to the effects of the
direct oral anticoagulants (DOACs) and has been
proposed as an assay that could be adapted to
DOAC monitoring, although this has not been
validated for this purpose and is not in clinical use
57. • Assays for specific coagulation factors
• Coagulation factor assays are mostly used to
diagnose specific factor deficiencies
• Inherited factor deficiencies, including
hemophilia A (factor VIII deficiency),
hemophilia B (factor IX deficiency), factor XI
deficiency, and other rare factor deficiencies
• Acquired factor inhibitors, based on the
finding of abnormal clotting times that fail to
correct in a mixing study
• In some cases, chromogenic assays may be
used to monitor therapy for hemophilia or to
monitor warfarin anticoagulation in a patient
with a baseline prolonged PT/INR
58. • Clot-based assays
• Factor activity can be measured by using
the aPTT (for intrinsic pathway factors) or
PT (for factor VII and common pathway
factors)
• These assays use a clotting endpoint and
are calibrated for individual factors using
factor-deficient plasmas and reported as
percent activity
• These assays, referred to as "one-stage"
clot-based assays, are the most commonly
used method to determine factor activity
levels
59. • Chromogenic assays — Chromogenic assays use cleavage of
a chromogenic (colored) substrate and a calibration curve to
assess factor activity.
• Factor VIII chromogenic assay — A chromogenic assay for
factor VIII activity can be useful in assessing factor VIII levels in
patients with aPTT interferences such as lupus anticoagulants.
Additionally, in some patients with hemophilia A, a chromogenic
factor VIII assay correlates better with bleeding phenotype than a
one-stage clot-based assay
• Chromogenic assays that use bovine reagents are required
when assessing underlying factor VIII activity in patients who are
receiving emicizumab
• For this reason, specialized coagulation laboratories and
hemophilia treatment centers often have both one-stage and
chromogenic factor VIII activity assays available for use
• Some of the recombinant or modified long half-life factor VIII and
factor IX products in many cases are better monitored with
chromogenic assays
60. • Factor X chromogenic assay — A chromogenic assay for
factor X activity is useful for monitoring warfarin therapy in
selected patients who have interferences with PT/INR
measurement, such as a prolonged PT/INR due to a lupus
anticoagulant
• The chromogenic factor X assay can also be used in
patients receiving argatroban or other direct thrombin
inhibitors who are being transitioned to warfarin
• The INR range of 2 to 3 corresponds to a chromogenic
factor X assay of approximately 20 to 30 percent.
• Of note, the chromogenic factor X assay is different from
the anti-factor Xa activity assay used to monitor
heparins, fondaparinux, and direct factor Xa inhibitors
61. • Antigenic assays
• Antigenic assays such as ELISAs
(enzyme-linked immunosorbent assays)
can also be used to measure clotting
factors
• This is typically done when there is a
need to distinguish quantitative from
qualitative factor deficiencies (decreases
in both antigen and functional activity
versus decreased functional activity with
preserved antigen levels)
• These assays are typically only available
at specialized referral centers.
62. • Fibrinogen
• Fibrinogen is the precursor to fibrin, the principle
component of a fibrin clot
• Abnormally low levels of fibrinogen (typically, <50 to 100
mg/dL) can result in impaired clot formation and increased
bleeding risk.
• Clinical uses of plasma fibrinogen levels include evaluation
for the following:
• Disseminated intravascular coagulation (DIC)
• Liver disease
• Inherited or acquired disorders of fibrinogen
64. • Clot solubility
• Tests that measure the solubility of clots in the
presence of a strong denaturing agent can detect
abnormalities of factor XIII,which crosslinks the fibrin
clot after it has formed
• Examples include solubility in 5M urea,1 percent
chloroacetic acid,or 2 percent acetic acid
• Lysis in any of these solutions within 24 hours
suggests factor XIII deficiency
• Factor XIII levels
65. • Fibrin D-dimer
• Fibrin D-dimer is one of the major fibrin degradation products
released upon cleavage of crosslinked fibrin by plasmin
• The dimer consists of two D domains from adjacent fibrin
monomers that have been crosslinked by activated factor XIII
• Normal plasma levels of D-dimer by ELISA testing are <500
ng/mL for fibrin equivalent units (FEU) or <250 ng/mL for D-
dimer units (DDU)
• Elevated concentrations of plasma D-dimer indicate recent or
ongoing intravascular coagulation and fibrinolysis
• Plasmin cleaves crosslinked fibrin at multiple sites, generating
other fibrin degradation products (FDPs), but D-dimer is the
best-studied and validated for clinical assessment
• Clinical uses of the D-dimer include evaluation for the following
• Deep vein thrombosis
• Pulmonary embolism
• DIC
• Primary hyperfibrinolysis
• Prognostic assessment in coronavirus disease 2019 (COVID-
19)
66. Laboratory diagnosis of abnormal
fibrinolysis
Test Primary
hyperfibrinolysis
DIC TTP
CBC and blood
smear
Normal Thrombocytopeni
a, MAHA
Thrombocytopeni
a, MAHA
PT and aPTT Normal or
prolonged
Prolonged Normal
Fibrinogen Decreased Decreased Normal
D-dimer or FDP* Increased Increased Normal
Antithrombin Normal Decreased Normal
Euglobulin clot
lysis time
Shortened Shortened Normal
ADAMTS13
activity
Normal Normal or mildly
reduced
Severely deficient
(usually <10%)
68. • HHT
• Hereditary hemorrhagic telangiectasia (HHT) is an
autosomal dominant disorder associated with
telangiectasias and arteriovenous malformations
(AVMs) of the small vessels in skin, oropharynx, lungs,
gastrointestinal tract, and other tissues
• Common findings include epistaxis, gastrointestinal
bleeding, telangiectasias on the lips and fingertips,
and iron deficiency anemia. Bleeding may begin in
childhood; by age 16, the majority of patients will
experience hemorrhagic symptoms
• Scurvy Scurvy is caused by vitamin C deficiency
• Symptoms are thought to be due to impaired collagen
synthesis and disordered connective tissue; these
generally occur when the vitamin C level is <0.2 mg/dL
(<11 micromol/L)
• Recent vitamin C intake can falsely normalize levels
69. Hereditary hemorrhagic telangiectasia Scurvy
petechiae on both legs, and corkscrew
hairs, gingival bleeding,
multiple bright red punctate macules and
papules a) Lips, b) Neck,c) Tongue,
d) Hardpalate, e) Finger tips, f) Nail folds
and nail beds
70. • Ehlers-Danlos syndrome
• Ehlers-Danlos syndrome (EDS) refers to a group of collagen
disorders characterized by easy bruising and hemorrhage from
ruptured blood vessels; there are several genes involved
• The classical EDS causing joint hypermotility and hyperextensibility
of the skin may cause bruising but is not likely to result in massive
bleeding
• The vascular type IV EDS is due to pathogenic variants in the
COL3A1 gene; bruising can be very extensive, and vascular rupture
can be fatal
• The skin may be thin and wrinkled, but joint hyperextensibility is rare
• Tests of platelet and clotting function are normal
• Diagnosis requires demonstration of the genetic abnormality or of
abnormal type III collagen
• Osteogenesis imperfecta
• Osteogenesis imperfecta (OI) is a connective tissue disorder
that primarily affects bone but can also be associated with
bleeding due to capillary fragility
71. • Isolated laboratory abnormalities with negative
bleeding history
• Certain coagulation factor abnormalities will affect
the coagulation studies but are not associated with
clinical bleeding
• These include deficiencies of high molecular
weight kininogen, prekallikrein, and factor XII,
many dysfibrinogenemias, as well as certain
mutations in the factor VII molecule that affect its
interaction with bovine but not human
thromboplastin (this occurs if the laboratory is
using bovine thromboplastin as the tissue factor
reagent in the PT assay).
• Some individuals with other factor deficiencies
may not have bleeding (eg, certain individuals with
factor XI deficiency or mild factor VII deficiency).
72. • Isolated family history of a bleeding
disorder
• Individuals who have a family history of a
bleeding disorder may request an evaluation
for the disorder even if they have not had
bleeding (eg, prior to surgery or to assist with
preconception counseling)
• In such individuals, it is reasonable to review
the family member's diagnosis to determine if
it is in fact a heritable condition. If so, it is
reasonable to test the individual, especially
prior to surgery. Often this will entail a
screening test plus specific testing (eg, for
hemophilia, aPTT plus factor VIII level)
73. CASE SCENARIO 1
• PT-Normal
• APTT-Normal
• TT-Normal
• Fibrinogen-Normal
• Platelet count-Normal
• Interpretation
• Normal hemostasis
• Disorders of platelet function
• Mild von Willebrand disease
• Factor XIII deficiency
• Disorders of fibrinolysis
• Vascular disorders of hemostasis
• Second line investigations
• Clot solubility test
• Specific factor assay for suspected factor deficiency factor XIII
• PFA 100
• Testing for VWD
74. CASE SCENARIO 2
• PT-Prolonged
• APTT-Normal
• TT-Normal
• Fibrinogen-Normal
• Platelet count-Normal
• Interpretation
• Factor VII deficiency
• Vitamin K deficiency
• Liver disease
• Warfarin
• Second line investigations
• Mixing test
• Factor VII assay
• Liver function test
75. CASE SCENARIO 3
• PT-Normal
• APTT-Prolonged
• TT-Normal
• Fibrinogen-Normal
• Platelet count-Normal
• Interpretation
• Deficiency of factor VIII,IX,XII,XI,Prekallikrein,HMWK
• Acquired inhibitor of factor VIII,IX,XI or XII
• Heparin,Dabigatran,Argatroban,Direct factor Xa inhibitors
• von Willebrand disease
• Lupus anticoagulant
• Second line investigations
• Mixing test
• Factor VIII,IX assay
• Lupus anticoagulant
76.
77.
78.
79.
80.
81. CASE SCENARIO 4
• PT-Prolonged
• APTT-Prolonged
• TT-Normal
• Fibrinogen-Normal
• Platelet count-Normal
• Interpretation
• Deficiency of prothrombin,factor V, or factor X
• Severe vitamin K deficiency
• Anticoagulants
• Acquired inhibitor of prothrombin,factor V or factor X
• Amyloidosis-associated factor X deficiency
• Anticoagulant rodenticide poisoning
• Second line investigations
• Mixing test
• Specific factor assay
• Liver function test
82. CASE SCENARIO 5
• PT-Prolonged
• APTT-Prolonged
• TT-Prolonged
• Fibrinogen -Normal/Abnormal
• Platelet count-Normal
• Interpretation
• Unfractionated heparin
• Hypofibrinogenemia
• Afibrinogenemia
• Dysfibrinogenemia
• Systemic hyperfibrinolysis with increased FDP e.g DIC
• Some cases of liver disease
• Second line investigations
• Reptilase or ancord time
• D-Dimer level
83. CASE SCENARIO 6
• PT- Prolonged
• APTT-Prolonged
• TT-Normal
• Fibrinogen-Normal/Low
• Platelet count-Low
• Interpretation
• Massive transfusion of stored/plasma depleted blood
• DIC
• Chronic liver disease esp.cirrhosis
• Second line investigations
• Specific factor assay
• Peripheral blood smear
• Bone marrow aspiration
84. CASE SCENARIO 7
• PT-Normal
• APTT-Normal
• TT-Normal
• Fibrinogen-Normal
• Platelet count-Low
• Interpretation
• Thrombocytopenia
• Futher work up for thrombocytopenia
(peripheral smear,bone marrow aspiration etc)
85. CASE SCENARIO 8
• PT-Prolonged
• APTT-Prolonged
• TT-Prolonged
• Fibrinogen-Low
• Platelet count-Low
• Interpretation
• DIC
• Acute liver necrosis with DIC
• Second line investigations
• FDP or D-Dimer assay
86. INDICATIONS FOR HEMATOLOGIST REFERRAL
• Active bleeding that requires administration of hemostatic
products
• Abnormal initial testing that cannot be easily explained (eg, by
presence of an anticoagulant) and/or that persists upon retesting
• Test results consistent with a specific hematologic disorder (eg,
von Willebrand disease [VWD], immune thrombocytopenia [ITP])
• Concern about a bleeding disorder based on personal or family
history, with normal initial testing
• Concern that a previous diagnosis may be inaccurate (eg,
individual with borderline von Willebrand factor [VWF] levels told
they have VWD)
• Concern about a bleeding disorder and upcoming surgery
• Question about whether anticoagulant or antiplatelet medications
can be used in an individual with a mild bleeding disorder
• Family history of a bleeding disorder with upcoming surgery,
delivery, or desire for prenatal counseling
87. DIFFERENTIAL DIAGNOSIS
• Trauma
• Trauma from frequent falls or other injuries (eg, due to
loss of balance, self-injury, or physical abuse) can
produce ecchymoses and/or internal bleeding.
• Vasculitis
• Some vasculitides (especially small-vessel) produce
palpable purpura. Unlike purpura from
thrombocytopenia, purpura associated with vasculitis
is palpable and the platelet count is typically normal
• Psychogenic purpura
• Psychogenic purpura refers to purpura or bleeding in
an unusual pattern that often resembles religious
stigmata; it is a diagnosis of exclusion. The cause is
not well understood, and psychiatric factors are often
implicated
• Dysproteinemias Bleeding due to dysproteinemias
such as seen in plasma cell dyscrasias
