Investigations
1st investigations to order
FBC
Test
An initial laboratory screening test for patients with symptoms of a bleeding disorder.
FBC may be normal in von Willebrand disease (VWD), or may show anaemia (due to ongoing blood loss and iron deficiency) or thrombocytopenia (e.g., in type 2B VWD or platelet type [pseudo]-VWD).[15][32]
Abnormal blood count may indicate acquired von Willebrand syndrome associated with a myeloproliferative neoplasm (e.g., essential thrombocythemia).
Result
usually normal; may be abnormal (e.g., low haemoglobin [anaemia]; low platelet count [in type 2B VWD]; elevated platelet count [in AVWS associated with essential thrombocythemia])
prothrombin time (PT)
Test
An initial laboratory screening test for patients with symptoms of a bleeding disorder.
Usually normal in von Willebrand disease.
Result
normal
activated partial thromboplastin time (aPTT)
Test
Usually normal in von Willebrand disease (VWD), but may be prolonged if patients have reduced factor VIII (FVIII) levels (e.g., in type 2N or type 3 VWD).[32]
A normal aPTT does not exclude the diagnosis of VWD.
Result
usually normal; may be prolonged if FVIII level is low (<0.40 IU/mL)
von Willebrand factor (VWF) antigen (VWF:Ag)
Test
An initial blood test to diagnose von Willebrand disease (VWD).[15]
Type 1 VWD is confirmed if VWF:Ag level is <0.30 IU/mL regardless of bleeding (or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is normal (i.e., >0.7).[15] If type 1C VWD is suspected, VWF levels are checked at 1 and 4 hours following a desmopressin infusion.[15][50]
Type 2A, 2B, or 2M VWD is confirmed if VWF:Ag level is <0.30 IU/mL regardless of bleeding (or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is abnormal (i.e., <0.7).[15] Additional tests (including VWF multimer analysis, VWF collagen binding assay [VWF:CB], low-dose ristocetin-induced platelet agglutination [RIPA], and genetic testing) are required to confirm the specific subtype of type 2 VWD.[15] In type 2N, VWF:Ag level may be normal.[52] Alternative tests are required to diagnose type 2N (e.g., FVIII coagulant activity assay [FVIII:C]).
A diagnosis of type 3 VWD can be made if VWF:Ag is undetectable (<0.01 IU/mL).[36]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous oestrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous oestrogen therapy.[36][51]
Result
<0.30 IU/mL (or 0.30 to <0.50 IU/mL with abnormal bleeding)
von Willebrand factor (VWF) platelet-binding activity assay (VWF:GPIbM, VWF:GPIbR, or VWF:RCo)
Test
An initial blood test to diagnose von Willebrand disease (VWD).[15]
Use of VWF mutant glycoprotein Ibɑ binding assay (VWF:GPIbM) or VWF recombinant glycoprotein Ibɑ binding assay (VWF:GPIbR) to assess VWF platelet-binding activity is recommended over the VWF ristocetin cofactor assay (VWF:RCo).[15] These newer assays have lower variability and higher reproducibility, and are not affected by the VWF D1472H polymorphism (a benign VWF gene variant that impairs binding of VWF to ristocetin and can lead to a false diagnosis).[49][50]
Type 1 VWD is confirmed if VWF platelet-binding activity is <0.30 IU/mL regardless of bleeding (or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is normal (i.e., >0.7).[15] Additional tests are required to confirm type 1C VWD if suspected (e.g., desmopressin trial with VWF levels checked at 1 and 4 hours post-infusion; or VWF propeptide analysis).[15][50]
Type 2A, 2B, or 2M VWD is confirmed if VWF platelet-binding activity is <0.30 IU/mL regardless of bleeding (or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is abnormal (i.e., <0.7).[15] Additional tests (including VWF multimer analysis, VWF collagen binding assay [VWF:CB], low-dose ristocetin-induced platelet agglutination [RIPA], and genetic testing) are required to confirm the specific subtype of type 2 VWD.[15] In type 2N, VWF platelet-binding activity may be normal.[52] Alternative tests are required to diagnose type 2N (e.g., FVIII coagulant activity assay [FVIII:C]).
In type 3 VWD, VWF platelet-binding activity is undetectable (<0.01 IU/mL).[36]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous estrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous estrogen therapy.[36][51]
Result
<0.30 IU/mL (or 0.30 to <0.50 IU/mL with abnormal bleeding)
factor VIII (FVIII) coagulant activity assay (FVIII:C)
Test
An initial blood test to diagnose VWD. VWF stabilises FVIII in the blood and prevents proteolytic degradation, so reduced VWF leads to a similar decrease in FVIII.
In type 1 VWD, FVIII:C level is often normal, but may be low in more severe disease. In patients with type 2A, 2B and 2M VWD, FVIII:C level may be normal to mildly reduced. FVIII:C level is often very low in patients with type 3 VWD.
In patients with type 2N VWD, FVIII:C level is low relative to VWF:Ag level (similar to patients with mild haemophilia A).[15][36][52] It is important to differentiate type 2N VWD and haemophilia A because these conditions are managed differently. Additional tests required to establish a diagnosis of type 2N VWD include FVIII binding assay (VWF:FVIIIB) and genetic testing (for type 2N variants).[15]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous oestrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous oestrogen therapy.[36][51]
Result
normal to decreased
Investigations to consider
fibrinogen
Test
May be ordered for patients with symptoms of a bleeding disorder.[15]
Usually normal in von Willebrand disease.
Result
normal
thrombin time
Test
May be ordered for patients with symptoms of a bleeding disorder.[15]
Usually normal in von Willebrand disease.
Result
normal
von Willebrand factor (VWF) multimer analysis
Test
Should be performed to confirm the specific subtype of type 2 VWD (e.g., type 2A, 2B, or 2M) if VWF platelet-binding activity is abnormal (i.e., <0.30 IU/mL regardless of bleeding; or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is abnormal (i.e., <0.7).[15]
A normal VWF multimer analysis confirms a diagnosis of type 2M VWD.
An abnormal VWF multimer analysis suggests type 2A or 2B VWD; these two subtypes can be differentiated using genetic testing (for type 2B variants) and low-dose ristocetin-induced platelet agglutination (RIPA) test.
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous estrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous estrogen therapy.[36][51]
Result
normal (VWF multimers present) in type 2M VWD; abnormal (VWF multimers absent) in type 2A and 2B VWD
von Willebrand factor (VWF) collagen binding assay (VWF:CB)
Test
Should be performed to confirm the specific subtype of type 2 VWD (e.g., type 2A, 2B, or 2M) if VWF platelet-binding activity is abnormal (i.e., <0.30 IU/mL regardless of bleeding; or 0.30 to <0.50 IU/mL for patients with abnormal bleeding), and VWF platelet-binding activity/VWF:Ag ratio is abnormal (i.e., <0.7).[15]
A normal VWF:CB/VWF:Ag ratio confirms a diagnosis of type 2M VWD.
An abnormal (reduced) VWF:CB/VWF:Ag ratio suggests type 2A or 2B VWD; these two subtypes can be differentiated using genetic testing (to identify type 2B VWD variants).[15][36]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous oestrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous oestrogen therapy.[36][51]
Result
normal VWF:CB/VWF:Ag ratio in type 2M VWD; abnormal (reduced) VWF:CB/VWF:Ag ratio in type 2A and 2B VWD
genetic testing (for type 2B or 2N VWD variants)
Test
Should be performed to confirm type 2B or 2N VWD if these subtypes are suspected.[15][36]
Genetic testing for type 2B variants can be used to differentiate type 2B VWD and platelet type (pseudo)-VWD.[36]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous oestrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous oestrogen therapy.[36][51]
Result
positive for type 2B or 2N VWD variants
low-dose ristocetin-induced platelet agglutination (RIPA) mixing studies
Test
Can be used to differentiate type 2B VWD and platelet type (pseudo)-VWD.[36]
An abnormal low-dose RIPA test (i.e., enhanced platelet agglutination at low ristocetin concentrations) is characteristic of type 2B VWD and platelet type (pseudo)-VWD.[36] However, low-dose RIPA mixing studies (combining normal donor plasma with patient platelets, and combining normal platelets with patient plasma) can determine if the cause is an underlying defect with the VWF protein (i.e., type 2B VWD) or with the platelets (i.e., platelet type [pseudo]-VWD).
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous estrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous estrogen therapy.[36][51]
Result
abnormal (enhanced platelet agglutination) in type 2B VWD
von Willebrand factor (VWF) FVIII binding assay (VWF:FVIIIB)
Test
Can be used to differentiate type 2N VWD and haemophilia A in patients with low factor VIII (FVIII) coagulant activity (FVIII:C) levels.[15][36][52]
It is important to differentiate type 2N VWD and haemophilia A because these conditions are managed differently.
A diagnosis of type 2N VWD is confirmed in those with low FVIII:C levels if VWF:FVIIIB is abnormal (i.e., reduced VWF:FVIIIB/VWF:Ag ratio) and genetic testing is positive for type 2N variants, or genetic testing alone is positive for type 2N variants.[15]
A diagnosis of type 2N VWD can be ruled out if genetic testing is negative for type 2N variants, or if VWF:FVIIIB is normal (i.e., normal VWF:FVIIIB/VWF:Ag ratio).
VWF:FVIIIB levels may vary in type 2N VWD depending on the inheritance pattern (e.g., lower in homozygotes and compound heterozygotes compared with heterozygotes).[36]
Diagnostic tests for VWD should be repeated to avoid misdiagnosis.[36] If the patient history or bleeding assessment tool (if used for initial screening) indicates an abnormality of haemostasis but initial blood tests for VWD are normal, then further investigations for platelet, coagulation, and blood vessel wall abnormalities should be pursued.
VWF is an acute phase reactant and may be elevated in patients who are pregnant or receiving exogenous estrogen therapy. Diagnostic testing for VWD in these patients may lead to inaccurate results and require repeating. Testing should be avoided during pregnancy or while the patient is receiving exogenous estrogen therapy.[36][51]
Result
abnormal (reduced) VWF:FVIIIB/VWF:Ag ratio in type 2N VWD; normal in haemophilia A
VWF propeptide assay (VWFpp/VWF:Ag)
Test
May be helpful for diagnosis of type 1C VWD, which is characterised by accelerated clearance (shortened half-life) of VWF. Observational studies suggest that a substantial increase in the VWFpp/VWF:Ag ratio is predictive of a significantly decreased VWF half-life, but the certainty of evidence was judged to be very low in one systematic review.[50]
The 2021 ASH ISTH NHF WFH (American Society of Hematology, International Society on Thrombosis and Haemostasis, National Hemophilia Foundation, and World Federation of Hemophilia) guideline recommends using a desmopressin trial over the VWF propeptide assay (VWFpp/VWF:Ag) to confirm increased clearance of VWF, but they acknowledge that the VWFpp/VWF:Ag ratio may be helpful for patients who cannot safely undergo a desmopressin trial (e.g., very young or very old patients).[15]
Result
VWF:FVIIIB/VWF:Ag ratio may be increased in type 1C VWD
platelet function analyser (e.g., PFA-100)
Test
A platelet function analyser (e.g., PFA-100) can be used to screen for Von Willebrand disease (VWD; particularly more severe types), but they are not thought to be sensitive and specific enough to be widely recommended for the diagnosis of VWD.[48][36]
A specific diagnosis does not follow directly from an abnormal screening result; therefore, further testing is needed.
Result
may be abnormal
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