Haemophilia (2014), 1–7
DOI: 10.1111/hae.12437
ORIGINAL ARTICLE
Evaluation of a rapid von Willebrand Factor Activity Latex Immuno Assay for monitoring of patients with von Willebrand disease (VWD) receiving DDAVPâ or VWF replacement therapy S . V I N A Y A G A M , L . R . S I M O N S , P . C H O W D A R Y , P . T H U R L O W , S . V . B R O O K S and A. F. RIDDELL Katharine Dormandy Haemophilia Centre & Thrombosis Unit, Royal Free London NHS Foundation Trust, London, United Kingdom
Summary. Haemostatic management of surgery in patients with von Willebrand disease (VWD) includes DDAVPâ or von Willebrand factor (VWF)-containing concentrates. Although the recommendations are for monitoring by VWF activity assays, it is quite common for clinicians to use factor VIII due usually to longer turnaround times required for VWF ristocetin cofactor assay (VWF:RCo) measurements. The aim of this study was to evaluate use of the rapid HaemosILTM VWF activity (VWF:Act) latex immuno assay (LIA) on an automated coagulometer (ACL TOPTM 700; Instrumentation Laboratory, Bedford, MA, USA) compared to platelet-based VWF:RCo assays in this setting. One hundred and sixty-seven plasma samples from 42 patients [Type 1 (n = 22), Type 2A (n = 2), Type 2B (n = 3), Type 2M (n = 10), Type 3 (n = 3)] and acquired von Willebrand syndrome (n = 2) with VWD treated with DDAVPâ or VWF-containing concentrates were included in the study. Method comparison and method bias were
Introduction von Willebrand factor (VWF), a plasma glycoprotein synthesized in megakaryocytes and vascular endothelial cells is secreted into plasma and subendothelial compartments in response to vascular injury or stimulation by endogenous or exogenous secretagogues. VWF plays a critical role in primary haemostasis, Correspondence: Anne Riddell, Katharine Dormandy Haemophilia centre and Thrombosis unit, Royal Free London NHS Foundation Trust, Pond Street, London, United Kingdom, NW3 2QG. Tel: +442078302274; fax: +442078302879; e-mail: [email protected] Accepted after revision 11 March 2014 © 2014 John Wiley & Sons Ltd
evaluated by Bland–Altman analysis (BA) and Passing and Bablok regression modelling respectively. BA of baseline samples (n = 39) showed a mean difference of 3.0 (1.96 SD 25.2 to +19.4). Post (treatment) samples (n = 120) were separated into two groups. Group 1 contained samples with VWF:RCo levels 10 to ≤175 IU dL 1 (n = 97) and group 2, samples with VWF:RCo levels >175 IU dL 1 (n = 23). BA of group 1 postsamples showed a mean difference of +3.4 (1.96 SD 44.6 to +51.5), and the BA of Group 2 samples was 23.9 (1.96 SD 136.1 to +88.3). In conclusion, use of HaemosIL VWF:Act LIA test on an automated coagulometer is a reproducible and rapid assay that can be used as an alternative test for monitoring VWF replacement therapy, facilitating dose adjustments on a real-time basis. Keywords: HaemosILTM von Willebrand factor activity, ristocetin cofactor assays, treatment monitoring, von Willebrand disease, von Willebrand factor
serving as a mediator for platelet adhesion and aggregation at sites of vascular injury, as well as acting as a carrier protein to stabilize factor VIII (FVIII) in the circulation [1]. von Willebrand disease (VWD) is a congenital bleeding disorder resulting from a quantitative or qualitative deficiency of VWF function. It is the most common hereditary bleeding disorder, with laboratory abnormalities of VWF present in up to 1% of the population [2]. The quantity of VWF protein present in plasma can be measured immunologically as VWF antigen (VWF:Ag). VWF activity is commonly measured by its ability to facilitate platelet agglutination in the presence of the antibiotic ristocetin, as VWF ristocetin cofactor assay (VWF:RCo). It is also important to measure FVIII activity (FVIII:C) in patients with VWD, as it is a reflection of the ability 1
2
S. VINAYAGAM et al.
of VWF to bind and maintain the level of FVIII in the circulation. Individuals with VWD are at risk for surgical bleeding, both intra-operatively and in the postoperative period. Haemostasis in these patients requires the correction of the defect in VWF-platelet-vessel wall interactions and the deficiency of FVIII. Correction of the defect in VWF function is most important perioperatively and in the immediate postoperative period, particularly when mucosal bleeding is possible. Therapeutic FVIII:C levels also need to be maintained peri- and postoperatively for 7–10 days [3]. Strategies for perioperative management of patients with VWD depend on the type of deficiency, baseline levels of VWF and FVIII:C, the patient’s historical responses to 1-deamino-8-D-arginine vasopressin, (DDAVPâ) and the site and invasiveness of the planned procedure. United Kingdom Haemophilia Centre Doctors’ organization guidelines recommend covering operative procedures with DDAVPâ in responsive patients unless contraindicated. Treatment of surgical bleeding with VWF-containing concentrates is advised for patients with VWD who do not respond adequately to DDAVPâ or in whom use of this agent is contraindicated. It is recommended that the VWF:RCo should be maintained at above 50 IU dL 1 in the perioperative period and FVIII:C maintained at around 100 IU dL 1 during major surgery and above 50 IU dL 1 in the postoperative period, until haemostasis is secure. The guidelines recommend that preferentially VWF:RCo should be monitored, in reality FVIII:C is commonly used, although it is a surrogate marker and does not truly reflect the kinetics of VWF replacement therapy, especially in patients with type 2 and 3 VWD. Whilst the VWF:RCo assay is the standard assay used to evaluate VWF activity, it is labour intensive with a high coefficient of variation [4] and in our laboratory is subject to an urgent turnaround time of at least one hour. Measurement of VWF collagen binding activity (VWF:CB) has been proposed as an alternative method of monitoring treatment in patients with VWD but it is also time consuming, requiring an enzyme linked immunosorbent assay (ELISA) technique. The availability of a rapid method of VWD treatment monitoring in VWD would enable more accurate and responsive prophylaxis of bleeding during major surgery, potentially avoiding excessive administration of concentrates and reducing the risk of perioperative haemorrhage and venous thromboembolic complications. The HaemosILTM VWF activity [Instrumentation Laboratory (IL), Bedford, MA, USA] latex immuno assay (LIA) contains a purified anti mouse monoclonal antibody (RFF VIII:R/1) directed against a functional epitope of VWF–Glycoprotein 1b (platelet binding) receptor site–adsorbed onto latex particles, also containing bovine serum albumin, stabilizers and preservative that interacts with VWF in a plasma Haemophilia (2014), 1--7
sample [5]and from now the test will be referred to as ‘VWF:Act’. The VWF:Act test is independent of ristocetin and can be fully automated and run on a number of different analysers. Previous studies of the assay have satisfactorily evaluated its use in identifying patients with VWD [6–8] but the use of this antibody in an ELISA setting has previously demonstrated limitations in its sensitivity to some qualitative variants of VWF [9]. However, this simple automated latex assay potentially enables laboratories to provide a more rapid evaluation of VWF function and shorter turnaround time for monitoring response to treatment in the perioperative setting. In our laboratory, the VWF:Act test takes 15 min (if test previously calibrated) in comparison to 1 h for a platelet-based VWF:RCo assay result. In this study, we compared the performance of HaemosILTM VWF Activity assay on an ACL TOPTM 700 coagulometer (IL) with VWF:RCo assays using lyophilized platelet reagents in VWD patients receiving VWF-containing concentrates and DDAVPâ for a variety of surgical procedures for accuracy and clinical usefulness.
Material and methods This was a retrospective study of patients with VWD receiving either DDAVPâ or VWF-containing concentrates such as Haemateâ P (CSL Behring, Marburg, Germany), Wilfactinâ (LFB, Les Ulis, France) or Alphanateâ (Grifols, Barcelona, Spain) for a range of reasons including major and minor surgeries, treatment of mucocutaneous bleeding episodes, gastrointestinal bleeding and excessive nose bleeding. Venous whole blood was collected into 0.106 M trisodium citrate tubes (Becton & Dickinson, UK), centrifuged at 2100 g for 12 min, plasma separated into two aliquots, and stored at 40°C until testing. In total 169 plasma samples from 42 patients with VWD with the following subtypes were included: Type 1 (n = 22), Type 2A (n = 2), Type 2B (n = 3), Type 2M (n = 10), Type 3 (n = 3) and acquired von Willebrand syndrome (n = 2). The median age of the patient group was 37 years (range 5–83 years) and included males (n = 21) and females (n = 19). Of the 169 Samples, 46 were baseline samples and 123 samples were taken after treatment at variable time intervals. The details of the treatment material and subtype of VWD are presented in Table 1. VWF:RCo levels were measured using lyophilizedplatelet-based VWF:RCo methods on either a platelet aggregation profiler 4 (PAP-4) aggregometer (Bio-Data, Horsham, PA, USA) or on an ACL9000 (IL). Briefly, diluted patient, standard or quality control (QC) plasma samples were added to either lyophilized platelets (Bio Data) and ristocetin (10 mg mL 1) (Abp, London, UK) and end point measured using the PAP-4 aggregometer or added to lyophilized platelets (BCâ VW reagent, Siemens Healthcare Diagnostics, Marburg, © 2014 John Wiley & Sons Ltd
EVALUATION OF A RAPID VWF ACTIVITY LATEX IMMUNO ASSAY
3
Table 1. Showing number of samples by type of VWD and type of treatment received. Type of VWD Treatment material
Type 1
Type 2A
Type 2B
Type 2M
Type 3
AVWS*
Total number (n) of samples
Alphanateâ Haemateâ P Wilfactinâ DDAVPâ Total (n) samples by VWD subtype
6 1 2 16 25
1 1 – – 2
2 – 1 – 3
3 5 2 1 11
1 2
– 2 – – 2
13 11 5 17 46
– 3
AVWS, acquired von Willebrand syndrome; VWD, von Willebrand disease. *Acquired von Willebrand disease.
Germany) with added ristocetin (50 mg mL 1) based on the method by Lattuada et al. [10] and the end point measured on an ACL9000. Prior to use of ACL9000 VWF:RCo method, a comparison with the PAP-4 VWF: RCo method was carried out. The correlation between VWF:RCo methods was good (r = 0.9562, P = 175 IU dL 1). It would be useful for laboratories to have a rapid assay to monitor VWF function in patients requiring VWF replacement therapy who are undergoing major and minor surgical episodes. Recently, newer types of Haemophilia (2014), 1--7
assays for measuring VWF function using recombinant glycoprotein 1b molecules have been developed, but there is limited literature on their use in monitoring VWF replacement therapy [27,28]. The VWF:Act is an easy to perform assay that is able to produce a result quickly when compared to lyophilized platelet VWF:RCo assays (15 min vs. 1 h respectively) and therefore will help facilitate a rapid turnaround of results in urgent settings especially prior to surgery. Because the VWF:Act test is very reproducible in comparison to the VWF:RCo assay on an automated coagulometer (intra- and inter-assay CVs 3.7% and 4.4% vs. 5.4% and 9.3% respectively) it therefore seems reasonable that this test can be the assay of choice for monitoring VWF replacement therapy, facilitating dose adjustments on a real-time basis allowing for optimization of treatment and preventing under-treatment in the first 48 h with resultant bleeding and over-treatment after 48 h with the attendant risk of thrombosis. Since the initial validation we now routinely use the VWF:Act test for monitoring VWF replacement © 2014 John Wiley & Sons Ltd
EVALUATION OF A RAPID VWF ACTIVITY LATEX IMMUNO ASSAY
therapy with occasional retrospective cross checking with VWF:RCo assay measurements on patient samples (especially when they first receive treatment and their response is not known). This helps to facilitate shorter turnaround times for monitoring patients receiving VWF treatments (DDAVPâ and VWF-containing concentrates) that are within acceptable clinical time frames. In conclusion, the HaemosILTM VWF: Activity test is a rapid and reliable automated assay that can be used for monitoring VWF functional activity of VWF-containing concentrates or DDAVPâ therapy in patients with VWD.
References 1 Wagner DD, Bonfanti R. von Willebrand factor and the endothelium. Mayo Clin Proc 1991; 66: 621–7. 2 Rodeghiero F, Castaman G, Dini E. Epidemiological investigation of the prevalance of von Willebrand’s disease. Blood 1987; 69: 454–9. 3 Pasi KJ, Collins PW, Keeling DM et al. Management of von Willebrand disease: a guideline from the UK Haemophilia Centre Doctors’ Organization. Haemophilia 2004; 10: 218–31. 4 Kitchen S, Jennings I, Woods TA, Kitchen DP, Walker ID, Preston FE. Laboratory tests for measurement of von Willebrand factor show poor agreement among different centers: results from the United Kingdom National External Quality Assessment Scheme for Blood Coagulation. Semin Thromb Hemost 2006; 32: 492–8. 5 De Vleeschauwer A, Devreese K. Comparison of a new automated von Willebrand factor activity assay with an aggregation von Willebrand ristocetin cofactor activity assay for the diagnosis of von Willebrand disease. Blood Coagul Fibrinolysis 2006; 17: 353–8. 6 Sucker C, Senft B, Scharf RE, Zotz RB. Determination of von Willebrand factor activity: evaluation of the HaemosIL assay in comparison with established procedures. Clin Appl Thromb Hemost 2006; 12: 305–10. 7 Trossaert M, Ternisien C, Lefrancois A et al. Evaluation of an automated von Willebrand factor activity assay in von Willebrand disease. Clin Appl Thromb Hemost 2011; 6: 25–9. 8 Pinol M, Sales M, Costa M, Tosetto A, Canciani MT, Federici AB. Evaluation of a new turbidimetric assay for von Willebrand factor activity useful in the general screening of von Willebrand disease. Haematologica 2007; 92: 712–3. 9 Nitu-Whalley IC, Riddell A, Lee CA et al. Identification of type 2 von Willebrand disease in previously diagnosed type 1 patients: a reappraisal using phenotypes, genotypes and molecular modelling. Thromb Haemost 2000; 84: 998–1004. 10 Lattuada A, Preda L, Sacchi E, Gallo L, Federici AB, Rossi E. A rapid assay for
© 2014 John Wiley & Sons Ltd
11
12
13
14
15
16
17
18
19
20
7
Author contributions SV and SB performed the assays and wrote first draft of the manuscript. LS and PC reviewed and wrote the manuscript. PT collected the patient samples and helped to collate treatment information. AR designed the study, analysed the data, and reviewed the manuscript. Thanks to Vincent P Jenkins for helpful discussion on the manuscript.
Disclosures The authors stated that they had no interests which might be perceived as posing a conflict or bias.
ristocetin cofactor activity using an automated coagulometer (ACL 9000). Blood Coagul Fibrinolysis 2004; 15: 505–11. Riddell AF, Jenkins PV, Nitu-Whalley IC, McCraw AH, Lee CA, Brown SA. Use of the collagen-binding assay for von Willebrand factor in the analysis of type 2M von Willebrand disease: a comparison with the ristocetin cofactor assay. Br J Haematol 2002; 116: 187–92. Bland JM, Altman DG. Measuring agreement in method comparison studies. Stat Methods Med Res 1999; 8: 135–60. Passing H, Bablok W. A new biometrical procedure for testing the equality of measurements from two different analytical methods. Application of linear regression procedures for method comparison studies in clinical chemistry, Part I. J Clin Chem Clin Biochem 1983; 21: 709–20. Mannucci PM. Treatment of von Willebrand’s disease. N Engl J Med 2004; 351: 683–94. Batlle J, Lopez-Fernandez MF, Fraga EL, Trillo AR, Perez-Rodriguez MA. Von Willebrand factor/factor VIII concentrates in the treatment of von Willebrand disease. Blood Coagul Fibrinolysis 2009; 20: 89– 100. Berntorp E, Archey W, Auerswald G et al. A systematic overview of the first pasteurised VWF/FVIII medicinal product, Haemate P/Humate-P: history and clinical performance. Eur J Haematol 2008; 80 (Suppl. 70): 3–35. Franchini M. Surgical prophylaxis in von Willebrand’s disease: a difficult balance to manage. Blood Transfus 2008; 6(Suppl. 2): 33–8. Michiels JJ, van Vliet HH, Berneman Z, Schroyens W, Gadisseur A. Managing patients with von Willebrand disease type 1, 2 and 3 with desmopressin and von Willebrand factor-factor VIII concentrate in surgical settings. Acta Haematol 2009; 121: 167–76. Hay CR, Hellmann A, Dmoszynska A et al. Experience with Optivate(R), a new high purity concentrate of factor VIII with von Willebrand factor, in patients undergoing surgery. Haemophilia 2011; 17: 428–32. Windyga J, Von Depka-Prondzinski M. Efficacy and safety of a new generation von
21
22
23
24
25
26
27
28
Willebrand factor/factor VIII concentrate (Wilate(R)) in the management of perioperative haemostasis in von Willebrand disease patients undergoing surgery. Thromb Haemost 2011; 105: 1072–9. Lethagen S, Kyrle PA, Castaman G, Haertel S, Mannucci PM. von Willebrand factor/ factor VIII concentrate (Haemate P) dosing based on pharmacokinetics: a prospective multicenter trial in elective surgery. J Thromb Haemost 2007; 5: 1420–30. Gill JC, Shapiro A, Valentino LA et al. von Willebrand factor/factor VIII concentrate (Humate-P) for management of elective surgery in adults and children with von Willebrand disease. Haemophilia 2011; 17: 895–905. Favaloro EJ, Thom J, Patterson D et al. Desmopressin therapy to assist the functional identification and characterisation of von Willebrand disease: differential utility from combining two (VWF:CB and VWF: RCo) von Willebrand factor activity assays? Thromb Res 2009; 123: 862–8. Van Vliet HH, Kappers-Klunne MC, Leebeek FW, Michiels JJ. PFA-100 monitoring of von Willebrand factor (VWF) responses to desmopressin (DDAVP) and factor VIII/ VWF concentrate substitution in von Willebrand disease type 1 and 2. Thromb Haemost 2008; 100: 462–8. Trossaert M, Ternisien C, Lefrancois A et al. Evaluation of an automated von Willebrand factor activity assay in von Willebrand disease. Clin Appl Thromb Hemost 2011; 17: E25–9. Chen D, Tange JI, Meyers BJ, Pruthi RK, Nichols WL, Heit JA. Validation of an automated latex particle-enhanced immunoturbidimetric von Willebrand factor activity assay. J Thromb Haemost 2011; 9: 1993–2002. Flood V, Cox Gill J, Morateck PA et al. Gain-of-function GPIb ELISA assay for VWF activity in the Zimmerman Program for the Molecular and Clinical Biology of VWD. Blood 2011; 117: 67–74. Cabrera N, Moret A, Caunedo P et al. Comparison of a new chemiluminescent immunoassay for von Willebrand factor activity with the ristocetin cofactor-induced platelet agglutination method. Haemophilia 2013: 19: 920–5.
Haemophilia (2014), 1--7
DOI: 10.1111/hae.12437
ORIGINAL ARTICLE
Evaluation of a rapid von Willebrand Factor Activity Latex Immuno Assay for monitoring of patients with von Willebrand disease (VWD) receiving DDAVPâ or VWF replacement therapy S . V I N A Y A G A M , L . R . S I M O N S , P . C H O W D A R Y , P . T H U R L O W , S . V . B R O O K S and A. F. RIDDELL Katharine Dormandy Haemophilia Centre & Thrombosis Unit, Royal Free London NHS Foundation Trust, London, United Kingdom
Summary. Haemostatic management of surgery in patients with von Willebrand disease (VWD) includes DDAVPâ or von Willebrand factor (VWF)-containing concentrates. Although the recommendations are for monitoring by VWF activity assays, it is quite common for clinicians to use factor VIII due usually to longer turnaround times required for VWF ristocetin cofactor assay (VWF:RCo) measurements. The aim of this study was to evaluate use of the rapid HaemosILTM VWF activity (VWF:Act) latex immuno assay (LIA) on an automated coagulometer (ACL TOPTM 700; Instrumentation Laboratory, Bedford, MA, USA) compared to platelet-based VWF:RCo assays in this setting. One hundred and sixty-seven plasma samples from 42 patients [Type 1 (n = 22), Type 2A (n = 2), Type 2B (n = 3), Type 2M (n = 10), Type 3 (n = 3)] and acquired von Willebrand syndrome (n = 2) with VWD treated with DDAVPâ or VWF-containing concentrates were included in the study. Method comparison and method bias were
Introduction von Willebrand factor (VWF), a plasma glycoprotein synthesized in megakaryocytes and vascular endothelial cells is secreted into plasma and subendothelial compartments in response to vascular injury or stimulation by endogenous or exogenous secretagogues. VWF plays a critical role in primary haemostasis, Correspondence: Anne Riddell, Katharine Dormandy Haemophilia centre and Thrombosis unit, Royal Free London NHS Foundation Trust, Pond Street, London, United Kingdom, NW3 2QG. Tel: +442078302274; fax: +442078302879; e-mail: [email protected] Accepted after revision 11 March 2014 © 2014 John Wiley & Sons Ltd
evaluated by Bland–Altman analysis (BA) and Passing and Bablok regression modelling respectively. BA of baseline samples (n = 39) showed a mean difference of 3.0 (1.96 SD 25.2 to +19.4). Post (treatment) samples (n = 120) were separated into two groups. Group 1 contained samples with VWF:RCo levels 10 to ≤175 IU dL 1 (n = 97) and group 2, samples with VWF:RCo levels >175 IU dL 1 (n = 23). BA of group 1 postsamples showed a mean difference of +3.4 (1.96 SD 44.6 to +51.5), and the BA of Group 2 samples was 23.9 (1.96 SD 136.1 to +88.3). In conclusion, use of HaemosIL VWF:Act LIA test on an automated coagulometer is a reproducible and rapid assay that can be used as an alternative test for monitoring VWF replacement therapy, facilitating dose adjustments on a real-time basis. Keywords: HaemosILTM von Willebrand factor activity, ristocetin cofactor assays, treatment monitoring, von Willebrand disease, von Willebrand factor
serving as a mediator for platelet adhesion and aggregation at sites of vascular injury, as well as acting as a carrier protein to stabilize factor VIII (FVIII) in the circulation [1]. von Willebrand disease (VWD) is a congenital bleeding disorder resulting from a quantitative or qualitative deficiency of VWF function. It is the most common hereditary bleeding disorder, with laboratory abnormalities of VWF present in up to 1% of the population [2]. The quantity of VWF protein present in plasma can be measured immunologically as VWF antigen (VWF:Ag). VWF activity is commonly measured by its ability to facilitate platelet agglutination in the presence of the antibiotic ristocetin, as VWF ristocetin cofactor assay (VWF:RCo). It is also important to measure FVIII activity (FVIII:C) in patients with VWD, as it is a reflection of the ability 1
2
S. VINAYAGAM et al.
of VWF to bind and maintain the level of FVIII in the circulation. Individuals with VWD are at risk for surgical bleeding, both intra-operatively and in the postoperative period. Haemostasis in these patients requires the correction of the defect in VWF-platelet-vessel wall interactions and the deficiency of FVIII. Correction of the defect in VWF function is most important perioperatively and in the immediate postoperative period, particularly when mucosal bleeding is possible. Therapeutic FVIII:C levels also need to be maintained peri- and postoperatively for 7–10 days [3]. Strategies for perioperative management of patients with VWD depend on the type of deficiency, baseline levels of VWF and FVIII:C, the patient’s historical responses to 1-deamino-8-D-arginine vasopressin, (DDAVPâ) and the site and invasiveness of the planned procedure. United Kingdom Haemophilia Centre Doctors’ organization guidelines recommend covering operative procedures with DDAVPâ in responsive patients unless contraindicated. Treatment of surgical bleeding with VWF-containing concentrates is advised for patients with VWD who do not respond adequately to DDAVPâ or in whom use of this agent is contraindicated. It is recommended that the VWF:RCo should be maintained at above 50 IU dL 1 in the perioperative period and FVIII:C maintained at around 100 IU dL 1 during major surgery and above 50 IU dL 1 in the postoperative period, until haemostasis is secure. The guidelines recommend that preferentially VWF:RCo should be monitored, in reality FVIII:C is commonly used, although it is a surrogate marker and does not truly reflect the kinetics of VWF replacement therapy, especially in patients with type 2 and 3 VWD. Whilst the VWF:RCo assay is the standard assay used to evaluate VWF activity, it is labour intensive with a high coefficient of variation [4] and in our laboratory is subject to an urgent turnaround time of at least one hour. Measurement of VWF collagen binding activity (VWF:CB) has been proposed as an alternative method of monitoring treatment in patients with VWD but it is also time consuming, requiring an enzyme linked immunosorbent assay (ELISA) technique. The availability of a rapid method of VWD treatment monitoring in VWD would enable more accurate and responsive prophylaxis of bleeding during major surgery, potentially avoiding excessive administration of concentrates and reducing the risk of perioperative haemorrhage and venous thromboembolic complications. The HaemosILTM VWF activity [Instrumentation Laboratory (IL), Bedford, MA, USA] latex immuno assay (LIA) contains a purified anti mouse monoclonal antibody (RFF VIII:R/1) directed against a functional epitope of VWF–Glycoprotein 1b (platelet binding) receptor site–adsorbed onto latex particles, also containing bovine serum albumin, stabilizers and preservative that interacts with VWF in a plasma Haemophilia (2014), 1--7
sample [5]and from now the test will be referred to as ‘VWF:Act’. The VWF:Act test is independent of ristocetin and can be fully automated and run on a number of different analysers. Previous studies of the assay have satisfactorily evaluated its use in identifying patients with VWD [6–8] but the use of this antibody in an ELISA setting has previously demonstrated limitations in its sensitivity to some qualitative variants of VWF [9]. However, this simple automated latex assay potentially enables laboratories to provide a more rapid evaluation of VWF function and shorter turnaround time for monitoring response to treatment in the perioperative setting. In our laboratory, the VWF:Act test takes 15 min (if test previously calibrated) in comparison to 1 h for a platelet-based VWF:RCo assay result. In this study, we compared the performance of HaemosILTM VWF Activity assay on an ACL TOPTM 700 coagulometer (IL) with VWF:RCo assays using lyophilized platelet reagents in VWD patients receiving VWF-containing concentrates and DDAVPâ for a variety of surgical procedures for accuracy and clinical usefulness.
Material and methods This was a retrospective study of patients with VWD receiving either DDAVPâ or VWF-containing concentrates such as Haemateâ P (CSL Behring, Marburg, Germany), Wilfactinâ (LFB, Les Ulis, France) or Alphanateâ (Grifols, Barcelona, Spain) for a range of reasons including major and minor surgeries, treatment of mucocutaneous bleeding episodes, gastrointestinal bleeding and excessive nose bleeding. Venous whole blood was collected into 0.106 M trisodium citrate tubes (Becton & Dickinson, UK), centrifuged at 2100 g for 12 min, plasma separated into two aliquots, and stored at 40°C until testing. In total 169 plasma samples from 42 patients with VWD with the following subtypes were included: Type 1 (n = 22), Type 2A (n = 2), Type 2B (n = 3), Type 2M (n = 10), Type 3 (n = 3) and acquired von Willebrand syndrome (n = 2). The median age of the patient group was 37 years (range 5–83 years) and included males (n = 21) and females (n = 19). Of the 169 Samples, 46 were baseline samples and 123 samples were taken after treatment at variable time intervals. The details of the treatment material and subtype of VWD are presented in Table 1. VWF:RCo levels were measured using lyophilizedplatelet-based VWF:RCo methods on either a platelet aggregation profiler 4 (PAP-4) aggregometer (Bio-Data, Horsham, PA, USA) or on an ACL9000 (IL). Briefly, diluted patient, standard or quality control (QC) plasma samples were added to either lyophilized platelets (Bio Data) and ristocetin (10 mg mL 1) (Abp, London, UK) and end point measured using the PAP-4 aggregometer or added to lyophilized platelets (BCâ VW reagent, Siemens Healthcare Diagnostics, Marburg, © 2014 John Wiley & Sons Ltd
EVALUATION OF A RAPID VWF ACTIVITY LATEX IMMUNO ASSAY
3
Table 1. Showing number of samples by type of VWD and type of treatment received. Type of VWD Treatment material
Type 1
Type 2A
Type 2B
Type 2M
Type 3
AVWS*
Total number (n) of samples
Alphanateâ Haemateâ P Wilfactinâ DDAVPâ Total (n) samples by VWD subtype
6 1 2 16 25
1 1 – – 2
2 – 1 – 3
3 5 2 1 11
1 2
– 2 – – 2
13 11 5 17 46
– 3
AVWS, acquired von Willebrand syndrome; VWD, von Willebrand disease. *Acquired von Willebrand disease.
Germany) with added ristocetin (50 mg mL 1) based on the method by Lattuada et al. [10] and the end point measured on an ACL9000. Prior to use of ACL9000 VWF:RCo method, a comparison with the PAP-4 VWF: RCo method was carried out. The correlation between VWF:RCo methods was good (r = 0.9562, P = 175 IU dL 1). It would be useful for laboratories to have a rapid assay to monitor VWF function in patients requiring VWF replacement therapy who are undergoing major and minor surgical episodes. Recently, newer types of Haemophilia (2014), 1--7
assays for measuring VWF function using recombinant glycoprotein 1b molecules have been developed, but there is limited literature on their use in monitoring VWF replacement therapy [27,28]. The VWF:Act is an easy to perform assay that is able to produce a result quickly when compared to lyophilized platelet VWF:RCo assays (15 min vs. 1 h respectively) and therefore will help facilitate a rapid turnaround of results in urgent settings especially prior to surgery. Because the VWF:Act test is very reproducible in comparison to the VWF:RCo assay on an automated coagulometer (intra- and inter-assay CVs 3.7% and 4.4% vs. 5.4% and 9.3% respectively) it therefore seems reasonable that this test can be the assay of choice for monitoring VWF replacement therapy, facilitating dose adjustments on a real-time basis allowing for optimization of treatment and preventing under-treatment in the first 48 h with resultant bleeding and over-treatment after 48 h with the attendant risk of thrombosis. Since the initial validation we now routinely use the VWF:Act test for monitoring VWF replacement © 2014 John Wiley & Sons Ltd
EVALUATION OF A RAPID VWF ACTIVITY LATEX IMMUNO ASSAY
therapy with occasional retrospective cross checking with VWF:RCo assay measurements on patient samples (especially when they first receive treatment and their response is not known). This helps to facilitate shorter turnaround times for monitoring patients receiving VWF treatments (DDAVPâ and VWF-containing concentrates) that are within acceptable clinical time frames. In conclusion, the HaemosILTM VWF: Activity test is a rapid and reliable automated assay that can be used for monitoring VWF functional activity of VWF-containing concentrates or DDAVPâ therapy in patients with VWD.
References 1 Wagner DD, Bonfanti R. von Willebrand factor and the endothelium. Mayo Clin Proc 1991; 66: 621–7. 2 Rodeghiero F, Castaman G, Dini E. Epidemiological investigation of the prevalance of von Willebrand’s disease. Blood 1987; 69: 454–9. 3 Pasi KJ, Collins PW, Keeling DM et al. Management of von Willebrand disease: a guideline from the UK Haemophilia Centre Doctors’ Organization. Haemophilia 2004; 10: 218–31. 4 Kitchen S, Jennings I, Woods TA, Kitchen DP, Walker ID, Preston FE. Laboratory tests for measurement of von Willebrand factor show poor agreement among different centers: results from the United Kingdom National External Quality Assessment Scheme for Blood Coagulation. Semin Thromb Hemost 2006; 32: 492–8. 5 De Vleeschauwer A, Devreese K. Comparison of a new automated von Willebrand factor activity assay with an aggregation von Willebrand ristocetin cofactor activity assay for the diagnosis of von Willebrand disease. Blood Coagul Fibrinolysis 2006; 17: 353–8. 6 Sucker C, Senft B, Scharf RE, Zotz RB. Determination of von Willebrand factor activity: evaluation of the HaemosIL assay in comparison with established procedures. Clin Appl Thromb Hemost 2006; 12: 305–10. 7 Trossaert M, Ternisien C, Lefrancois A et al. Evaluation of an automated von Willebrand factor activity assay in von Willebrand disease. Clin Appl Thromb Hemost 2011; 6: 25–9. 8 Pinol M, Sales M, Costa M, Tosetto A, Canciani MT, Federici AB. Evaluation of a new turbidimetric assay for von Willebrand factor activity useful in the general screening of von Willebrand disease. Haematologica 2007; 92: 712–3. 9 Nitu-Whalley IC, Riddell A, Lee CA et al. Identification of type 2 von Willebrand disease in previously diagnosed type 1 patients: a reappraisal using phenotypes, genotypes and molecular modelling. Thromb Haemost 2000; 84: 998–1004. 10 Lattuada A, Preda L, Sacchi E, Gallo L, Federici AB, Rossi E. A rapid assay for
© 2014 John Wiley & Sons Ltd
11
12
13
14
15
16
17
18
19
20
7
Author contributions SV and SB performed the assays and wrote first draft of the manuscript. LS and PC reviewed and wrote the manuscript. PT collected the patient samples and helped to collate treatment information. AR designed the study, analysed the data, and reviewed the manuscript. Thanks to Vincent P Jenkins for helpful discussion on the manuscript.
Disclosures The authors stated that they had no interests which might be perceived as posing a conflict or bias.
ristocetin cofactor activity using an automated coagulometer (ACL 9000). Blood Coagul Fibrinolysis 2004; 15: 505–11. Riddell AF, Jenkins PV, Nitu-Whalley IC, McCraw AH, Lee CA, Brown SA. Use of the collagen-binding assay for von Willebrand factor in the analysis of type 2M von Willebrand disease: a comparison with the ristocetin cofactor assay. Br J Haematol 2002; 116: 187–92. Bland JM, Altman DG. Measuring agreement in method comparison studies. Stat Methods Med Res 1999; 8: 135–60. Passing H, Bablok W. A new biometrical procedure for testing the equality of measurements from two different analytical methods. Application of linear regression procedures for method comparison studies in clinical chemistry, Part I. J Clin Chem Clin Biochem 1983; 21: 709–20. Mannucci PM. Treatment of von Willebrand’s disease. N Engl J Med 2004; 351: 683–94. Batlle J, Lopez-Fernandez MF, Fraga EL, Trillo AR, Perez-Rodriguez MA. Von Willebrand factor/factor VIII concentrates in the treatment of von Willebrand disease. Blood Coagul Fibrinolysis 2009; 20: 89– 100. Berntorp E, Archey W, Auerswald G et al. A systematic overview of the first pasteurised VWF/FVIII medicinal product, Haemate P/Humate-P: history and clinical performance. Eur J Haematol 2008; 80 (Suppl. 70): 3–35. Franchini M. Surgical prophylaxis in von Willebrand’s disease: a difficult balance to manage. Blood Transfus 2008; 6(Suppl. 2): 33–8. Michiels JJ, van Vliet HH, Berneman Z, Schroyens W, Gadisseur A. Managing patients with von Willebrand disease type 1, 2 and 3 with desmopressin and von Willebrand factor-factor VIII concentrate in surgical settings. Acta Haematol 2009; 121: 167–76. Hay CR, Hellmann A, Dmoszynska A et al. Experience with Optivate(R), a new high purity concentrate of factor VIII with von Willebrand factor, in patients undergoing surgery. Haemophilia 2011; 17: 428–32. Windyga J, Von Depka-Prondzinski M. Efficacy and safety of a new generation von
21
22
23
24
25
26
27
28
Willebrand factor/factor VIII concentrate (Wilate(R)) in the management of perioperative haemostasis in von Willebrand disease patients undergoing surgery. Thromb Haemost 2011; 105: 1072–9. Lethagen S, Kyrle PA, Castaman G, Haertel S, Mannucci PM. von Willebrand factor/ factor VIII concentrate (Haemate P) dosing based on pharmacokinetics: a prospective multicenter trial in elective surgery. J Thromb Haemost 2007; 5: 1420–30. Gill JC, Shapiro A, Valentino LA et al. von Willebrand factor/factor VIII concentrate (Humate-P) for management of elective surgery in adults and children with von Willebrand disease. Haemophilia 2011; 17: 895–905. Favaloro EJ, Thom J, Patterson D et al. Desmopressin therapy to assist the functional identification and characterisation of von Willebrand disease: differential utility from combining two (VWF:CB and VWF: RCo) von Willebrand factor activity assays? Thromb Res 2009; 123: 862–8. Van Vliet HH, Kappers-Klunne MC, Leebeek FW, Michiels JJ. PFA-100 monitoring of von Willebrand factor (VWF) responses to desmopressin (DDAVP) and factor VIII/ VWF concentrate substitution in von Willebrand disease type 1 and 2. Thromb Haemost 2008; 100: 462–8. Trossaert M, Ternisien C, Lefrancois A et al. Evaluation of an automated von Willebrand factor activity assay in von Willebrand disease. Clin Appl Thromb Hemost 2011; 17: E25–9. Chen D, Tange JI, Meyers BJ, Pruthi RK, Nichols WL, Heit JA. Validation of an automated latex particle-enhanced immunoturbidimetric von Willebrand factor activity assay. J Thromb Haemost 2011; 9: 1993–2002. Flood V, Cox Gill J, Morateck PA et al. Gain-of-function GPIb ELISA assay for VWF activity in the Zimmerman Program for the Molecular and Clinical Biology of VWD. Blood 2011; 117: 67–74. Cabrera N, Moret A, Caunedo P et al. Comparison of a new chemiluminescent immunoassay for von Willebrand factor activity with the ristocetin cofactor-induced platelet agglutination method. Haemophilia 2013: 19: 920–5.
Haemophilia (2014), 1--7
