RESUMO
BACKGROUND: The antiphospholipid syndrome (APS) is an autoimmune condition characterised by vascular thromboses and/or pregnancy morbidity. Diagnosis of APS typically requires laboratory evidence of antiphospholipid antibodies (aPL). Depending on their clinical presentation, affected individuals might be seen by a variety of clinical specialities. AIM: To evaluate clinical ordering patterns for aPL/APS at a tertiary level public facility. METHODS: We performed an audit of internal clinical requests for aPL tests at our institution for a 6-month period. RESULTS: We identified a wide variety of clinical ordering background for aPL, of predominantly obstetric (72/268; 26.9%) or thrombophilic (78/268; 29.1%) patients. Only 11/268 samples (4.1%) were positive for lupus anticoagulant (LA) and 14/268 (5.2%) were positive for anticardiolipin antibody (aCL). The percentage of aCL positivity in the LA-positive group was 46% (5/11). None of the 72 obstetric patients tested was identified to have aPL. Of the 11 LA-positive patients, the reasons identified for testing comprised: prolonged Activated Partial Thromboplastin Time (assay) (n= 3), thrombosis (n= 3), APS (n= 2), systemic lupus erythematosus (n= 2), vasculitis (n= 1). CONCLUSION: We determined a wide variety of clinical ordering background for aPL at a tertiary level institution, with an overall low rate (<10%) of aPL positivity among a hospital population of predominantly obstetric or thrombophilic patients. That no positive obstetric aPL cases were identified suggests local clinical ordering guidelines may need review, as also potentially practised at other institutions. We also observed a moderate rate (46%) of coincidence of aCL and LA, in agreement with guidelines indicating that multiple tests are required to identify APS.
Assuntos
Anticorpos Anticardiolipina/sangue , Anticorpos Antifosfolipídeos/sangue , Síndrome Antifosfolipídica/diagnóstico , Trombofilia/diagnóstico , Síndrome Antifosfolipídica/imunologia , Auditoria Clínica , Feminino , Humanos , Masculino , Gravidez , Estudos Retrospectivos , Trombofilia/imunologiaRESUMO
Platelet Factor VIII-related antigen (VIIIR:Ag) represents a significant proportion of the total circulating VIIIR:Ag pool. However, its participation in the events of primary hemostasis has not been shown. We now report that platelet-contained VIIIR:Ag is released from platelets by collagen, ADP and thrombin. The concentrations of these agonists, required for VIIIR:Ag release, are the same or lower than those required for release of serotonin, lysosomal enzymes, or fibrinogen. This release has the features of an energy-dependent secretory response because it is blocked by the metabolic inhibitors, antimycin A and 2-deoxy-D-glucose. The electrophoretic characteristics of the VIIIR:Ag released by collagen and ADP are similar to those of plasma VIIIR:Ag. However, thrombin-released platelet VIIIR:Ag differs from that of plasma in that the less anodal forms are relatively depleted. These differences do not appear to be the result of proteolytic degradation of platelet-derived VIIIR:Ag, but may reflect interactions between specific molecular forms of VIIIR:Ag and the platelet membrane. These studies suggest mechanisms by which platelet-contained VIIIR:Ag may contribute to the primary events of hemostasis.
Assuntos
Difosfato de Adenosina/farmacologia , Plaquetas/metabolismo , Colágeno/farmacologia , Fator VIII/metabolismo , Trombina/farmacologia , Antígenos/análise , Antígenos/imunologia , Autorradiografia , Plaquetas/imunologia , Fator VIII/imunologia , Fibrinogênio/metabolismo , Hemostasia , Humanos , Imunoeletroforese , Fatores de Tempo , Doenças de von Willebrand/sangue , Doenças de von Willebrand/metabolismoRESUMO
Phospholipid procoagulant material mainly derived from platelets interferes or "bypasses" the more specific tests for lupus anticoagulants. Such material in test plasmas can be inactivated with recovery of lupus anticoagulant activity by simple extraction with chloroform. This solvent treatment damages mainly factors VIII, V, VII and IX. Ether and various other solvents were less damaging to these clotting factors but not quite as effective as chloroform in the recovery of lupus anticoagulant activity.
Assuntos
Autoanticorpos/isolamento & purificação , Fatores de Coagulação Sanguínea/imunologia , Plasma/imunologia , Solventes , Fatores de Coagulação Sanguínea/isolamento & purificação , Humanos , Inibidor de Coagulação do Lúpus , Valores de Referência , Solventes/efeitos adversosRESUMO
We report an evaluation of current laboratory practice for the diagnosis of von Willebrand's disease (VWD) by means of a multilaboratory survey. This assessment was undertaken with the RCPA Quality Assurance Program (QAP) in Haematology, which covers a wide geographic area encompassing Australia, New Zealand and Asia. A total of 25 laboratories actively involved in testing for VWD were selected to participate in a sample testing assessment exercise. Samples comprised 10 plasmas: (i) a normal plasma pool (in duplicate), (ii) this pool diluted to 50% (in duplicate), (iii) a normal individual (X1), (iv) severe Type 1 VWD (X1), (v) Type 2B VWD (x2 unrelated donors), (vi) Type 3 VWD (x1), (vii) Type 2A VWD (x1). Laboratories were asked to perform all tests available to them in order to establish a laboratory diagnosis of VWD, and then to comment on the possibility or otherwise of VWD. Overall findings indicated a wide variation in test practice, in the effectiveness of various test procedures in detecting VWD, and in the ability of various composite test panels to identify type 2 VWD subtypes. Firstly, while all laboratories (n = 25) performed tests for FVIII:C activity, von Willebrand factor 'antigen' (VWF:Ag) and a functional VWF assay [using the ristocetin cofactor assay (VWF:RCo; n = 23) and/or the collagen binding assay (VWF:CBA; n = 12)], only three laboratories carried out VWF:Multimer analysis. Secondly, for the three quantitative VWF assays, 10/25 (40%) laboratories performed all three, whereas 15/25 (60%) performed only two [VWF:Ag and VWF:RCo (n = 13); VWF:Ag and VWF:CBA (n = 2)]. Thirdly, a variety of assay methodologies were evident for VWF:Ag [ELISA, electro-immuno diffusion (EID), latex immuno-assay (LIA), and VIDAS assay] and VWF:RCo (platelet agglutination/'aggregometry' and a 'functional VWF:RCo-alternative' ELISA assay). Between method analysis for the quantitative VWF assays showed that the VWF:RCo yielded the greatest degree of inter-laboratory assay variation, and had the poorest overall performance with respect to sensitivity to low levels of VWF. The VWF:CBA also performed better than the VWF:RCo in terms of ability to detect functional VWF 'discordance' (i.e. Type 2 VWD). Within VWF:Ag method analysis showed that the EID assay procedure was associated with the greatest variation in assay results, while the EID and LIA test methods showed poorer sensitivity at low VWF levels compared to the ELISA method. Within the VWF:RCo assay procedure, greatest variation in assay results and poorest sensitivity to low VWF levels was obtained using the agglutination method; however, the agglutination procedure showed better performance than the 'functional VWF:RCo-alternative' ELISA assay in identifying Type 2 VWD plasma samples. Finally, despite identified variations, most laboratories appeared to understand the complexities involved in the VWD-diagnostic process, and made appropriate diagnostic predictions regarding tested samples. From a total possible 246 interpretation events, laboratories in most cases correctly identified normal samples as normal (67/75 events = 89%), and VWD samples as derived from individuals with VWD (117/121 events = 97%). Moreover, when VWD was suggested by laboratory findings, laboratories usually correctly predicted the general subtype of VWD present (96/109 events = 88%). When 'misinterpretations' occurred, these could often be linked to the test panels utilised by laboratories. That is, laboratories using the VWF:Ag and VWF:RCo combination were more likely to incorrectly identify samples derived from Type 2 VWD patients as being Type 1, Type 1 VWD patients as being Type 2, and normal plasma samples as potentially derived from patients with VWD, compared to those using the VWF:Ag and VWF:CBA.
Assuntos
Técnicas de Laboratório Clínico/normas , Doenças de von Willebrand/diagnóstico , Estudos de Avaliação como Assunto , Inquéritos Epidemiológicos , Humanos , Padrões de ReferênciaRESUMO
Human endothelial cells, cultured from umbilical cord veins, have been evaluated for expression of a large number of cell surface antigens with known haemopoietic, particularly myeloid, cell distribution. This was achieved by evaluating endothelial reactivity (using non-fixed cells) with groups of monoclonal antibodies (MAB) belonging to distinct Clusters of Differentiation (CD), as defined by the Third International Workshop on Leukocyte Differentiation Antigens (ILWS). Results indicate that many antigens known to be present on haemopoietic cells, including those on platelets, are co-expressed on endothelial cells. The most intense degree of reactivity was seen using MAB to CD-9 and CD-13, although significant reactivity was also observed using MAB to CD-31 and CD-32. Data also suggests weak binding to endothelial cells of MAB belonging to CD-14, CD-15 and CD-16. A number of unclustered MAB reactive with haemopoietic antigens can also be shown to bind to endothelial cells.
Assuntos
Antígenos de Superfície/análise , Endotélio Vascular/imunologia , Células-Tronco Hematopoéticas/imunologia , Anticorpos Monoclonais , Endotélio Vascular/citologia , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , HumanosRESUMO
In a cross-sectional analytic study, we examined the differences in coronary heart disease (CHD) risk factors, including coagulation factors and platelet aggregation, among males from southern European countries and those of Anglo-Celtic descent who had widely different CHD standardized mortality ratios. The participants included 169 men aged 40 to 49 years, 27% of whom were born in southern European countries. The subjects had no history of heart disease and no other clinical conditions, or were not taking medications known to affect hemostasis. Data obtained included their medical history and CHD-related risk behaviors, blood pressure, height, weight, abdominal and pelvic circumference, and coagulation, fibrinolysis, platelet activity, lipids, and lipoproteins profiles. There were significant differences between the two groups in the prevalence of a positive family history, mean apolipoprotein A1 levels, and platelet aggregation responses to ADP. Other established risk factors, including coagulation factor levels, were not significantly different.
Assuntos
Doença das Coronárias/mortalidade , Agregação Plaquetária , Difosfato de Adenosina/farmacologia , Adulto , Austrália/epidemiologia , Coagulação Sanguínea , Colágeno/farmacologia , Doença das Coronárias/sangue , Doença das Coronárias/etnologia , Estudos Transversais , Europa (Continente)/etnologia , Humanos , Lipídeos/sangue , Lipoproteínas/sangue , Masculino , Pessoa de Meia-Idade , Agregação Plaquetária/efeitos dos fármacos , Fatores de Risco , Reino Unido/etnologiaRESUMO
Aging of the Australian population, as in other developed nations, will ensure that stroke remains one of the most important causes of death and disability. The Stroke Risk in the Elderly (SITE) study aims to measure prospectively the independent contribution of dietary, sociodemographic, blood lipid, blood pressure, and hemostatic factors to risk of stroke and other cardiovascular outcomes. The target population included all independently living men and women aged 65 years and over, residents in several retirement villages in western metropolitan Sydney, New South Wales, Australia. The study cohort consists of 225 men and 787 women, selected as a convenience sample from all eligible residents in the local government areas (LGAs) adjacent to Westmead Hospital. Participants attended a baseline session to complete dietary, life-style, medical, and sociodemographic questionnaires. Anthropomorphic variables and blood pressure were measured. Blood was taken for measurement of serum lipid, glucose, and hemostatic factors. Questionnaire results were compared with an age/sex-stratified, randomly selected sample drawn from the community (in the same LGAs), in order to quantify potential sampling and selection biases. The study cohort will be followed for a minimum of 5 years. The attendance rate of eligible residents for a baseline medical, dietary, life-style, and sociodemographic assessment was 72% for males and 69% for females. The study cohort was older, better educated, less ethnically diverse, and among women, less likely to have ever been married compared to people aged over 65 years in the comparison group. The baseline results suggest that hemostatic factors may be of importance in assessing risk of cardiovascular disease, (CVD), particularly in older men.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Transtornos Cerebrovasculares/epidemiologia , Vigilância da População , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Causas de Morte , Inquéritos sobre Dietas , Pessoas com Deficiência , Feminino , Humanos , Hiperlipidemias/sangue , Hiperlipidemias/complicações , Hipertensão/complicações , Estilo de Vida , Masculino , New South Wales/epidemiologia , Estudos Prospectivos , Fatores de Risco , Viés de Seleção , Fatores SocioeconômicosRESUMO
We have investigated the compartmentalization of aminopeptidase-N-like activity in various blood fractions obtained from patients with acute lymphoid (ALL) or myeloid (AML) leukemia. The primary difference appears not to be the absolute level of overall activity, but rather the relative proportions of the different forms of activity detected. Thus, despite similar levels of total aminopeptidase-N-like activity detected in cells from different leukemic groups, true aminopeptidase-N/CD13 activity was only detected in cells derived from AML patients. Even in these patients, however, most of the detected aminopeptidase-N-like activity ( > 80%) could not be attributed to aminopeptidase-N/CD13. In marked contrast, plasma from leukemic patients also contained substantial total aminopeptidase-N-like activity, of which (irrespective of leukemic group) most could be attributed to aminopeptidase-N/CD13. Whilst slightly higher levels of total activity were obtained in plasma from AML patients compared to ALL patients, there was no difference in the relative proportion attributable to aminopeptidase-N/CD13 (approximately 80% of total aminopeptidase-N-like activity). Evaluation of total aminopeptidase-N-like activity present in whole blood gave differential patterns, and whilst only a proportion (20-40% of total aminopeptidase-N-like activity) could be attributed to true aminopeptidase-N/CD13, blood from patients with CD13+ AML showed the greatest activity so attributable. In total, our results outline the complexities of peptidase activities present within blood of leukemic individuals, and may, in part, explain the variability of previous studies attempting to associate prognostic features with phenotypic expression of CD13.
Assuntos
Antígenos CD13/sangue , Leucemia Mieloide Aguda/enzimologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/enzimologia , Anticorpos Bloqueadores/imunologia , Anticorpos Monoclonais/imunologia , Humanos , Leucemia Mieloide Aguda/sangue , Leucemia Mieloide Aguda/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/sangue , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologiaRESUMO
A case of thrombotic thrombocytopenic purpura (TTP) is reported in a 40-year-old man 6 months after allogeneic bone marrow transplantation for multiple myeloma. The features of TTP included microangiopathic haemolytic anaemia, severe thrombocytopenia, fluctuating neurological abnormalities, and progressive renal impairment. Despite treatment with anti-platelet agents, prostacyclin infusion, intensive immunosuppression and prolonged plasma exchange, the patient developed end-stage renal failure and is now on maintenance haemodialysis 18 months after the onset of TTP. Graft-versus-host disease and cytomegalovirus infection could not be implicated as aetiological factors, and cyclosporin medication had ceased 1 week before the clinical onset of his disease. The unusually intensive pre-transplant chemotherapy and radiotherapy protocol used in this patient appear to be most likely cause of the generalized endothelial damage resulting in TTP in this patient.
Assuntos
Transplante de Medula Óssea/efeitos adversos , Púrpura Trombocitopênica Trombótica/etiologia , Adulto , Humanos , Falência Renal Crônica/etiologia , Masculino , Mieloma Múltiplo/cirurgia , Transplante HomólogoRESUMO
We present a 42-year-old man with acute lymphoblastic leukemia and hypodiploidy at diagnosis. Chromosome count was 37, with a mixture of numerical and structural abnormalities. The patient died 9 months post diagnosis, during which time three further cytogenetic tests were performed. The core abnormalities seen upon diagnosis were present at 7 and 9 months after diagnosis, with a duplication of the abnormal hypodiploid karyotype on the last specimen. While considerable imbalances were present as a result of whole chromosome aneuploidy, no region was obviously nullisomic.
Assuntos
Aneuploidia , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Adulto , Células da Medula Óssea/citologia , Humanos , Hibridização in Situ Fluorescente , MasculinoRESUMO
Fifty-four adolescent and adult patients with newly-diagnosed acute lymphoblastic leukaemia (ALL) were treated with combination chemotherapy at three Australian hospitals. The protocol consisted of one month of induction therapy with five cytotoxic agents, followed by consolidation therapy and prophylactic treatment to the central nervous system, then maintenance chemotherapy for 30 months on an outpatient basis. Complete remission was achieved in 47 (87%) patients, with 5 deaths due to treatment-related toxicity. Two patients had drug-resistant disease. Twenty-two patients subsequently relapsed in the bone marrow (18) or in the central nervous system (4). The median survival for all 54 patients is 45.6 months, while the median duration of remission for the 47 complete responders is 39.0 months, with 38.1% projected to be disease-free at 5 years. Age at diagnosis was found to be the only parameter at presentation with a significant predictive effect on outcome. Patients between 10 and 20 years of age had a median survival of 120.6 months, with the median duration of remission not yet reached. In contrast, patients aged 20 years or more had a significantly poorer outcome, with median survival and remission of 25.8 and 20.8 months respectively. These results would support the use of intensive chemotherapy for adolescent patients with ALL. The poor results in adults however justify the use of alternative approaches such as bone marrow transplantation.
RESUMO
Activation of washed platelets in the presence of EDTA with either 1 U/ml of alpha-thrombin or 2 microM calcium ionophore (A23187) caused the release of one-third to one-half of the platelet factor VIII/von Willebrand factor (FVIII/vWF) into the supernatant. When calcium was present in excess, only 10% of the platelet FVIII/vWF was detected free in the supernatant, regardless of whether calcium was present before stimulation or added to the platelets after thrombin activation. Release of [14C]serotonin and beta-thromboglobulin were not affected by divalent cations indicating that reduced supernatant levels of FVIII/vWF in the presence of calcium were not due to differential release, but were probably due to a calcium-dependent association of released FVIII/vWF with the platelet surface. The presence or absence of intact glycoprotein Ib on the platelet surface made no significant difference to the observed FVIII/vWF partition. Platelets from a patient with Glanzmann's thrombasthenia, however, failed to show a calcium effect with respect to released FVIII/vWF. The combined results suggest that as well as the ristocetin-dependent, divalent cation-independent binding of FVIII/vWF to glycoprotein Ib, there is a divalent cation-dependent binding of FVIII/vWF to the activated platelet surface which is mediated via the glycoprotein IIb/IIIa complex.
Assuntos
Fatores de Coagulação Sanguínea/metabolismo , Plaquetas/metabolismo , Cálcio/farmacologia , Fator VIII/metabolismo , Fator de von Willebrand/metabolismo , Plaquetas/efeitos dos fármacos , Calcimicina/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Glicoproteínas/sangue , Humanos , Técnicas In Vitro , Trombina/farmacologiaRESUMO
PURPOSE: To determine whether associations exist between cataract and established cardiovascular risk factors (other than smoking) - hypertension, body mass index, serum lipids and plasma fibrinogen. METHODS: The Blue Mountains Eye Study is a large (n=3654) population-based cross-sectional study conducted among people aged 49-97 years residing in the Blue Mountains, a region west of Sydney, Australia. Risk factor data were collected using standardised clinical procedures. Lens photographs were taken and graded for presence and severity of cortical, nuclear, and posterior subcapsular cataracts. RESULTS: Cortical cataract was associated with a history of myocardial infarction, higher plasma fibrinogen, and higher serum cholesterol. Nuclear cataract was associated with a higher platelet count but hypertension was associated with lower prevalence of nuclear cataract. Posterior subcapsular cataract was associated with higher plasma fibrinogen and lower body mass index. Some of these associations appeared to be stronger in women than in men: fibrinogen and cortical cataract and body mass index and posterior subcapsular cataract. CONCLUSIONS: Several risk factors for cardiovascular disease are associated with presence of cataract, perhaps explaining the observation in several studies that people with cataract have increased mortality rates. The possibility of strong associations between plasma fibrinogen and cataract merits further epidemiological and laboratory research.
Assuntos
Doenças Cardiovasculares/complicações , Catarata/complicações , Fibrinogênio/metabolismo , Idoso , Idoso de 80 Anos ou mais , Índice de Massa Corporal , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/epidemiologia , Catarata/sangue , Catarata/epidemiologia , Estudos Transversais , Feminino , Humanos , Incidência , Lipídeos/sangue , Masculino , Pessoa de Meia-Idade , New South Wales/epidemiologia , Razão de Chances , Estudos Retrospectivos , Fatores de Risco , Fatores Sexuais , Inquéritos e Questionários , Taxa de SobrevidaRESUMO
The key to an adequate assessment of the hemostatic status of an individual patient is a full and adequate medical history and proper physical examination, and no set of screening tests can replace this. The vast majority of patients thus assessed will require no tests and the coagulation laboratory staff can concentrate on the full assessment of the small number of patients who will be identified as being at risk. The assessment of disturbances of hemostasis is a logical, and basically quite simple, sequence of tests.
Assuntos
Transtornos da Coagulação Sanguínea/diagnóstico , Tempo de Sangramento , Contagem de Células Sanguíneas , Fatores de Coagulação Sanguínea/análise , Fatores de Coagulação Sanguínea/uso terapêutico , Testes de Coagulação Sanguínea , Hemorragia/etiologia , Hemostasia , Humanos , Complicações Intraoperatórias , Anamnese , Exame Físico , Complicações Pós-Operatórias , Cuidados Pré-Operatórios , Tempo de Protrombina , Tempo de Trombina , Trombocitopenia/complicaçõesRESUMO
Appropriate investigation of a patient with suspected von Willebrand's disease (VWD) involves a clinical assessment of the patient followed by laboratory testing. A variety of laboratory assays may be performed, not necessarily restricted to an assessment of von Willebrand factor (VWF). Due to the limitations of each assay, and because of VWD heterogeneity, no single test procedure is sufficiently robust to permit detection of all VWD variants. Indeed, these factors often lead to considerable confusion in the process of laboratory interpretation regarding the likelihood of VWD, and the subtype of VWD. This paper attempts to clarify some of the issues that lead to this confusion. It analyses the relative contribution of four separate assays for VWF [VWF:Multimer analysis, VWF:Antigen (VWF:Ag), VWF:Ristocetin Cofactor (VWF:RCof), and VWF:Collagen Binding Activity assay (VWF:CBA)] to the diagnosis and classification of VWD. Although each assay detects VWF, it is important to recognise that each assay provides different information on the VWF so detected. For example, while the VWF:Ag assay is a quantitative assay and provides a very good measure of the overall level of VWF present in a patient's plasma, it is not a functional assay and yields no information concerning the quality of the VWF present. Thus, the VWF:Ag on its own will not permit detection of many qualitative defects (consequently, use of this assay alone will lead to many Type 2 VWD patients being missed by the laboratory). In contrast, the VWF:CBA is a functional assay which provides very useful information on the quality of VWF present. Although the VWF:CBA is also a quantitative assay, it is less sensitive than the VWF:Ag assay in terms of its ability to measure the overall level of VWF (i.e.; the VWF:CBA detects only highly adhesive VWF, and therefore only a proportion of overall VWF). In essence, the VWF:Ag and VWF:CBA assays are complementary assays and should be used in combination. The VWF:Multimer assay is a qualitative procedure, but at best is only semi-quantitative. The VWF:Multimer assay essentially provides a snap-shot of the VWF present. Unfortunately considerable technical and interpretive problems limits its overall applicability and usefulness. The VWF:RCof assay is both a quantitative and qualitative assay that provides information about the presence of VWF that lies between that provided individually by the VWF:Ag and VWF:CBA assays. Unfortunately, the VWF:RCof suffers considerable technical problems, including considerable assay variability, that also limits its overall usefulness. Laboratories performing assays for VWF need to develop diagnostic strategies which include the use of appropriate multiple test combinations so as to ensure that VWD is properly detected.
Assuntos
Técnicas de Laboratório Clínico , Doenças de von Willebrand/diagnóstico , Fator de von Willebrand/análise , Colágeno/metabolismo , Humanos , Agregação Plaquetária , Fator de von Willebrand/imunologia , Fator de von Willebrand/metabolismoRESUMO
This report details extensive studies investigating an improved screening procedure for the laboratory confirmation of clinically suspected von Willebrand's disease (vWD). Over the past two years, over 400 plasma samples, comprising samples derived from both normal individuals (n = 112) and from patients undergoing investigation on clinical grounds, underwent analysis in this screening procedure, comprising three distinct assays: a standard ELISA assay for von Willebrand Factor (vWF) antigen levels (vWF:Ag), a standard ristocetin cofactor (R Cof) assay, and a functional collagen based ELISA assay for vWF ('CBA'). Normal individual plasma samples yielded normal reference values (mean +/- 2SD) approximating 50-200% (vWF:Ag, R Cof) or 50-250% (CBA). In order to permit comparative analysis, and based upon derived assay values, and subsequent multimer analysis, patient samples were either deemed to derive from persons unlikely to suffer vWD ('non-vWD' patient group) or those potentially suffering vWD. The latter group was further separated into subgroups based upon the likelihood, and probable subtype of vWD. In conjunction with the vWF:Ag assay, the CBA provides the basis by which an effective predictor system (likelihood and probable subtype of vWD) can be offered on the basis of preliminary screening procedures. To date, there has been no overlap in vWF:Ag to CBA ratios (vWF:CBA) between patients yielding Type II vWD like multimer patterns and those yielding Type I vWD, or normal, multimer patterns. Thus, high vWF:CBA (i.e. > or = 3.0) would suggest a Type II, or pseudo, -vWD like defect, whereas low vWF:CBA (< or = 2.5) would likely derive from either normal individuals, or persons suffering from Type I vWD.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Ensaio de Imunoadsorção Enzimática/métodos , Doenças de von Willebrand/imunologia , Fator de von Willebrand/análise , Humanos , Doenças de von Willebrand/sangueRESUMO
We report here on the usefulness of the 14C-serotonin release assay for the laboratory confirmation of the clinical diagnosis of heparin-induced thrombocytopenia syndrome (HITS). Over the past 3 yrs, some 140 individual serum samples have been tested in our laboratory for heparin-associated anti-platelet activity ('heparin antibodies'). These included sera from 54 selected (4 positive, 50 negative) controls and a group of 86 patients where the test was requested on clinical grounds. Of 20 patients derived from within our institution, 7 out of 8 patients (88%) with good clinical probability of HITS were confirmed to have heparin platelet antibodies by the serotonin release assay. In contrast, only 2 out of 9 patients (22%) with a low clinical probability of HITS were shown to be positive by this procedure, as were 2 out of 3 patients (66%) deemed to have an 'intermediate' clinical probability of HITS. In addition, screening of 50 serum samples forming a 'negative-control non-HITS' group (either patients on heparin therapy without thrombocytopenia, patients with non-heparin associated thrombocytopenia [eg. ITP*, other drug related], or normal laboratory volunteers), consistently failed to display heparin associated anti-platelet activity by the 14C-serotonin release assay. In addition to the good specificity and sensitivity described above, the 14C-serotonin release assay was found to be nearly twice as sensitive when compared to the platelet aggregation procedure, and it is therefore a useful diagnostic test for the confirmation of clinically suspected HITS.
Assuntos
Heparina/efeitos adversos , Trombocitopenia/diagnóstico , Plaquetas/metabolismo , Radioisótopos de Carbono , Humanos , Agregação Plaquetária , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Serotonina , Trombocitopenia/sangue , Trombocitopenia/induzido quimicamenteRESUMO
A simplified dilute Russell's viper venom time (DRVVT) test--in which the venom, trace phospholipid and calcium were combined into a single reagent--was evaluated for the detection of lupus anticoagulants (LA) in 28 plasma samples containing non-specific circulating anticoagulants. In agreement with previous studies, the DRVVT was found to be insensitive to defects in contact and haemophilic factors and was only marginally affected by antibodies directed against factor VIII. Thus, the use of a DRVVT test in investigations of anticoagulants reduces the risk of confusing a haemorrhagic inhibitor of factor VIII with a non-haemorrhagic LA. In comparisons of sensitivity against activated partial thromboplastin time tests (APTT-Actin FSL and Organon-Teknika reagents) the simplified DRVVT was prolonged slightly more than the APTT in most of the test plasmas containing various non-specific circulating anticoagulants. Three anticoagulants affecting APTTs more than the DRVVT were found to be associated with anticardiolipin IgMs. APTT-prolonging anticoagulants, whether prolonging DRVVT tests or not, showed similar 'correction' of their APTTs by the addition of platelets or phospholipid. Thus, phospholipid-dependent or LA show heterogeneity. Those affecting only the APTT and not DRVVT should perhaps be classified differently.
Assuntos
Fatores de Coagulação Sanguínea/imunologia , Tempo de Protrombina , Adulto , Idoso , Transtornos da Coagulação Sanguínea/imunologia , Fatores de Coagulação Sanguínea/análise , Fator VIII/antagonistas & inibidores , Feminino , Humanos , Inibidor de Coagulação do Lúpus , Lúpus Eritematoso Sistêmico/imunologia , Masculino , Pessoa de Meia-Idade , Tempo de Tromboplastina ParcialRESUMO
We have developed and evaluated an ELISA-based collagen binding assay (CBA) as an aid in the diagnosis and classification of von Willebrand's disease (vWD). The assay is simple to perform, and appears capable of differentiating Type II vWD from Type I vWD. Using plasma samples from both affected and non-affected patients, or from normal individuals, data obtained using the CBA were directly compared to data simultaneously derived from a standard von Willebrand factor antigen (protein; vWFAg) ELISA, and from a standard ristocetin cofactor (RCof) assay. Plasma derived from vWD patients (both Type I and Type II) showed overall reduced levels of vWF as detected by all three assays. Mean levels as a per cent of normal for vWFAg, CBA, RCof were 47.3, 60.7, 31.1 for Type I patients (n = 37), and 34.9, 1.6, 11.9 for Type II patients (n = 16) respectively. However despite the reduced vWF levels detected in Type I vWF binding values for both the CBA and vWFAg showed near comparability (i.e. vWFAg:CBA ratio generally less than or equal to 1.0). These ratio values were thus similar to those observed using plasma derived from either individual normal donors, or from non-vWD affected patients. On the other hand, plasma from Type II vWD affected patients showed markedly disparent values, with increased (greater than 8.0) vWFAg:CBA ratios coincident with virtually absent CBA binding in these patients. Thus, the CBA as reported here does appear to constitute a novel functional assay capable of detecting qualitative vWF differences in plasma of affected vWD patients.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Colágeno/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Doenças de von Willebrand/diagnóstico , Fator de von Willebrand/metabolismo , Antígenos/metabolismo , Humanos , Valores de Referência , Doenças de von Willebrand/sangue , Doenças de von Willebrand/classificaçãoRESUMO
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