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Biomedical subjects

D M Keeling

Publications and source records attributed to D M Keeling.

At least 19 recordsLinked to original sources
PubMed2000

Evaluation of the von Willebrand factor antigen (vWF-Ag) assay using an immuno-turbidimetric method (STA Liatest vWF) automated on the MDA 180 coagulometer.

The standard enzyme-linked immunosorbent assay (ELISA) test for von Willebrand factor antigen (vWF-Ag), though sensitive and specific, does not deliver the flexibility to handle single sample assessments economically or provide rapid, emergency testing capability. The present study examined the performance characteristics of an immuno-turbidimetric assay kit (STA Liatest) modified for automation on the MDA(R)180 coagulation analyser using a supplied protocol. One hundred and sixteen patient samples were assessed by both the standard and the modified method. The correlation coefficient was 0.98 across the range of values 1-487 IU/dl. Above 200 IU/dl, where specimen dilution was indicated, there was greater diversity (r = 0.86) between the techniques. Plotting the difference between methods against the mean of both showed excellent agreement between methods below 100 IU/dl vWF. The intra-assay and interassay coefficients of variation were less than 3% at both low and normal range levels. The MDA(R)180 automated vWF assay merits consideration as an alternative to ELISA testing that provides random access and result availability within 30 min.

Antigens↗
PubMed1999

Diagnostic importance of the two-stage factor VIII:C assay demonstrated by a case of mild haemophilia associated with His1954-->Leu substitution in the factor VIII A3 domain.

In some families with mild haemophilia higher results are obtained for factor VIII activity (FVII:C) determined by one-stage assay than by two-stage or chromogenic assays. Amino-acid substitutions in the A1, A2 and A3 domains of factor VIII have been described in affected individuals with this phenotype. We describe a case of mild haemophilia A in which FVIII:C measured by one-stage assay was normal at 106%. However, FVIII:C levels measured by two-stage and chromogenic assays were 18% and 35% respectively. DNA analysis revealed a novel mutation in the A3 domain of factor VIII, His1954-->Leu. In a molecular model of the FVIII A domains, His1954 is placed in close proximity to two other mutations that have previously been shown also to be associated with one-stage/two-stage discrepancies. In this patient the diagnosis of haemophilia A would be missed if only the one-stage assay was used.

Amino Acid Substitution↗
PubMed1999

D-dimer for the exclusion of venous thromboembolism: comparison of a new automated latex particle immunoassay (MDA D-dimer) with an established enzyme-linked fluorescent assay (VIDAS D-dimer).

The use of D-dimer tests to exclude venous thromboembolism is an important advance in clinical practice and also has economic benefits. Ideally the test should be objective and a test that could be run on the routine coagulometer would obviate the need for additional investment in alternative hardware. A new automated latex particle immunoassay (MDA D-dimer) that can be run on a routine coagulometer was compared with a well established enzyme linked fluorescent assay (VIDAS D-dimer) on the basis of their ability to exclude venous thromboembolism. The assays were compared in 49 patients presenting to the emergency department with clinically suspected deep vein thrombosis or pulmonary embolism. After objective diagnostic imaging, 20 patients were confirmed to have venous thromboembolism. There was strong agreement between the assays in individual patients. Using a cut-off of 500 micrograms/l, both tests had a sensitivity of 100% and therefore a negative predictive value of 100%, however the MDA test would have spared more patients (20% vs. 12%) from further testing if a negative D-dimer was used to rule-out the diagnosis. It was concluded that a rapid, objective latex D-dimer test run on a routine coagulometer (MDA D-dimer) can be used to exclude the diagnosis of venous thromboembolism.

Enzyme-Linked Immunosorbent Assay↗
PubMed1997

Cryoprecipitate prepared from plasma virally inactivated by the solvent detergent method.

There remains a small risk of viral transmission from single-donor blood components such as fresh frozen plasma (FFP) and cryoprecipitate which have not been subjected to a viral inactivation procedure. It is now possible to subject pooled FFP to viral inactivation by the solvent detergent (SD) treatment method, but with some loss of coagulation factors. To establish whether cryoprecipitate prepared from SD plasma would be suitable for the treatment of hypofibrinogenaemia and von Willebrand's disease (VWD), control and SD cryoprecipitate were assayed for factor VIII. von Willebrand factor (VWF) and fibrinogen content. In SD cryoprecipitate, levels of VWF activity and antigen were only 36% and 37% of control values respectively, whereas fibrinogen was 72%. The highest molecular weight multimer of VWF:Ag were absent from both SD plasma and SD cryoprecipitate. SD cryoprecipitate would thus be unsuitable for treating VWD, but would provide an alternative to untreated individual donor units of cryoprecipitate for the treatment of hypofibrinogenaemic states.

Blood Preservation↗
PubMed1994

Platelet aggregation in response to four low molecular weight heparins and the heparinoid ORG 10172 in patients with heparin-induced thrombocytopenia.

A simple rapid platelet aggregation test was used to evaluate cross-reactivity of four low molecular weight heparins and the heparinoid ORG 10172 in three patients with heparin-induced thrombocytopenia. The low molecular weight heparins cross-reacted in 11 out of 12 tests. The heparinoid ORG 10172 did not cross-react in any of the patients. One patient was treated with ORG 10172 and thrombocytopenia resolved.

Adult↗
PubMed1993

Haematological manifestations of systemic lupus erythematosus.

Haematological involvement is common in systemic lupus erythematosus (SLE). Whilst anaemia is most often due to chronic disease, other causes such as autoimmune haemolytic anaemia and hypoplastic anaemia need to be considered. The increased risk of infection in patients with SLE is due in part to changes in the white blood cells though treatments do not yet aim to modify these. Thrombocytopenia occurs frequently and is almost invariably autoimmune. It is often of little consequence, but may occasionally be severe and serious, requiring aggressive treatment. Patients with SLE have an increased risk of thrombosis, increased further in the presence of antiphospholipid antibodies (aPL). Changes in the haemostatic system and new insights into the nature of aPL are described.

Anemia↗
PubMed1993

Role of beta 2-glycoprotein I and anti-phospholipid antibodies in activation of protein C in vitro.

AIMS: To investigate the effect of beta 2-glycoprotein I (beta 2 GPI) on the thrombin/thrombomodulin dependent activation of protein C; and to determine whether beta 2 GPI dependent anticardiolipin antibodies have any effect. METHODS: Protein C was activated by thrombin in the presence of thrombomodulin and phospholipid vesicles in an in vitro system. The effect of adding purified beta 2 GPI to this system was observed. Affinity purified anticardiolipin antibodies and total IgG from patients with anticardiolipin antibodies and the lupus anticoagulant were studied for their effects on protein C activation in the presence and absence of beta 2 GPI. RESULTS: beta 2-Glycoprotein I had no effect on the activity of preformed activated protein C. When the phospholipid vesicles were incubated with beta 2 GPI before the addition of protein C, the activation of protein C was inhibited in a dose dependent manner. With phosphatidylserine:phosphatidylcholine vesicles at a concentration of 1 microM:2 microM, beta 2 GPI began to inhibit the reaction at a concentration of 15 nM, and at 4 microM (the normal plasma concentration) the activation of protein C was reduced to 40%. Anticardiolipin antibodies had no demonstrable effect. CONCLUSIONS: beta 2-Glycoprotein I inhibits protein C activation in an in vitro system. Its physiological role is unknown but it has potential procoagulant as well as anticoagulant properties. An effect of antiphospholipid antibodies on protein C activation, which might explain their association with thrombosis, could not be shown.

Antibodies, Antiphospholipid↗
PubMed1993

Lupus anticoagulant activity of some antiphospholipid antibodies against phospholipid bound beta 2 glycoprotein I.

AIMS: To determine whether beta 2 glycoprotein I (beta 2GPI) dependent anticardiolipin (aCL) antibodies detected in solid phase enzyme linked immunosorbent assays can also have lupus anticoagulant activity. METHODS: Six anticardiolipin antibodies were affinity purified from patients with these antibodies and lupus anticoagulant activity in their plasma. RESULTS: The anticardiolipin antibodies bound only to anionic phospholipids in the presence of beta 2GPI and bound to beta 2GPI in the absence of phospholipids. Four out of six had lupus anticoagulant activity in the dilute Russell viper venom time test. CONCLUSIONS: The results show that some beta 2GPI dependent aCL are lupus anticoagulants. It is unclear why only some should have lupus anticoagulant activity while others do not.

Anions↗
PubMed1992

Treponemal infection coexisting with systemic lupus erythematosus.

A patient with systemic lupus erythematosus (SLE)/erosive arthritis overlap is described who was also shown to have late latent treponemal infection. The possibility of serological reactivation is discussed. Since antiphospholipid antibodies are the basis of the reagin tests for syphilis as well as being present in patients with SLE, difficulties with diagnosis may ensue.

Female↗
PubMed1992

Some 'antiphospholipid antibodies' bind to beta 2-glycoprotein I in the absence of phospholipid.

Some antiphospholipid antibodies (aPL) only bind to anionic phospholipids in the presence of a serum cofactor, beta 2-glycoprotein I (beta 2GPI). Whether these aPL can bind to beta 2GPI in the absence of phospholipid is controversial. We have purified anticardiolipin antibodies (aCL) from the plasma of four patients and beta 2GPI from normal plasma by solid phase affinity methods. All four aCL bound to cardiolipin and phosphatidylserine in the presence of beta 2GPI but not in its absence. The binding of two of the antibodies to cardiolipin and phosphatidylserine at various concentrations of human beta 2GPI was compared with that obtained using 10% bovine serum. The two antibodies responded differently to increasing beta 2GPI concentrations, and binding to phosphatidylserine was relatively greater than to cardiolipin using human beta 2GPI. All four aCL bound to plastic plates coated with beta 2GPI in the absence of phospholipid, and beta 2GPI in the fluid phase had no effect on binding. Binding to beta 2GPI coated plates was increased equally when bovine serum or bovine albumin were used as the sample diluent in place of gelatine. These findings and those of others have important implications for the design of assays for antiphospholipid antibodies.

Antibodies, Anticardiolipin↗