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D B Rylatt

Publications and source records attributed to D B Rylatt.

At least 37 records · Page 2Linked to original sources
PubMed1988

Serial D-dimer levels in the assessment of tumor mass and clinical outcome in ovarian cancer.

D-dimer (DD)--an end product of fibrinolysis--was measured in serum by enzyme immunoassay using a monoclonal antibody to the gamma gamma crosslink in patients with ovarian cancer, before primary surgery and during chemotherapy for 12 months or more. Serial DD levels were found to have a high sensitivity for the detection of tumor in patients with subclinical disease (91%) as well as for predicting progression of disease (100%). As determined by a careful second-look laparotomy in patients with complete clinical remission the DD marker was highly predictive of tumors less than 1 cm. The negative predictive value (82%) was higher than the positive predictive value (69%). However, there were 31% of the patients who showed a false-positive result; a close examination of the clinical data of these 9 patients failed to reveal an explanation for the positive DD levels. Despite the lack of specificity (50% in the present series), the findings support the use of DD in the assessment of patients with ovarian cancer, especially those with subclinical disease.

Antibodies, Monoclonal↗
PubMed1988

Production and characterization of monoclonal antibodies specific for Mycobacterium bovis.

A series of monoclonal antibodies (MAbs), specific for Mycobacterium bovis and BCG strains, were tested extensively for cross-reactivity to a wide range of mycobacterial species using ELISA, Western blotting and dot-blot analysis. The MAbs bound specifically to M. bovis and BCG and showed limited cross-reactivity with some strains of M. tuberculosis. All these MAbs recognized a 22 kDa protein previously termed MPB70, and by competitive ELISA analysis appeared to detect at least three M. bovis-specific determinants on the MPB70 molecule.

Animals↗
PubMed1987

Monoclonal antibody-based enzyme immunoassay for trypsinogen in neonatal screening for cystic fibrosis.

An enzyme immunoassay (EIA) with monoclonal antibodies against human trypsinogen in neonatal blood-spots has been evaluated for screening for neonatal cystic fibrosis (CF). In a retrospective study, 36 of 39 CF samples were distinguished from controls matched for age and storage time. 7 infants with CF were detected in 16,500 infants screened in a prospective study. The EIA is quicker and less labour intensive than conventional assays for the detection of immunoreactive trypsin and may have further advantages of specificity and sensitivity for monitoring the release of pancreatic zymogens in CF.

Antibodies, Monoclonal↗
PubMed1987

Measurement of crosslinked fibrin derivatives--use in the diagnosis of venous thrombosis.

The measurement of crosslinked fibrin derivatives in plasma has received evaluation as a screening test in the diagnosis of venous thrombosis. Plasma samples were taken from 104 patients undergoing venography because of clinical suspicion of lower limb venous thrombosis. The samples were assayed using a monoclonal antibody identifying an epitope on D dimer and larger crosslinked fibrin derivatives in an enzyme immunoassay. 100% of patients with positive venograms had elevated levels of these molecules. While a percentage of patients with negative venograms also had increased levels, alternative clinical explanations were apparent in most. A normal D dimer value excludes the diagnosis of venous thrombosis, while an increased value supports it. The measurement of crosslinked fibrin derivatives in plasma may play a role in the selection of patients for venography.

Adolescent↗
PubMed1987

Use of an antigen detection assay to determine mortality of Dirofilaria immitis after thiacetarsamide therapy.

The use of an antigen detection enzyme immunoassay (EIA) to determine the post-treatment infection status of 16 dogs naturally infected with Dirofilaria immitis was investigated. Dogs were treated with thiacetarsamide at a dose rate of 12mg/4.5kg twice daily for 2 days, bled at regular intervals and necropsied 9 weeks later. The infection status of all dogs at necropsy was compared to the ratios of optical density (OD) values from the EIA using fresh plasma samples (day 60/day 0 = R60) and dogs were divided into 2 groups. Using the R60 ratios, those dogs with fewer than 2 live adult worms or immature worms at necropsy ("cleared" dogs) could be differentiated with 95% confidence from those dogs with more than 1 live adult worm ("non-cleared" dogs). Changes in the average OD values from the plasma of "cleared" dogs and "non-cleared" dogs were similar up to 46 days after treatment but diverged significantly thereafter. The efficacy of thiacetarsamide was 50% if all worms were considered and 75% if the presence of immature worms was ignored. The benefits of antigen detection assays for diagnosis and improved patient assessment and the use of an R60 ratio to assess the efficacy of adulticides such as thiacetarsamide are discussed in relation to their practical significance for clinicians.

Animals↗
PubMed1987

A latex agglutination assay for D dimer: evaluation and application to the diagnosis of thrombotic disease.

Although latex agglutination assays have been used for some years to diagnose thrombotic disorders, only recently has it been possible to measure specifically the products of fibrin breakdown in the presence of fibrinogen degradation products, by using monoclonal antibodies. We have evaluated a preparation of latex particles coupled to the monoclonal antibody DD-3B6/22, which is specific for cross-linked fibrin degradation products (XDP) and allows accurate discrimination between normal and pathological conditions. Of samples from 515 apparently healthy volunteers, 97.7% failed to agglutinate the latex; the normal reference interval for XDP determined by enzyme immunoassay was less than 78-320 micrograms/L. The use of different anticoagulants with or without the addition of a protease inhibitor had no significant effects on the results of the latex assay. The latex preparation provides a useful, rapid diagnostic tool for assaying small numbers of samples or as an emergency test.

Antibodies, Monoclonal↗
PubMed1986

Rapid detection of cross-linked fibrin degradation products in plasma using monoclonal antibody-coated latex particles.

Conformational and structural changes on conversion of fibrinogen to fibrin and its cross-linking by Factor XIIIa lead to the development of new antigenic determinants that permit differentiation between their plasminolytic cleavage products. A monoclonal antibody (DD-3B6/22) that is specific for cross-linked fibrin derivatives containing the D dimer configuration has been used in developing a latex agglutination procedure that can detect fibrin degradation products in either plasma or serum. Fibrinogen or its degradation products do not cross-react with this antibody. Results were calibrated with an enzyme immunoassay, which used a purified D dimer standard. Plasmas from 40 normal subjects, all having D dimer levels below 250 ng/mL measured by enzyme immunoassay, were all negative by latex assay. In contrast, positive latex agglutination titers were obtained with 87 of 88 patients with demonstrated deep venous thrombosis, pulmonary embolism, or disseminated intravascular coagulation. Compared to enzyme immunoassay, latex agglutination assay is less sensitive, but this latex procedure provides a rapid and less elaborate test for elevated levels of cross-linked fibrin degradation products in patients with thrombosis. Plasma assays for fibrin degradation products are preferable to those using serum.

Antibodies, Monoclonal↗
PubMed1985

Evidence from the use of monoclonal antibody probes for structural heterogeneity of the growth hormone receptor.

We describe the use of four monoclonal antibodies (MAbs) to the rabbit liver growth hormone (GH) receptor and one raised against purified rat liver GH receptor to characterize liver receptor subtypes which differ in their hormone-binding regions. The anti-(rat liver GH receptor) MAb both inhibited and precipitated rat and rabbit GH receptors, but only one-half of 125I-oGH (ovine GH) binding to liver microsomes could be inhibited by excess antibody. Conversely, only one-half of 125I-anti-(rat GH receptor) MAb binding was inhibited by excess oGH and Scatchard plots for this MAb exhibited two components. Although only 50% of 125I-oGH binding to membranes was inhibited by this MAb, all solubilized receptor could be immunoprecipitated. We postulate two epitopes for the anti-(rat GH receptor) MAb, one located at the hormone-binding site (inhibitory site) and one elsewhere (immunoprecipitating site). A second, rabbit-specific antibody (MAb 7) inhibited 85% of hormone binding but only 30% of 125I-anti-(rat GH receptor) MAb binding to rabbit liver microsomes. A combination of this MAb with the anti-(rat GH receptor) MAb totally inhibited 125I-oGH binding. MAb 7 alone totally inhibited 125I-rat GH binding to rabbit liver microsomes, as it did with 125I-oGH binding to purified receptor. On the basis of these results and others we postulate three types of GH receptor in rabbit liver membranes and ascribe approximate extents of 125I-oGH binding to each. A cytosolic 'GH receptor' which is not poly(ethylene glycol)-precipitable is shown to share five epitopes with 'type 2' microsomal receptors. Purified plasma membrane and endoplasmic reticulum fractions derived from a rabbit liver microsomal preparation have identical antigenic characteristics with respect to the GH-binding region, indicating that the heterogeneity we describe is not related to receptor processing. Of the three types of GH receptor in the plasma membrane of the rabbit (and possibly rat) we postulate that one (type 1) corresponds to the GH receptor involved in stimulating growth and possesses all of the epitopes studied here. A second (type 2) appears to be identical with the cytosolic 'GH receptor' and lacks the epitope for the anti-(rat GH receptor) MAb in the hormone binding site region. A third (type 3) does not possess the epitope for the inhibitory anti-(rabbit GH receptor) MAb, appears not to bind rat GH and is lost during purification. The availability of type-specific MAbs will facilitate assignment of specific functions to liver receptor subtypes which mediate the multiple functions of GH.

Animals↗
PubMed1985

Rapid screening of monoclonal antibodies by spin adherence double immunosorbent test (SADIST).

The spin adherence double immunosorbent test (SADIST) is a simple, rapid immunoassay with sensitivity similar to the enzyme-linked immunosorbent assay (ELISA). A 1-step SADIST has been found suitable for rapid screening of hybridomas for antigen-specific monoclonal antibodies (MAb). In this procedure hybridoma supernatants are added to antigen coated microplates followed by commercially available antiglobulin beads. The microplate is immediately centrifuged. Wells containing antigen-specific MAb produce a mat of beads whilst wells without antigen-specific MAb produce a button of beads. No washing or incubation steps are necessary and results are read within minutes of adding beads to test supernatants. By comparison, ELISA tests require several hours to perform with multiple wash steps and further reagent additions. A 2-step SADIST was also assessed. Supernatants are incubated in the microplate as for an ELISA and a wash step precedes the addition of antiglobulin beads. A panel of 117 hybridoma supernatants was selected to assess the suitability of the SADIST techniques for hybridoma screening. The supernatants were added to antigen-coated microplates and SADIST and ELISA tests performed. The SADIST correctly discriminated most hybridoma supernatants that were clearly positive or negative by ELISA. It was also found possible to perform SADIST followed by ELISA tests on the same microplate well without significantly affecting ELISA values.

Antibodies, Monoclonal↗
PubMed1985

Monoclonal antibodies directed against Brucella abortus cell surface antigens.

A panel of monoclonal antibodies (MAb) has been raised against Brucella abortus cell surface antigens from mice immunized with either heat/phenol treated or UV killed bacterial suspensions of B. abortus. The hybridomas were screened by either a microagglutination procedure or by an indirect enzyme immunoassay (EIA) on sonicated bacterial preparations. From a large number of MAb generated by various procedures, two distinct types of MAb emerged. The most numerous type was capable of agglutinating B. abortus and reacting with a soluble preparation of lipopolysaccharide (LPS). A second type was not capable of agglutinating the bacterial suspensions or of binding to the soluble LPS preparation but reacted with an antigen present in bacterial sonicates. Two MAb of this type react differentially with sonicates prepared from virulent and avirulent strains of B. abortus. There appeared to be sufficient evidence from our analysis of the relative degree of cross reaction with antigens present on a range of B. abortus strains and Brucella and xenogenic bacterial species to conclude that each of the seven MAb was recognising a separate antigenic site on the B. abortus cell surface.

Animals↗
PubMed1985

A competitive enzyme immunoassay for the detection of bovine antibodies to Brucella abortus using monoclonal antibodies.

A competitive enzyme immunoassay (EIA) for the detection of circulating bovine antibodies to Brucella abortus has been developed using horseradish peroxidase conjugated monoclonal antibodies (MAb) raised against B. abortus cell surface antigens. Antibodies present in the serum of either vaccinated or infected cattle can apparently displace the conjugated MAb from the lipopolysaccharide antigen (LPS) in a quantitatively different manner allowing an assessment of immune status of the animal. The results from a panel of sera from animals with a known status of vaccination or infection indicated that the test was more selective in the detection and discrimination of infected from uninfected or immunized animals, than conventional complement fixation, agglutination or indirect enzyme immunoassay procedures.

Animals↗
PubMed1985

Serum crosslinked fibrin (XDP) and fibrinogen/fibrin degradation products (FDP) in disorders associated with activation of the coagulation or fibrinolytic systems.

Soluble crosslinked fibrin derivatives (XDP) in serum were determined by enzyme immunoassay utilizing monoclonal antibodies and compared with serum fibrinogen/fibrin degradation products (FDP) assayed by conventional techniques. In healthy subjects and patients with miscellaneous disorders not usually associated with activation of the haemostasis mechanism, mean XDP levels were 45 and 70 ng/ml respectively. However, elevated levels of XDP occurred in conditions commonly associated with intravascular and possibly extravascular activation of the coagulation system. Markedly raised mean XDP values (677-6900 ng/ml) occurred in treated pulmonary embolism, disseminated neoplasia, severe inflammatory disorders and complicated postoperative states, and lesser but significant elevation (mean 150-400 ng/ml) in treated venous thrombosis, uneventful postsurgical states, localized neoplasia, liver disease and symptomatic arterial disease. Levels during initial streptokinase therapy (mean 24 000 ng/ml) fell tenfold as treatment was continued. The degree of XDP elevation over normal values was significantly higher than that of FDP in conditions with a propensity for venous thrombosis (post-operative states, disseminated neoplasia and inflammatory diseases) than in liver disease, localized neoplasia or patients receiving heparin therapy for venous thromboembolism.

Adult↗
PubMed1984

Measurement of cross linked fibrin derivatives in plasma: an immunoassay using monoclonal antibodies.

Fibrinogen degradation, fibrin polymerisation, and the insertion of cross links into fibrin by fibrin stabilising factor lead to the appearance of new antigenic determinants. Antibodies against these antigenic sites may react specifically with the derivatives but not with the parent molecules. We have utilised a monoclonal antibody, which interacts with the cross linked fragment D dimer and related high molecular weight fibrin derivatives, to develop an enzyme immunoassay which measures cross linked fibrin derivatives in plasma and serum using D dimer as standard. Mean concentration in plasma from normal subjects was 75 ng/ml with an upper limit of about 144 ng/ml. Concentrations in patients with pulmonary embolism, deep venous thrombosis, arterial thromboembolism, and disseminated intravascular coagulation were raised in all cases. Confirmation of the specific increase of cross linked fibrin derivatives in patients with disseminated intravascular coagulation was obtained by parallel monitoring of their fibrin degradation products in serum using affinity chromatography and sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis. In many patients the plasma concentrations greatly exceeded the serum values of cross linked fibrin degradation products, suggesting that the procedure can measure fibrin derivatives in plasma which are absent from serum.

Antibodies, Monoclonal↗
PubMed1984

Monoclonal antibodies to the rabbit liver growth hormone receptor: production and characterization.

Monoclonal antibodies to the GH receptor (GHR) have been produced by the application of hybridoma technology to splenic lymphocytes from BALB/C mice immunized with a human (hGH) affinity purified preparation of rabbit liver GHR. Primary screening of 384 wells yielded 4 antibodies able to immunoprecipitate [125I]iodo-hGH complexes with purified GHR and one able to inhibit binding of [125I]iodoovine GH ([125I]iodo-oGH) to rabbit liver microsomes. These cells were cloned and grown as ascitic tumors with loss of 1 of the 4 precipitators. Ascitic fluids contained monoclonal antibodies of high titer (inhibitor and 2 precipitators, 1:2.0 X 10(5); one precipitator, 1:2.0 X 10(4)) and high affinity (precipitators, 2.5-6.0 X 10(9) M-1; inhibitor, high affinity component, 6.4 X 10(10) M-1), which were isotyped as IgG1K and IgG2aK (1 precipitator). These antibodies did not cross-react with rabbit insulin or PRL receptors in the appropriate receptor assays and did not possess antihormone activity. Binding of [125I]iodo-MAb7, the inhibitory antibody, was totally blocked by the addition of excess unlabeled oGH or hGH, although these hormones had no effect on binding of the 125I-labeled precipitators. Scatchard analysis of [125I]iodo-oGH binding in the presence of MAb7 showed decreased binding by loss of sites rather than affinity. Antibody dilution curves and Scatchard plots for MAb7 binding provided evidence for two types of GHR in the rabbit liver, in accord with previously published data based on hormone binding studies. All precipitating antibodies gave an enhancement of [125I] iodo-oGH binding with purified receptor (up to 360% of polyethylene glycol-precipitated control), but only minimal enhancement with solubilized microsomal membranes. This enhancement was shown to be due to an increase in receptor number rather than affinity. After examining a number of hypotheses, we concluded that the enhancement was an artifact resulting from a nonpolyethylene glycol-precipitable species of GHR which could be totally precipitated by the monoclonal antibodies. We have produced and characterized four monoclonal antibodies to the GHR which will be of value in characterizing the structure and function of this receptor.

Animals↗
PubMed1984

Demonstration of lymphocyte surface lectins that recognize sulphated polysaccharides.

Cholate extracts of murine lymphocytes were shown to contain haemagglutinating activity against autologous erythrocytes. The species specificity and sugar inhibition pattern of the haemagglutinin closely paralleled the specificity of autorosetting, an interaction that had been shown previously to involve the recognition by lymphocytes of carbohydrate structures on autologous erythrocytes. The probable identity of the haemagglutinin and autorosetting receptors was confirmed by experiments utilizing the unique plasma protein autorosette inhibition factor, which appears to block both interactions by masking carbohydrate acceptor sites on erythrocytes. Detailed sugar inhibition studies revealed that the haemagglutinin and autorosetting receptors have a high affinity for certain sulphated polysaccharides, such as heparin and dextran sulphate. Since similar sulphated polysaccharides have been shown previously to inhibit lymphocyte recirculation, a possible role for these receptors in lymphocyte homing and recirculation is discussed.

Agglutinins↗
PubMed1983

An immunoassay for human D dimer using monoclonal antibodies.

Monoclonal antibodies (MAb) were raised against human D dimer. The hybridomas were screened with a solid phase enzyme immunoassay against D dimer and fibrinogen degradation products. Among the panel of MAb identified, two distinct patterns emerged; the majority belonging to a panspecific class reacting against epitopes present on both D dimer and fibrinogen degradation product Dcate and a monospecific class reacting with determinants apparently present only on D dimer. A number of MAb were further characterised for their ability to specifically capture antigen in a solid phase enzyme immunoassay and assays were developed which have a sensitivity of 10 ng/ml for D dimer or crosslinked fibrin derivatives and may be suitable for detection of crosslinked derivatives in serum and plasma samples in a clinical situation.

Animals↗
PubMed1983

Measurement of crosslinked fibrin degradation products - an immunoassay using monoclonal antibodies.

We have prepared a monoclonal antibody which recognises an antigenic determinant on D dimer, a specific fragment resulting from the degradation of crosslinked fibrin. This antibody has been used in the development of an enzyme-linked immunoassay for D dimer and related degradation products containing crosslinked gamma-gamma chains, to provide a simple assay of circulating crosslinked fibrin degradation products suitable for clinical use. Since these crosslinked fibrin degradation products are characteristic of fibrinolysis, as distinct from fibrinogenolysis, their measurement should aid in the diagnosis, evaluation and monitoring of thrombotic and thrombolytic states. In preliminary studies, low concentrations of crosslinked fibrin derivatives were detected in normal sera. High levels were found in 30/30 patients with disseminated intravascular coagulation and in the majority of patients having deep venous thrombosis or pulmonary embolism.

Animals↗