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Biomedical subjects

A L Bloom

Publications and source records attributed to A L Bloom.

At least 55 records · Page 3Linked to original sources
PubMed1985

Immunoassays of factor IX antigen using monoclonal antibodies.

Monoclonal antibodies to factor IX were produced by immunization of balb/c mice with purified factor IX and fusion of spleen cells with SP-1 murine myeloma cells. Antibody producing hybrids were detected by an enzyme linked immunoassay (ELISA) and by coagulation inhibitor (Bethesda type) methods. Monoclonal antibodies with titres of greater than 1 X 10(5) tested by the ELISA and 5000-8000 inhibitor units were obtained. A new ELISA method was developed using one of these monoclonal antibodies to quantitate factor IX antigen (IXAg) in plasma samples from patients with hereditary factor IX deficiency. In addition a two-site solid phase immunoradiometric assay (IRMA) for factor IX was established. The results of these new methods were compared with those obtained on the same plasma samples using conventional factor IX coagulation assays and the Laurell rocket method. The lower limit for the detection of IXAg by the ELISA was approximately 0.01 unit per ml whilst that for the IRMA was about 0.001 unit per ml (normal plasma = 1 unit per ml). The lower detection limit for IXAg using the Laurell rocket method was about 0.06 units per ml. The improved sensitivity of the new immunoassays enabled quantitation of low levels of IXAg in patients with moderately severe factor IX deficiency and confirmed the presence of excess IXAg compared to IX activity (IXC) in a relatively high proportion of cases (28 out of 51 tested). Results of testing plasma from obligate carriers confirm the suggestion that measurements of IXAg and IXC may improve the classification of carrier status in these kindred.

Animals↗
PubMed1985

A study of cell mediated and humoral immunity in haemophilia and related disorders.

Helper (OKT4) and suppressor (OKT8) T lymphocyte populations and functional assays of cellular immunity were studied in 37 patients with haemophilia and related disorders in parallel with age matched control subjects. The study included 26 patients with factor VIII (FVIII:C) deficiency, eight patients with factor IX deficiency and three patients with severe von Willebrand's disease (vWd). In patients with factor VIII deficiency low helper T lymphocyte counts, low T helper: suppressor ratios and a diminished response to the lymphocyte mitogen phytohaemagglutinin with decreased natural killer cell activity were observed. Individuals with factor IX deficiency had low absolute T lymphocyte (OKT3) counts and T helper cell counts. Patients with severe factor VIII deficiency (FVIII:C less than 1 u/dl) had lower T helper suppressor ratios and lower killer cell and natural killer cell activity in comparison to mildly affected individuals and all of the severely affected factor IX deficient cases. Studies of humoral immunity revealed a generalized increase in immunoglobulin levels in patients with coagulation disorders of any type. The total haemolytic complement activity was reduced in a significant proportion of haemophilic subjects. Levels of alpha-1-interferon were elevated in the groups of haemophilic subjects studied. The abnormalities of cellular and humoral immunity observed did not correlate with the amount or type of coagulation factor administered to individual patients in the preceding 2 years. The most marked abnormalities of immune function occurred in the one patient diagnosed as suffering from AIDS on clinical grounds.

Adolescent↗
PubMed1985

AIDS and haemophilia.

The prevalence of overt AIDS in haemophilia A in the USA is about 3% treated patients and about 2% in UK. Nevertheless up to 90% of patients with haemophilia A treated with commercial concentrates of factor VIII prepared from the blood of paid donors are seropositive to the AIDS-associated virus LAV/HTLV III. Infection seems to have commenced in 1978 in USA and in 1980 in Europe. The accession rate to the symptomatic group is not known. Seropositivity is much lower in patients treated with blood products derived from voluntarily donated blood but such patients, even when seronegative, often show cellular immune defects. An additional immunosuppressive agent in factor VIII concentrates may be responsible for this and could predispose to LAV infection. AIDS, immune defects and seropositivity are less common in patients with haemophilia B even when treated with factor IX concentrate derived from paid donors. The reasons for this anomaly are not known. The diagnosis, social, family and counselling problems of infected patients are surveyed and the prospects for effective anti-viral or immunotherapy assessed. Treatment policies using heat-treated factor concentrates and the impact of donor screening are reviewed. It is concluded that the outlook for uninfected haemophiliacs with regard to AIDS is now good.

Acquired Immunodeficiency Syndrome↗
PubMed1984

Acquired immunodeficiency syndrome and other possible immunological disorders in European haemophiliacs.

A questionnaire concerning the occurrence of acquired immunodeficiency syndrome (AIDS) and possible related clinical syndromes in treated haemophiliacs was sent to directors of 201 European haemophilia centres. 135 replies were received from eighteen countries, covering 8928 patients with haemophilia A, 1889 with haemophilia B, and 2330 with von Willebrand's disease treated with blood products per year; 8 cases of AIDS were identified, one of which was in a patient with haemophilia B; 7 suffered from infections and 1 from pleomorphic non-Hodgkin's lymphoma. 3 other patients with lymphoma were recorded. The median age was 20 years with range 12-57 years and 3 were aged 14 years or less; 3 of the patients had died. 179 patients with other possible immunological disorders were recorded including 86 with lymphadenopathy but 70 of these occurred at one centre. At least 48 patients with thrombocytopenia were identified, suggesting that this condition is more common in treated haemophiliacs than is "idiopathic thrombocytopenic purpura" in the general population. None of the abnormalities could be related to use of any particular type of factor VIII concentrates. These results, together with 3 cases of AIDS previously reported and not returned in this study, point to an incidence of 11 cases in 13 000 treated haemophiliacs.

Acquired Immunodeficiency Syndrome↗
PubMed1984

Carrier detection by direct gene analysis in a family with haemophilia B (factor IX deficiency).

DNA from a patient with severe factor IX deficiency (haemophilia B) in whom an inhibitor to factor IX had developed was studied with four genomic gene probes specific for the factor IX gene. All gave a negative result, indicating at least a partial gene deletion. Eight female relatives, covering four generations, were also studied. Restriction-enzyme-fragmented DNA was probed in each case and the level of binding assessed by darkening of the autoradiograph. The DNA of this patient's sister and mother had reduced signals, when compared with that of normal female subjects, indicating the presence of the defective gene. However, the other six female subjects (grandmother, great-grandmother, maternal aunt, and three female cousins) had normal signals. Levels of factor IX and factor IX antigen were also normal in these subjects. By direct gene analysis in this family, the point of mutation has been identified (mother) and diagnosis of the sister as a carrier confirmed.

Autoradiography↗
PubMed1984

Atherosclerosis and von Willebrand factor. I. Prevalence of severe von Willebrand's disease in western Europe and Israel.

The prevalence of von Willebrand's disease (VWD) in Western European countries and Israel was assessed. Possible patients were identified initially by questionnaire and those with plasma levels of von Willebrand factor antigen ( WFAg ; factor VIII related antigen) less than 1 u/dl (less than 1%) were confirmed by immunoradiometric assay performed at four reference centres. The prevalences of severe VWD in Scandinavian countries were similar to each other and about 10 times greater than those in other Western European countries. The prevalence in Israel was intermediate. The Registry will form the basis for future non- invasive epidemiological studies of atherosclerosis in these individuals.

Adolescent↗
PubMed1984

The effect of desamino-D-arginine vasopressin (DDAVP) and naloxone infusions on factor VIII and possible endothelial cell (EC) related activities.

Studies on the effect of DDAVP both in vitro and in vivo are reported. In order to define the extent of the DDAVP induced rise of circulating endothelial cell proteins in normal individuals and the endothelial cell defect in von Willebrand's disease (vWd) we have measured the effect of intravenous DDAVP on a range of possible endothelial cell markers in normal subjects and in patients with mild haemophilia and vWd. In a series of double blind cross over studies on normal volunteers we have tested the effect of naloxone, DDAVP or saline on circulating levels of factor VIII related activities (VIIIR) and plasminogen activator (PA). The results confirmed the effect of DDAVP on circulating levels of VIIIR and PA but showed that it did not induce release of these activities from cultured endothelial cells in vitro nor did it influence circulating levels of other endothelial cell markers including fibronectin, antithrombin III and platelet factor 4. Infusion of nalaxone did not significantly alter circulating levels of VIIIR or PA nor the response of these to DDAVP suggesting that normally these activities are not subjected to a vasopressin drive.

Adolescent↗
PubMed1983

Differential localisation and synthesis of porcine factor VIII related antigen (VIIIR:AG) in vascular endothelium and in endothelial cells in culture.

Porcine and human umbilical vein and adult blood vessels were studied for the presence and synthesis of factor VIII related antigen (VIIIR:Ag) and fibronectin (Fn) by immunofluorescence histology and immunoautoradiography. Investigation of human tissue confirmed the widespread distribution of VIIIR:Ag on the endothelium of all blood vessels examined but observations on porcine tissue gave different results. Porcine umbilical vein and porcine adult veins were positively stained for VIIIR:Ag whilst porcine aorta and other pig arteries appeared to be negative or only weakly positive. Some blood vessels (?venous) in the adventitia of porcine aorta were positively stained whilst adjacent ones (?arterial) were negative. Radiolabelled methionine was added to culture medium and proteins synthesised by cultured EC were examined by two dimensional crossed immunoelectrophoresis and autoradiography. Identification of radiolabelled precipitin arcs provided a highly sensitive method for confirming the specificity of antisera and for detecting VIIIR:Ag and Fn. Examination of cultured human umbilical vein EC confirmed the synthesis of VIIIR:Ag and distinguished between VIIIR:Ag and Fn. Studies of porcine umbilical vein EC in culture gave similar results to those observed with corresponding human EC. However, cultures derived from porcine aorta did not demonstrate synthesis of VIIIR:Ag and microscopy failed to locate VIIIR:Ag in these cells with certainty. The results confirmed the synthesis of VIIIR:Ag by human and porcine umbilical vein EC but differences in staining reactions and the apparent inability to synthesise VIIIR:Ag by cells derived from porcine aorta suggested that porcine EC at different anatomical sites may subserve different functions.

Animals↗
PubMed1983

Renal papillary necrosis in haemophilia and christmas disease.

Haematuria is common in haemophilia and Christmas disease. A variety of radiological abnormalities in the urinary tract has been described in the literature, the commonest being hydronephrosis and obstructive uropathy. The findings in 26 cases seen in this centre were reported by Beck and Evans in 1972; although urinary-tract obstruction was relatively common, it was noted that signs suggestive of renal papillary necrosis (RPN) were present in some cases. In the present study, 20 of these patients were reinvestigated up to 10 years after the original radiological studies; a total number of 46 past and present intravenous urograms were examined. There was evidence of RPN in 11 of the 20 patients, and in five of these the signs had changed over the intervening period. The relevance of these findings is discussed; none of the patients showed evidence of renal failure. It is suggested that episodes of urinary-tract obstruction could be related to papillary sloughing, in some cases, rather than clot obstruction or obstruction due to retroperitoneal bleeding.

Adult↗
PubMed1983

The interaction between factor VIII clotting antigen (VIIICAg) and phospholipid.

The interaction between factor VIII clotting antigen (VIIICAg) and phospholipid (PL) was studied using a two-site solid phase immunoradiometric assay (IRMA) for VIII CAg. Incubation (2 h, 37 degrees C) of normal plasma, cryoprecipitate or factor VIII concentrate with Diagen PL (0.5 mg/unit VIIICAg) resulted in 80-90% loss of IRMA-measurable VIIICAg. No loss of factor VIII related antigen (VIIIRAg) or factor VIII clotting activity (VIIIC) was seen. Treatment of factor VIII concentrate with purified PLs showed greatest VIIICAg reduction with phosphatidylserine, less with phosphatidylethanolamine, and very little with phosphatidylcholine. The action of phospholipase-C (PL-C) on VIIICAg-PL complexes was investigated, with enzyme activity being monitored by thin-layer chromatography. Treatment of normal plasma, cryoprecipitate or factor VIII concentrate with PL-C (5 u/unit VIIICAg) resulted in 25%, 25% and 30% increases in VIIICAg. No increase in VIIIC or VIIIRAg was seen. Preincubation of factor VIII concentrate with PL, followed by PL-C incubation, resulted in 70-80% recovery of measurable VIIICAg. Incubation of 'activated' prothrombin complex with PL-C increased VIIICAg by 44% (Autoplex) and 80% (FEIBA), indicating VIIICAg-PL complexes are present. Incubation of factor VIII concentrate with fresh platelet lysate led to a reduction in VIIICAg (100 u/dl to 21 u/dl). In fresh platelet lysate alone low VIIICAg levels were detectable (0.71 X 10(-3) u/10(9) plt). After PL-C incubation VIIICAg levels increased to 9.8 X 10(-3) u/10(9) plt) (14-fold increase). Thus VIIICAg in platelets may be hidden by VIIICAg-PL complexes.

Antigens↗
PubMed1983

Human factor IX inhibitors: immunochemical characteristics and treatment with activated concentrate.

Plasma was obtained from two patients with severe factor IX deficiency who had developed specific inhibitors of factor IX. Immunochemical characterization of the inhibitors by coagulation inhibitor neutralization assays and by immunoelectrophoretic methods demonstrated that both were IgG antibodies. One of the antibodies appeared to be monoclonal in origin with IgG subclass 4 heavy chains and lambda light chains. The other appeared to be oligoclonal and contained IgG subclass 1 and subclass 4 heavy chains and kappa and lambda light chains. One of the patients was treated with conventional, non-activated factor IX concentrate and with activated factor IX concentrate (Feiba) for repeated bleeding episodes. Administration of Feiba resulted in a progressive shortening of the kaolin cephalin clotting time and was followed by a good clinical response. Infusion of non-activated factor IX concentrate failed to induce clinical resolution of haemarthroses and had minimal effect on laboratory tests. The presence of circulating immune complexes could not be demonstrated in this patient.

Adult↗
PubMed1982

Immunohistological comparison of platelet factor 4 (PF4), fibronectin (Fn) and factor VIII related antigen (VIIIR:Ag) in human platelet granules.

Immunofluorescence microscopy was used to study the localization of platelet factor 4 (PF4), fibronectin (Fn) and factor VIII related antigen in washed normal platelets and those from patients with severe von Willebrand's disease (vWd). Platelets were prepared by an improved multi-unit modification of the Sayk chamber. This facilitated the preparation of samples with minimum disruption of platelets and readily permitted the demonstration of intact platelet granules by the immunological techniques used. The antigens were demonstrated by treating the same preparations sequentially with appropriate heterologous antisera and species specific fluorescein or rhodamine conjugated antisera. Comparison of the different antigens in identical platelets indicated that Fn and VIIIR:Ag were localized to the same granules as PF4 and the results were thus consistent with their presence in alpha granules. Fn and PF4, but not VIIIR:Ag, were present in platelet granules of patients with severe vWd. The antigens were always detected in the same granules, and major sub-populations of differently stained granules were not observed. The methods were applied to investigate normal platelets aggregated with collagen. Fn and VIIIR:Ag were detected in platelet granules after aggregation although the granules themselves were possibly differently distributed in these samples compared with the non-aggregated platelets. The localization of PF4 could not be reliably assessed in aggregated platelets by these methods. The techniques may be useful in localizing platelet antigens and studying release during aggregation.

Antigens↗
PubMed1982

Quantitation of coagulant antigens and inhibition of activated protein C in combined factor V VIII deficiency.

Inhibition of activated human protein C was assessed in an amidolytic assay system using normal human plasma and samples from patients with hereditary coagulation abnormalities. In eight experiments normal plasma inhibited 63.5% (+/- 15.6%) protein C activity. Plasma from patients with haemophilia A or isolated factor V deficiency gave results which were not significantly different from normal. However, plasma from patients with combined factor V and factor VIII deficiencies inhibited an average of 24.5% (+/- 13.6%) of the amidolytic activity (P less than 0.01). Two of these plasma samples failed to inhibit any protein C activity. The relationship between the level of inhibitor and those of factor V and factor VII coagulant antigens (VCAg and VIIICAg) in the combined defect was investigated. There was no significant correlation between the level of inhibitor and any of the coagulation immunoassays on these stored samples but there was significant correlation between VCAg and VIIICAg in some assay systems. The levels of VCAg and VII CAg was low in most samples from patients with the combined defect which was in contrast to the results obtained when normal plasma was incubated with activated protein C in vitro. The findings are consistent with the presence of biochemical similarities between factors V and VIIIC molecules, but the role of activated protein C and its inhibitor in hereditary combined factor V/VIII deficiency remains to be firmly established.

Antigens↗
PubMed1981

Platelet and coagulation function in patients with abnormal cardiac valves treated with sulphinpyrazone.

Eight patients on warfarin with rheumatic heart disease and prosthetic cardiac valves were selected for study on the basis of persistently elevated plasma beta-thromboglobulin (beta-tg) and platelet factor 4 (PF4) concentrations. Platelet mean lifespan and fibrinogen half life were short, and positively correlated, and both were inversely related to the plasma concentration of the platelet specific proteins. Antithrombin III (ATIII) levels were also reduced. Treatment with sulphinpyrazone resulted in lengthening of both platelet and fibrinogen survival, a rise in ATIII but no change in the beta tg or PF4 concentrations. It is concluded that patients with abnormal cardiac valves and raised plasma levels of beta tg or PF4 have, despite warfarin, a consumption coagulopathy that can be inhibited by sulphinpyrazone.

Adult↗
PubMed1981

Factor VIII clotting antigen (VIIICAg) in haemophilia measured by two immunoradiometric assays (IRMA) using different antibodies, and the measurement of inhibitors to procoagulant factor VIII (VIIIC) by IRMA.

Factor VIII clotting antigen (VIIICAg) was measured by immunoradiometric assay (IRMA) using two different antibodies. Both antibodies arose in polytransfused severe haemophiliacs and had similar titres against VIIIC. In 12 normal plasmas there was no significant difference in VIIICAg values obtained (VIIICAg (AbI) = VIIICAg (AbII)). In the majority of 15 severe haemophiliacs tested VIIICAg was undetectable by both antibodies. In 28 mild to moderate haemophiliacs VIIICAg (AbII) was significantly greater than VIIICAg (AbI) (P less than 0.01) suggesting different antigenic determinants. The difference, however, was small and does not affect diagnosis of haemophilia. A modified IRMA has been used to measure anti VIIIC inhibitors by competition of the inhibitor with 125I labelled VIIICAg antibodies for common antigenic determinants. Using an inhibitor of 225 Bethesda units as a standard, results by IRMA of inhibitors in severe haemophiliacs have been similar to those obtained by clotting assay, but with a sensitivity of 0.01 u/ml suggesting the possible use in the detection of weak inhibitors.

Antibodies↗