Novel mesenchymal and haematopoietic cell isoforms of the SHP-2 docking receptor, PZR: identification, molecular cloning and effects on cell migration.

SHP-2 (Src homology phosphatase type-2) is essential for haematopoietic skeletal and vascular development. Thus the identification of its binding partners is critically important. In the present study, we describe a unique monoclonal antibody, WM78, which interacts with PZR, a SHP-2 binding partner.

In vitro testing of the leukaemia monoclonal antibody WM-53 labeled with alpha and beta emitting radioisotopes.

We report the preparation and testing of a new alpha emitting radio-immunoconjugate (RIC) against acute myeloid leukaemia (AML) using CD33 positive monoclonal antibody WM-53 (specific for HL-60 cell line). Using cyclic anhydride of diethylenetriaminepentacetic acid (cDTPAa) as chelator, antibody was labeled with 213Bi (alpha), 149Tb (alpha), 153Sm (beta) and 152Tb (positron). In vitro testing showed high labeling efficiency (90-95%) and stability (11-19% leaching) with immunoreactivity virtually the same before and after labeling.

Discrimination of von Willebrands disease (VWD) subtypes: direct comparison of von Willebrand factor:collagen binding assay (VWF:CBA) with monoclonal antibody (MAB) based VWF-capture systems.

Discrimination of von Willebrand's Disease (VWD) subtypes is important since it influences management. Qualitative [ie Type 2A, 2B, 2M] defects exhibit von Willebrand factor (VWF) discordance and give high VWF:Ag to VWF:'activity' ratios. Classically, VWF:'activity' is assessed using the VWF:RCof assay.

Novel molecular mechanisms of dendritic cell-induced T cell activation.

In this study we have re-examined the molecular mechanisms involved in activation of T cells by dendritic cells (DC). Human peripheral blood DC (PBDC) were derived by 2 h adhesion followed by 7 day culture in a combination of granulocyte macrophage colony stimulating factor and IL-4, and depletion of residual T and B cells. These PBDC were used to induce autologous T cell proliferation in a CD3-dependent response, and antibodies against CD11a/18 and CD86 were used as control inhibitors of accessory function.

Use of a novel platelet function analyzer (PFA-100) with high sensitivity to disturbances in von Willebrand factor to screen for von Willebrand's disease and other disorders.

The PFA-100 is a new platelet function analyzer which uses whole blood and high shear stress blood flow to simulate primary hemostasis and assess platelet function. A small volume of blood is introduced into a disposable cartridge, and forced through a capillary tube. Platelet adhesion and aggregation is then initiated following exposure to either collagen/ADP [C/ADP] or collagen/epinephrine [C/Epi] coated membranes.

MUC18, a member of the immunoglobulin superfamily, is expressed on bone marrow fibroblasts and a subset of hematological malignancies.

Despite the importance of bone marrow stromal cells in hemopoiesis, the profile of surface molecule expression is relatively poorly understood. Mice were immunized with cultured human bone marrow stromal cells in order to raise monoclonal antibodies to novel cell surface molecules, which might be involved in interactions with hemopoietic cells. Three antibodies, WM85, CC9 and EB4 were produced, and were found to identify a 100-110 kDa antigen on bone marrow fibroblasts.

Assessment of the validity of the Adams and Fujita approximation for the higher oligomers of human spectrin.

Analysing the self-association behaviour of human erythrocyte spectrin is complicated by a large degree of nonideality. Adams and Fujita [1] proposed that, as a first order approximation, the logarithm of the activity coefficient of the protomer of a self-associating system can be considered to be linearly dependent on the total concentration of the protein, and that the same second virial coefficient could be considered to apply to all species. As a consequence of the Adams and Fujita approximation, the apparent equilibrium constant is equal to the thermodynamic equilibrium constant.

Reinvestigation of the thermodynamics of spectrin self-association.

The thermodynamics of the self-association reactions of human spectrin have been reinvestigated by means of sedimentation equilibrium over the temperature range 18-40 degrees C. The experimental data were analysed in terms of a cooperative isodesmic model of association. The van't Hoff plot showed that the standard change in enthalpy for the heterodimer-tetramer step was temperature-dependent, leading to an estimate of -8.

A pilot clinical trial of two murine monoclonal antibodies fixing human complement in patients with chronic lymphatic leukaemia.

The use of monoclonal antibodies (MABs) for the therapy of malignant diseases offers the potential advantage of greater target cell specificity, and therefore less toxicity. A major limitation of this therapeutic approach has been the inability of most MABs to kill the cell once bound to the target antigen. We have previously reported the development of two murine IgM MABs, WM63 (CD48) and WM66 (unclustered), that react with panleucocyte antigens widely expressed on cells from lymphoproliferative disorders, and are lytic with human complement.

Characterization of a novel leucocyte surface membrane antigen recognized by the monoclonal antibody WM65.

WM65 is a murine mAb which recognizes a novel surface membrane antigen present on leukaemic and normal leucocytes. The present study further investigates the nature of this antigen, especially those features which relate to the possible therapeutic applications of the WM65 antibody. There are 1-3 x 10(4) molecules of this antigen present on normal leucocytes, and the same or greater numbers of antigen molecules are present on a variety of leukaemic cells.

Purification of two murine monoclonal antibodies of the IgM class by hydroxylapatite chromatography and gel filtration.

A two-step method using hydroxylapatite chromatography and gel filtration is described for the purification of two murine monoclonal antibodies of the IgM class. Ascites fluid from each hybridoma was diluted in sodium phosphate buffer (0.01 M, pH 6.

Reaction coupling of chelation and antigen binding in the calcium ion-dependent antibody binding of cyclic AMP.

Polyclonal antibodies, raised against cyclic AMP (cAMP) by the immunization of animals with a 2'-O-succinyl cAMP/bovine albumin conjugate, have been reported to be dependent upon the presence of calcium ion (Ca2+) for antigen binding. They also exhibit a major "bridge" effect whereby 2'-O-succinyl and 2'-O-acetyl derivatives are bound more avidly than the parent nucleotide. Since cAMP and these derivatives bind Ca2+ very weakly, they do not present substantially in the chelated form over the range of Ca2+ concentrations used.

A novel human leucocyte surface membrane antigen defined by murine monoclonal antibody.

A murine monoclonal antibody has been produced which identifies a novel human leucocyte differentiation antigen. The antibody, designated WM-66, of IgM subclass, was cytolytic with human complement. WM-66 was shown to react with virtually all normal T and B lymphocytes from peripheral blood and lymphoid tissues, as well as blood monocytes and approximately 40% of bone marrow mononuclear cells.

A novel non-lineage antigen on human leucocytes: characterization with two CD-48 monoclonal antibodies.

Two murine monoclonal antibodies have been produced which identify a novel surface antigen expressed on human leucocytes in a non-lineage-restricted distribution. Antibodies WM-63 and WM-68 were derived after immunization of mice with human T-CLL cells and the leukaemic cell line HSB-2. Both antibodies were shown to react with over 90 per cent of normal T and B lymphocytes from peripheral blood and tonsil, and also with monocytes from peripheral blood.

Expression of a novel human pan leucocyte differentiation antigen on leukaemic cells.

A murine monoclonal antibody has been produced which identifies a novel surface membrane antigen present on virtually all normal human leucocytes and leukaemic cells. The antibody, designated WM-65, reacted with over 95% of peripheral blood, tonsil and thymic lymphocytes, and with a similar proportion of monocytes and granulocytes. A majority of nucleated normal bone marrow cells were also reactive with WM-65; however, these included only a small proportion of myeloid progenitor cells.

Further studies on the heterogeneity of antigens recognised by CD-1 monoclonal antibodies: distribution of epitopes and analysis of serological binding patterns.

The binding patterns of 28 monoclonal antibodies (MAB) recognizing antigens belonging to Cluster of Differentiation One (CD-1) were analyzed in order to investigate heterogeneity within this cluster. Competitive binding assays using radiolabelled MAB provided detailed information on CD-1 antigenic heterogeneity, and demonstrated that at lease six epitopic regions are recognised as CD-1 MAB. Further studies, based on single cell suspension immunofluorescence assays (using thymocytes and subclones on the cell line Molt-4), suggested that the majority of MAB studied could be serologically separated into three groups.

The effect of o,p'-DDD on adrenal steroid replacement therapy requirements.

Two patients with adrenal carcinoma treated with 2,2-bis (2-chlorophenyl-4-chlorophenyl)-1,1-dichloroethane (o,p'-DDD) as adjuvant therapy were studied. Both patients developed hypoadrenalism while on o,p'-DDD and apparently adequate dexamethasone replacement therapy. The hypoadrenalism was overcome by increasing steroid replacement therapy.

The measurement of antithyroglobulin autoantibodies in the presence of thyroglobulin.

An homogeneous phase radioassay (HRA) for antithyroglobulin autoantibodies (TgAb) in serum was investigated. In this method TgAb is allowed to react with 125I-Tg in solution and the immune complexes formed are separated by precipitation with sheep anti-human gammaglobulin. HRA proved to be suitable for the screening of sera prior to thyroglobulin (Tg) radioimmunoassay; being both sensitive, and unaffected by high endogenous levels of Tg.

Venous ACTH sampling in Cushing's syndrome.

A case of Cushing's syndrome in a 36-year-old woman, in whom the corticosteroid levels remained elevated in peripheral blood despite high doses of dexamethasone (8 mg/day), is reported. Although, on her skull X-ray film, the pituitary fossa was seen to be asymmetrically enlarged and had signs of possible erosion and depression of the floor on the left, the question whether this was an anatomical variant was raised. The correct diagnosis was made after measuring ACTH levels in venous blood samples taken from multiple sites, including the left petrosal sinus, in which a significant elevation of ACTH level, compared with the level in peripheral blood, was found.

The prenatal determination of fetal sex: amniotic fluid testosterone as a preliminary screening test.

Amniotic fluid testosterone levels were measured by radioimmunoassay without chromatography on 101 specimens obtained at amniocenteses between 15 and 19 wk gestation. For the male fetus, the amniotic fluid testosterone level of 553 +/- 23 pmol/l (mean +/- SE) was significantly higher (P less than 0.0005) than the concentration found for the female fetus (206 +/- 9 pmol/l).