We have identified the Xga antigen, encoded by the XG blood group gene, by employing rabbit polyclonal and mouse monoclonal antibodies raised against a peptide derived from the N-terminal domain of a candidate gene, referred to earlier as PBDX. In indirect haemagglutination assays, these anti-peptide antibodies react with Xg(a+) but not Xg(a-) erythrocytes. In antibody-specific immobilization of antigen (ASIA) and immunoblot assays, the anti-peptide antibodies react with the same molecule as does human anti-Xga. Therefore, by its identity with PBDX, Xga is identified as a cell-surface protein that is 48% homologous to CD99 (previously designated the 12E7 antigen), the product of MIC2 which is tightly linked to XG. PBDX is renamed here XG.

No abstract

The morphology of the bone marrow of 21 dentists who habitually used nitrous oxide in their surgeries was investigated. Exposure to nitrous oxide was measured with an atmospheric sampling device, and each dentist was invited to fill in a questionnaire giving details of medical history, diet, and intake of alcohol. During the trial a full neurological and haematological investigation was carried out and a bone marrow aspirate was examined both morphologically and by the deoxyuridine suppression test. Mean exposures to nitrous oxide ranged from 159 to 4600 parts per million. In all subjects serum vitamin B12 and folate concentrations were within normal limits. Abnormal results of deoxyuridine suppression tests were obtained in three of the 20 dentists tested; two of these three had abnormal white cells in their peripheral blood films.This study provides direct evidence that occupational exposure to nitrous oxide may cause depression of vitamin B12 activity resulting in measurable changes in bone marrow secondary to impaired synthesis of deoxyribonucleic acid.

In aspirin-treated platelets labelled by preincubation with [(3)H]-choline, enhanced release of both [(3)H]-choline and [(3)H]-choline phosphate resulted from stimulation by collagen or thrombin. No such release accompanied stimulation by ADP, platelet-activating factor or adrenaline. Release of [(3)H]-choline phosphate was entirely dependent on aggregate formation whereas release of [(3)H]-choline was reduced but not eliminated, if aggregation was prevented. The properties of [(3)H]-choline and [(3)H]-choline phosphate release indicated that both collagen and thrombin induced activation of phospholipase D in the absence of aggregate formation. Such activation was augmented if aggregate formation occurred. Aggregation induced by these two agonists also caused activation of phosphatidylcholine-specific phospholipase C. These effects were prevented in the presence of staurosporine and could also be induced by addition of a synthetic 1,2-diacylglycerol indicating a role for protein kinase C.

The monoclonal antibody-specific immobilization of erythrocyte antigens (MAIEA) technique is an immunoassay devised primarily for locating blood group antigens on specific red-cell membrane proteins. The assay involves the incubation of intact red cells with two antibodies, one human alloantibody, the other a nonhuman antibody, usually a rodent monoclonal antibody, but polyclonal antibodies of rabbit origin have been utilized. For a positive result, both antibodies must bind to the same membrane protein. The red cells are lysed, the membrane solubilized and the trimolecular complex of two antibodies and membrane protein is captured in a well coated with goat antirodent (or rabbit) immunoglobulin. The immobilized complex is then detected by the use of peroxidase-conjugated goat antihuman (or rodent) immunoglobulin. Negative results, due to mutual blocking between the human and animal antibodies when their epitopes are close together on the same molecule, have permitted a degree of localization of epitopes on some proteins. This has been most effective in the mapping of Cromer blood group system antigens on the complement control protein domains of decay-accelerating factor (DAF, CD55), but has also proved informative in the clustering of antigens on the Lutheran and Kell glycoproteins. MAIEA is an effective tool for the identification of antibodies to Knops-system antigens on complement receptor 1 (CR1, CD35) in immunohaematology reference laboratories. These antibodies are clinically unimportant, but must be identified before they can be ignored for transfusion purposes.

The MAIEA (monoclonal-antibody-specific immobilisation of erythrocyte antigens) assay has recently been developed for the assignment of red cell antigens, recognised by human alloantisera, to particular membrane components of the red cell membrane. This technique detects trimolecular complexes formed by the reaction of a human antibody and a mouse antibody with a particular red cell protein. A positive reaction, in an ELISA-type detection procedure, occurs if the epitopes to the human and mouse antibodies are present on the same membrane component but at different regions. In this report, we show how the MAIEA assay can be used to confirm the relationship between Cromer system antigens and the complement-regulatory protein, decay-accelerating factor (DAF, CD 55). In addition, the location of the antigens along the protein is postulated by using three anti-DAF monoclonal antibodies with specificities to different regions of DAF. Tca and Esa are assigned provisionally to the first short-consensus repeat (SCR), UMC to the second SCR, Dra to the third SCR and Cra, WESa and WESb to the fourth SCR or to the serine/threonine rich region of the DAF protein.

The monoclonal antibody-specific immobilisation of erythrocyte antigens (MAIEA) test has been employed to investigate the Kell system using five monoclonal antibodies which recognise high frequency epitopes on the 93,000-molecular weight Kell glycoprotein: BRIC 18, BRIC 68, BRIC 107, BRIC 203 and 6-22. BRIC 107, which has anti-k-like (KEL2) specificity, identifies a distinct epitope, whilst competitive binding assays suggested that BRIC 203 (anti-Kpbc), BRIC 18, BRIC 68 and 6-22 (anti K14) comprise an overlapping set of epitopes. The MAIEA assay has been very successful in confirming the assignment of most of the Kell and para-Kell antigens to the Kell protein. Due to the competitive nature of the assay and the fact that the monoclonal antibodies bind to different regions, the results also suggest the relative positions of some of the Kell antigens on the Kell protein; these appear to be located in at least five spatially distinct regions.

The biochemical relationship between the red cell antigens Xg^a and the MIC2 gene product, CD99 - previously designated the 12E7 antigen - has been examined by immunoblotting and immunoprecipitation analyses of the protein molecules bearing these antigens. Immunoblotting of membrane components and Xg^a-immunoprecipitates with anti-Xg^a has shown that Xg^a antigen is carried on a broad band of apparent molecular weight (M(r)) 24,500-29,500, which consists of a darkly stained component at M(1) 24,500 and a more diffusely stained component at approximately M(r) 26,500-29,500. Immunoblotting of membrane components and 12E7-immunoprecipitates with 12E7, and RFB-l and NaM123 which also recognise CD99, distinguished two bands of M(1) 30,000 and 32,000. A non-radioactive immunoprecipitation technique was employed, which uses chemiluminescence detection of biotin-labelled red cell proteins. The protein of M(1) 32,000, which carries CD99, was identified by this method and the red cell quantitative polymorphism of CD99 was demonstrated. When the Xg^a protein was precipitated from biotin-labelled red cells, a protein of M(r) 32,000 was coprecipitated. This suggests that the proteins carrying the Xg^a antigen and CD99 are associated in the membrane.