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Biomedical subjects

V van Heyningen

Publications and source records attributed to V van Heyningen.

At least 109 records · Page 6Linked to original sources
PubMed1986

Molecular and physical arrangements of human DNA in HRAS1-selected, chromosome-mediated transfectants.

We used mitotic chromosomes isolated from a human EJ bladder carcinoma cell line for morphological transformation of mouse C127 cells. These chromosome-mediated transformants were analyzed for cotransfer of markers syntenic with c-Ha-ras-1 on human chromosome 11. We also used cloned, dispersed human DNA repeats, in a general mapping strategy, to quantitate the amounts and molecular state of human DNA transferred along with the activated c-Ha-ras-1 gene. In situ hybridization was used to visualize the physical state of the transfected human chromatin. The combined use of these various techniques revealed the occurrence of both chromosomal and DNA rearrangements. However, our analysis also demonstrated that, in general, very substantial lengths of DNA are transferred intact. Closely linked markers are likely to cosegregate. Therefore, these transformants should be invaluable sources for the complete molecular cloning of isolated fragments of the short arm of human chromosome 11.

Cell Line↗
PubMed1985

Prenatal diagnosis of neural-tube defects with a monoclonal antibody specific for acetylcholinesterase.

An immunoassay for acetylcholinesterase (AChE), based on a monoclonal antibody (AE-2), gave the following results when applied to a panel of amniotic fluids: (a) among 651 samples with normal outcome and normal alphafetoprotein (AFP) values there were 2 (0.31%) false positives; (b) of 9 samples with normal outcome and raised AFP values 1 had a raised AChE titre; (c) all 48 samples from anencephaly cases had raised AChE values; (d) among 49 samples from open spina bifida cases (2 of which had normal AFP values), 48 had raised AChE titres. It is suggested that a monoclonal-antibody-based immunoassay may displace polyacrylamide gel electrophoretic analysis of AChE as a complementary test to AFP in prenatal diagnosis of neural-tube defects, since it is a quantitative test largely independent of operator skill and experience.

Acetylcholinesterase↗
PubMed1985

Molecular analysis of chromosome 11 deletions in aniridia-Wilms tumor syndrome.

We describe five individuals who have constitutional deletions of the short arm of one chromosome 11, including all or part of the band p13. All of these individuals suffer from aniridia; two have had a Wilms tumor removed. We have established lymphoblastoid cell lines from these and in three cases constructed somatic cell hybrids containing the deleted chromosome 11. Analysis of DNA from the cell lines and hybrids with a cloned cDNA probe has shown that the catalase gene is deleted in four of five patients. The catalase locus must be proximal to the Wilms and aniridia-related loci. We have not detected a deletion of the beta-globin or calcitonin genes in any of these individuals; we conclude these genes are likely to be outside the region 11p12-11p15.4. In addition, we have used monoclonal antibodies in fluorescence-activated cell sorting analysis to measure expression in the hybrids of two cell surface markers encoded by genes that map to the short arm of chromosome 11. The genes for both of these are deleted in two individuals but are present in the individual with the smallest deletion.

Antigens, Surface↗
PubMed1984

A screening assay for monoclonal antibodies based on the availability of a polyclonal antiserum.

A double-sandwich enzyme-linked immunospecific assay (ELISA) has been developed for screening hybridoma clones for monoclonal antibodies against human alpha-fetoprotein (AFP). The assay uses commercial polyclonal antiserum to bind alpha-fetoprotein from biological fluids, and the complex then binds the specific monoclonal antibodies in hybridoma culture supernatants. The sandwich is completed with commercial peroxidase-linked anti-mouse immunoglobulin. In exploring the variables of this system, it was found that the quality of the polyclonal anti-alpha-fetoprotein antiserum was the most critical factor. The assay functioned with as little as 10 ng alpha-fetoprotein from cord serum or amniotic fluid. A variety of commercial peroxidase-linked second antibodies was satisfactory, and both IgG and IgA class monoclonals were detected. Comparison of the ELISA with a standard radioimmunoassay showed the former to perform very adequately as a general hybridoma screening assay.

Antibodies, Monoclonal↗
PubMed1984

Production of high affinity monoclonal antibodies to deoxycorticosterone.

Monoclonal antibodies to deoxycorticosterone were produced. Mice were immunised with deoxycorticosterone-3-mono-oxime-BSA conjugate and spleen cells were then hybridised with NS1/1Ag4-1 mouse myeloma cells using 1500 mol. wt polyethylene glycol. The hybrids were grown in RPMI 1640 medium containing HAT to facilitate selection of positive clones. The clones and subclones were screened by using deoxycorticosterone-3-mono-oxime-[125I]iodohistamine. Dextran-coated charcoal was used for separation of antibody bound and free fractions. Two independent clones producing antibody which specifically binds labelled deoxycorticosterone were obtained. Cells from the two best sub-clones were used to raise ascites fluid. Comparison of these antibodies with one of the best conventional antisera previously raised in rabbits showed that the affinity constants were almost comparable (0.49-1.4 X 10(10) l/mol). Cross-reactivity of monoclonal antibodies with cortisol, corticosterone, testosterone and pregnenolone was lower than for polyclonal antisera, but for progesterone the cross-reactivity was similar in both cases. The assay sensitivity obtained with ascites fluid was comparable to that of conventional antibody (2.5 pg/ml). The dilution of ascites fluid which produced 50% binding of the label was 1:4,000,000. These results confirm that it is possible to produce monoclonal antibodies to corticosteroids.

Animals↗
PubMed1982

Enzyme-linked immunospecific assays for human alphafetoprotein using monoclonal antibodies.

Two sandwich-type enzyme-linked immunospecific assays for human alphafetoprotein (AFP) have been developed. In both assays monoclonal antibodies were immobilised by absorption to the wells of microtitre plates and used to trap AFP in the samples offered. Bound AFP was then quantitated by completing the sandwich with peroxidase-conjugated anti AFP, which was either of polyclonal or monoclonal type. The monoclonal to monoclonal sandwich enable AFP measurements to be made in the range 100 to 1000 kU/l. The polyclonal to monoclonal sandwich covered an AFP range of 10 to 1000 kU/l. Both assays were rapid and robust and extremely sparing in the use of antibody.

Antibodies, Monoclonal↗
PubMed1982

Human MHC class II molecules as differentiation markers.

DA6.231 and DA6.164 are mouse monoclonal antibodies that immunoprecipitate HLA-DR-like p34,29 glycoprotein dimers from surface- and metabolically-labeled cells. On lymphoblastoid cell lines the distribution of the 231 epitope is completely nonpolymorphic, while the 164 epitope is present on all cells except on those that are DR7 homozygous. Binding-inhibition studies show that the 231 and 164 epitopes are spatially close to each other when present on the same molecule. The mutual inhibition pattern and the absence of the 164 epitope from the 231+ cells of a few leukemia patients suggest, however, that 231 and 164 epitopes are not invariably present together. Most DR-positive cells possess 231+ and 164+ and 231+ 164- class II molecules in approximately a 2:1 ratio. This has been confirmed by immune depletion studies. Thus DA6.231 appears to define a supralocus epitope. The 164 epitope may be a marker for a subset of class II molecules exhibiting differential expression on various cell types immortalized by malignant transformation.

Antibodies, Monoclonal↗
PubMed1981

Serological distinction between DR antigens and lymphocyte activating determinants.

The Burkitt lymphoma (BL)-derived, HLA-DR antigen positive B cell line, EB1, is a consistently low stimulator in MLC. A rabbit antiserum raised against the strongly stimulating BL line DAUDI, after appropriate absorption with EB1, inhibits MLC stimulation by both B cell lines and allogeneic lymphocytes, whilst lectin-induced proliferation is not significantly affected. Indirect immunofluorescence and 125I-staphylococcal protein A binding to cells pre-incubated with this antiserum suggest that the antigen is present on both peripheral B and T cells, as well as on B lymphoblastoid and myeloma lines. We suggest that this antiserum is directed against lymphocyte activating determinant(s) (LADs) and that these are distinct from the serologically defined DR antigens.

Animals↗
PubMed1980

Assignment to chromosome 16 of a gene necessary for the expression of human mitochondrial glutamate oxaloacetate transaminase (aspartate aminotransferase) (E.C. 2.6.1.1.).

A gene necessary for the expression of human mitochondrial glutamate oxaloacetate transaminase (GOT-2) has been assigned to chromosome 16 on the basis of an immunochemical analysis of human-mouse somatic cell hybrids. Mitochondrial GOT cosegregates with adenine phosphoribosyl transferase (E.C. 2.4.2.7.).

Adenine Phosphoribosyltransferase↗
PubMed1979

Association between pregnancy-associated alpha2-glycoprotein (alpha2-PAG) and mixed leucocyte reaction determinants on the leucocyte surface.

alpha2-PAG is present on the surface on mononuclear blood leucocytes and can be demonstrated predominantly on B-lymphocytes and monocytes. Pretreatment of cells with antibody to alpha2-PAG leads to a marked reduction in Fc-rosette formation. Competitive blocking experiments with specific antisera reveal a particularly close association between alpha2-PAG and MLR (mixed leucocyte reaction) determinants on the cell surface. These findings suggest one mechanism whereby alpha2-PAG may modify cell-mediated immune responses.

B-Lymphocytes↗
PubMed1979

The development of resistance to methotrexate in a mouse melanoma cell line. I. Characterisation of the dihydrofolate reductases and chromosomes in sensitive and resistant cells.

PG19T3 mouse melanoma cells were selected for resistance to methotrexate. Nine sub-lines that are resistant to concentrations of methotrexate ranging from 1.27 x 10(-7) M, to 1 x 10(-4) M methotrexate were selected and characterised in terms of their content of dihydrofolate reductase activity and their chromosomes. The intracellular level of dihydrofolate reductase activity increases with increasing resistance such that at the highest level of resistance PG19T3:MTXR10(-4)M cells contain approximately 1,000 fold more enzyme activity than the parental PG19T3 cells. It is shown that the enhanced activity is due to an increase in the amount of the enzyme rather than any structural change to the enzyme in resistant cells. Comparisons of pH activity profiles, profiles under different activating conditions and titrations with methotrexate suggest that the sensitive and resistant cells contain identical dihydrofolate reductases. Analysis of the chromosomes of resistant cells shows the presence of up to 5 large marker chromosomes which contain homogeneously staining regions after G-banding. These same regions stain intensely after C-banding and fluoresce brightly after staining with Hoechst 33258. The size of homogeneously staining regions increases throughout the process of selection. For one marker chromosome this increase may have been mediated via a ring chromosome.

Animals↗
PubMed1975

Chromosome assignment of some human enzyme loci: mitochondrial malate dehydrogenase to 7, mannosephosphate isomerase and pyruvate kinase to 15 and probably, esterase D to 13.

Eleven independent man-mouse hybrids and 40 subclones from four to them were analysed for up to 42 enzyme markers. Nine subclones from three hybrid lines were fully karyotyped. The data presented suggest that the gene for the human enzyme MOR-M can be assigned to chromosome 7, whilst those for MPI and PK-3 are on chromosome 15. The use of a small number of well-characterized hybrids for gene assigments is discussed as well as the significance of some known human linkage relationships.

Animals↗