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Biomedical subjects

R O'Kennedy

Publications and source records attributed to R O'Kennedy.

At least 19 recordsLinked to original sources
PubMed1992

Rapid and sensitive determination of coumarin and 7-hydroxycoumarin and its glucuronide conjugate in urine and plasma by high-performance liquid chromatography.

A rapid and sensitive high-performance liquid chromatographic method was developed for the analysis of coumarin, 7-hydroxycoumarin and its glucuronide conjugate in urine and plasma. This method was used to monitor the urinary excretion of these compounds following a single oral dose of coumarin (100 mg). This new method gives excellent chromatographic separation and includes an internal standard. The method was validated and shown to be both accurate and precise in the range 0.5-100 micrograms/ml.

Administration, Oral↗
PubMed1992

A binary plasmid system for shuffling combinatorial antibody libraries.

We have used a binary system of replicon-compatible plasmids to test the potential for promiscuous recombination of heavy and light chains within sets of human Fab fragments isolated from combinatorial antibody libraries. Antibody molecules showed a surprising amount of promiscuity in that a particular heavy chain could recombine with multiple light chains with retention of binding to a protein antigen. The degree to which a given heavy chain productively paired with any light chain to bind antigen varied from 43% to 100% and depended strongly on the heavy-chain sequence. Such productive crosses resulted in a set of Fab fragments of similar apparent binding constants, which seemed to differ mainly in the amount of active Fab fragment produced in the bacterial cell. The dominance of the heavy chain in the antibody-antigen interaction was further explored in a set of directed crosses, in which heavy and light chains derived from antigen-specific clones were crossed with nonrelated heavy and light chains. In these crosses, an Fab fragment retained antigen binding only if it contained a heavy chain from an antigen-specific clone. In no case did the light chain confer detectable affinity when paired with indifferent heavy chains. The surprising promiscuity of heavy chains has ramifications for the evaluation of the diversity of combinatorial libraries made against protein antigens and should allow the combination of one such promiscuous heavy chain with an engineered light chain to form an Fab fragment carrying synthetic cofactors to assist in antibody catalysis.

Amino Acid Sequence↗
PubMed1992

Bifunctional antibodies and their potential clinical applications.

Bifunctional antibodies are monovalent, bispecific, antibody-derived molecules. They have been produced by both chemical and biological means. They are thought to have several advantages over monoclonal antibodies in both immunotherapy and immunodiagnosis. Bifunctional antibodies have been shown to be efficient in the targeting of drugs, toxins, radiolabelled haptens and effector cells on to diseased tissues, primarily cancer cells. In addition, bifunctional antibodies have been used to develop novel immunoassays. The full potential of bifunctional antibodies has yet to be realised.

Antibodies↗
PubMed1991

A critical analysis of the use of sialic acid determination in the diagnosis of malignancy.

The measurement of serum sialic acid in sera from individuals with leukemia, breast, gastrointestinal, lung and bladder cancers, brucellosis and manic depression is described. The serum sialic acid levels were also examined in mice bearing Landschutz ascites tumours. The presence of sialic acid-enriched glycopeptides in serum was shown. The results indicate that while sialic acid may be significantly elevated in sera of some tumour-bearing individuals, it is not solely tumour-derived. Its application as a tumour marker is limited and requires critical analysis.

Animals↗
PubMed1991

Use and detection of nonradioactive iodine-labeled antibodies for immunoassay.

Traditionally radioactively labeled iodine has been used as a reporter group for the detection of antibodies in immunoassay. We have recently developed a microassay system for the detection of very low concentrations of iodide which eliminates the need for the use of radiolabeled iodine (O'Kennedy, R. et al. 1989, Anal. Biochem. 179, 138-144). The successful application of this assay for the detection of mouse IgG is described. The performance of this system compared with enzyme immunoassay was evaluated.

Animals↗
PubMed1991

In vitro immunization of human tonsilar lymphocytes: effects of PWM and rIL-6.

We previously described a system for the in vitro immunization of human tonsilar lymphocytes under serum-free conditions in the presence of muramyl dipeptide and rIL-2 (1). Further studies with this system reveal possible roles for PWM and rIL-6 in the immunization process. The addition of these factors led to an increase in the levels of antibody produced in lymphocyte cultures. The time of addition of these factors was found to be important in determining the extent of their effects on the overall yields of antigen-reactive cells.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1991

Antibody engineering: an overview.

We can now isolate and manipulate antibody genes. Mouse antibodies can be humanized, resulting in chimaeric or reshaped antibodies. Antibody engineering is useful in large scale production of antibodies, in production of active antibody fragments, bifunctional, single-domain and catalytic antibodies, and has lead to the production of novel expression systems useful in many other areas. It allows production of new antibody conjugates, e.g. antibody-toxin or antibody-enzyme linked proteins. Engineered antibodies have many potential applications e.g. imaging, therapy and biosensors.

Animals↗
PubMed1990

Investigation of the properties of bovine heart creatine kinase cross-linked with dimethyl suberimidate.

Dimeric bovine heart creatine kinase (EC 2.7.3.2, ATP: creatine N-phosphotransferase) has been cross-linked with the bifunctional reagent dimethyl suberimidate at several concentrations to yield modified enzyme with enhanced stability towards heat denaturation. The degree of thermal stability is dependent on the degree of cross-linking with optimal stabilization occurring when approx. half of all the available amino groups are covalently attached to dimethyl suberimidate. Accelerated storage studies were performed and the results used to predict the storage time of the native and modified enzyme at lower temperatures. The cross-linked derivative was predicted to have a longer shelf-life at 4 degrees C than the native enzyme. Modification caused a reduction in the specific activity of the enzyme. The pH profile was altered following cross-linking, but the Michaelis constants were not changed. The modified enzyme exhibited a marked resistance to the action of some denaturing agents.

Animals↗
PubMed1990

Bifunctional antibodies: concept, production and applications.

Immunoglobulins, or antibodies, are monospecific, bivalent antigen-binding molecules. Bifunctional antibodies are bispecific, with each arm binding to a different antigen, and may be produced by biological or chemical methods. Biological production involves the fusion of two monoclonal antibody-producing hybridomas or of an immunised spleen cell and a hybridoma. The resulting hybrid hybridomas (quadromas or triomas) secrete a mixture of parenteral monoclonal antibodies and bifunctional antibody. In chemical production, the parental monoclonal antibodies can be 'chopped up and reconstituted' to produce the bifunctional antibody only. Bifunctional antibodies have a variety of potential uses. They were originally proposed as an aid to cancer chemotherapy where one of the arms of the antibody would bind to a tumour marker and the other to a drug, toxin, or cytotoxic cell. Functional agents can thus be target directly onto tumour cells, accumulating with higher density, yet with reduced side effects for the patient. Further applications have been proposed involving enzyme immobilization and novel immunoassay techniques. This review describes developments that have taken place in bifunctional antibody technology to date.

Animals↗
PubMed1990

Effects of coumarins, haematoporphyrins and acridine orange on the viability and growth of Landshütz ascites tumour cells, in the presence and absence of photoradiation.

The effects of haematoporphyrin (Hp), haematoporphyrin derivative (HpD), coumarin, 7-hydroxycoumarin, 4-hydroxycoumarin, amino-methyl-coumarin acetic acid and acridine orange, with and without photoradiation, on Landschutz ascites tumour (LAT) cells were examined. Hp, HpD and acridine orange showed a significant effect on LAT cell viability following photoradiation. However, coumarin or its derivatives had no significant effect on LAT cell viability, with or without photoradiation, over 18 h. Coumarin alone had a significant effect on viability of LAT cells, in vitro, over a period of 9 days. This effect was verified by transplantability studies. Coumarin and 7-hydroxycoumarin both exhibited cytostatic effects on LAT cell growth.

Acridine Orange↗
PubMed1990

Application of adsorptive voltammetry to assess the stability of modified alanine aminotransferases.

Alanine aminotransferase has been stabilized by using chemical modification with both bis(imidates) (of varying length) and succinic anhydride. The voltammetric behavior of the native enzyme and its various modified forms has been studied by using both cyclic voltammetry and differential pulse adsorptive voltammetry. A distinctive accumulation pattern was found for each of the stabilized enzymes at the static mercury drop electrode with respect to the native alanine aminotransferase. Adsorptive voltammetry was demonstrated to be a useful technique to assess the extent of chemical modification of this enzyme, which is indirectly related to their stability for use in biotechnological processes. The sue of differential pulse adsorptive voltammetry, after a preconcentration of the enzyme for 300 s at the electrode surface, has yielded a detection limit of 1.0 x 10(-9) M.

Alanine Transaminase↗