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Biomedical subjects

D M Goodall

Publications and source records attributed to D M Goodall.

32 records · Page 2Linked to original sources
PubMed1993

Ultrastructural evidence for intramolecular double stranding in iota-carrageenan.

Kinetic studies of primary processes of conformational ordering in gel-forming biopolymers have suggested that a change in mechanism from intermolecular to intramolecular multistrand formation occurs on lowering the concentration of biopolymer. We report here ultrastructural observations consistent with intramolecular double stranding in a carbohydrate polymer, iota-carrageenan, by arresting this process of primary conformational ordering by an ultra-rapid freeze fixation technique. High-resolution transmission electron microscopy (TEM) revealed isolated iota-carrageenan chains showing a range of morphologies (linear, circular, and hairpin) consistent with intramolecular stranding. Control experiments in which iota-carrageenan was frozen in the disordered form revealed longer and thinner strands.

Carbohydrate Conformation↗
PubMed1993

Differential binding of tioconazole enantiomers to hydroxypropyl-beta-cyclodextrin studied by capillary electrophoresis.

Capillary electrophoresis has been used to determine binding constants of tioconazole enantiomers with hydroxypropyl-beta-cyclodextrin (HP-beta-CD), after correcting for changes in mobility with increasing viscosity. The predicted and observed values of enantiomeric mobility difference were found to be maximum at a concentration of HP-beta-CD equal to the reciprocal of the average binding constant.

2-Hydroxypropyl-beta-cyclodextrin↗
PubMed1993

Enantioselective enzymatic biotransformation of 2'-deoxy-3'-thiacytidine (BCH 189) monitored by capillary electrophoresis.

Concentrations of the enantiomers of 2'-deoxy-3'-thiacytidine (BCH 189) have been monitored during enantiospecific biotransformation catalyzed by cytidine deaminase. This enzymatic reaction is monitored by capillary electrophoresis (CE) over a 51-h time period. The use of dimethyl-beta-cyclodextrin as a chiral electrolyte additive in a pH 2.3 phosphate buffer system gives enantiomeric separation with results for peak efficiencies, resolution, and analysis time of the order of 25,000 plates, > 1.5 and 30 min, respectively. The features of the CE method are compared with those of the previously favored chiral HPLC method.

Antiviral Agents↗
PubMed1993

Improved visualization of folded collagen alpha-chains by ultra-rapid freezing.

Transmission electron microscopy techniques are commonly employed to examine the folded polymer structure of collagen polypeptides. These techniques include deposition of a sample by spraying, slow freeze-fixation, air drying and vacuum drying the specimen at room temperature, and using additives such as glycerol in the collagen preparation. Here we report preliminary observations of type I collagen alpha-chains, folded in water, at a concentration of 35 micrograms ml-1 and 10 micrograms ml-1, visualized by an ultra-rapid, freeze-fixation technique designed to minimize structural deformation caused by spraying, additives and poor freeze-fixation. The technique also allows the use of submicrolitre sample volumes of known concentrations with negligible loss and shearing, while at the same time providing excellent contrast to the collagen polymer for electron microscopy. This technique can be employed to study the structure of a wide range of macromolecules (proteins and carbohydrates).

Animals↗
PubMed1992

Evaluation of monoclonal antibodies having specificity for human IgG subclasses: results of the 2nd IUIS/WHO collaborative study.

Following the 1st IUIS/WHO Collaborative Study of monoclonal anti-IgG subclass antibodies, a panel of WHO Specificity Reference Reagents (SRR) was established [Jefferis, R., et al. (1985) Immunol. Lett., 10, 223]. At the time, the hope was expressed that further reagents particularly for IgG2, and other allotypic specificities would become available which could be applied in a wide range of assay protocols. The 2nd study reports the evaluation of nineteen anti-subclass and seven anti-allotype monoclonal antibodies. The anti-IgG1 antibody HP6187 was equivalent in performance to the SRR. Others, that were not of the mouse IgG1 isotype, may be useful for particular applications. The anti-IgG2 antibody HP6200 could be a valuable addition to the WHO SRR; it is specific for an epitope in the Fab region but does not have the light chain bias of HP6014. Antibodies of putative allotype specificity exhibited the claimed specificity when used within protocols similar to those employed by the originating laboratory. It appears to be inherent in the nature of the epitopes (allotopes) recognized that it will take several years before reagents applicable to a wide range of techniques will become available.

Antibodies, Anti-Idiotypic↗
PubMed1991

Assay restriction profiles of three monoclonal antibodies recognizing the G3m(u) allotype. Development of an allotype specific assay.

Three monoclonal antibodies raised against a purified human IgG3 paraprotein were found to exhibit a restriction profile for IgG3/G3m(u) and pan-IgG specificity which was dependent on the assay system. When adapted to an IgG3 subclass capture ELISA, all three McAbs discriminated between paraproteins expressing G3m(u) and antithetical markers G3m(st). One of the antibodies (PNF69C) was selected and conditions were optimised for Gm typing purposes. Using this system G3m(u) could be detected on captured IgG3 derived from human sera. This system may prove useful in the elucidation of Gm allotype profiles.

Animals↗
PubMed1990

A highly conserved conformational idiotope on human IgM rheumatoid factor paraproteins of the Wa cross-reactive idiotype family defined by a monoclonal antibody.

Human IgM rheumatoid factors have been classified into major (Wa) and minor (Po) idiotypically cross-reactive families on the basis of reactivity with polyclonal anti-idiotypic antisera. Extensive structural studies have revealed that RFs from the Wa CRI family have homologous light chains. The structural basis of this CRI however is not known. In this study we have defined a conformational idiotope requiring the association of the VH and VL of a number of RF from the Wa CRI family for its expression and which thus comprise part of the Wa CRI. A murine monoclonal antibody, G8 specific for the idiotope reacted with 67% of RFs from the Wa CRI family. G8 also reacted with one IgG2K paraprotein with unknown specificity suggesting that the idiotope per se does not confer RF reactivity. Comparative studies of G8 expression relative to previously described sequence-dependent heavy and light chain associated idiotopes suggest that G8 recognises a separate determinant. Furthermore, the G8 idiotope is found on RFs expressing the VHI subgroup of heavy chain. Study of G8 expression in polyclonal IgM in normal sera demonstrated that the idiotope is widely expressed in the population. However, significantly higher levels of IgM bearing G8 were detected in the sera of patients with rheumatoid arthritis.

Amino Acid Sequence↗
PubMed1990

Determination of enantiomeric purity of ephedrine and pseudoephedrine by high-performance liquid chromatography with dual optical rotation/UV absorbance detection.

A reversed-phase high-performance liquid chromatography (HPLC) method with dual optical rotation/UV absorbance detection has been developed for the determination of enantiomeric purity of ephedrine hydrochloride and pseudoephedrine hydrochloride using an achiral column. The method gave a correlation coefficient of 0.9997 for the plot of log(optical rotation response) versus log (concentration) over the range of 0.06-10 mg ml(-1) of (+)-ephedrine hydrochloride (20 microliters injection). The limit of detection was 1.0 micrograms. Enantiomeric purity is shown to be most readily determined by measuring optical rotation, alpha, and absorbance, A, responses for standard and unknown samples, and using the equation (alpha/A)u/(alpha/A)s = (2xu - 1)/(2xs - 1), where x is the mole fraction of one of the enantiomers and subscripts s and u refer to standard and unknown, respectively. In blind trials using unknown mixtures of (+)- and (+/-)-ephedrine hydrochloride and a (+)-ephedrine hydrochloride standard, enantiomeric purities were determined to +/- 0.4% (95% confidence level) with five or six replicate 50 micrograms injections. The method has also been applied to the determination of the enantiomer mole fraction of (+)-pseudoephedrine hydrochloride in a cough linctus, giving xu = 0.99 +/- 0.01 with seven replicate injections of 20-fold diluted linctus samples containing 7.5 micrograms of the chiral compound being assayed. Unlike conventional polarimetry, the method does not require chemically-pure samples and can be orders of magnitude more economical in material.

Antitussive Agents↗
PubMed1989

Polarimetric detection in high-performance liquid chromatography.

Chiroptical detection for HPLC is particularly useful as a selective detection method for chiral molecules, and in enantiomeric purity determination with partial chiral separation or without chiral separation. The recent development of laser-based polarimeters with microdegree sensitivity has increased the applicability of optical rotation detection in HPLC. The detection limit of these instruments is submicrogram on-column for many chiral compounds in analytical HPLC. A variety of applications of the selective detection of optically active molecules are reviewed. The use of polarimetric detection with partial chiral separation is considered, both as an aid to method development and for enantiomeric purity determination. Finally applications to enantiomeric purity determination without chiral separation are reviewed, with the dual use of nonchirally selective and chiroptical detectors to determine the total amount and optical purity of the analyte. Determinations of chiral purity for samples of high enantiomeric excess are described, which with laser-based instrumentation may give accuracies of better than +/- 1% with sample loadings of 50 micrograms on an achiral column. Applications to the study of enantioselective reactions are also considered, with determination of enantiomeric excess in near-racemates to better than +/- 0.1%.

Chromatography, High Pressure Liquid↗
PubMed1986

Immunogenic and antigenic epitopes of immunoglobulins. XVII--Monoclonal antibodies reactive with common and restricted idiotopes to the heavy chain of human rheumatoid factors.

Two mouse hybridoma antibodies, G6 and H1, with specificity for idiotypic determinants on the heavy chain of human IgM-RF have been produced. Idiotopes recognized by G6 and H1 were expressed on 5/12 and 2/12 RF paraproteins from the Wa idiotype group respectively, but not on paraproteins lacking rheumatoid factor (RF) activity. Inhibition experiments demonstrated that G6 and H1-specific epitopes were located on (or close to) the antibody binding site of RFs, and measurable on the heavy chain but not their light chains. However, it was concluded that the quaternary structure of the antibody-binding site was important for the optimal expression of both idiotopes. The idiotope recognized by G6 was also detected on polyclonal RFs in unfractionated sera from 8/14 patients with rheumatoid arthritis (RA). The results suggest that the G6 idiotope is a highly conserved determinant on the antibody-binding site of RF. These monoclonal antibodies will allow characterization of RF clonality in RA.

Animals↗
PubMed1986

Immunogenic and antigenic epitopes of immunoglobulins. XIV. Antigenic variants of IgG4 proteins revealed with monoclonal antibodies.

Two IgG4 paraproteins having heavy chains of normal molecular weight are shown to be antigenically distinct in their reactivity profiles with 18 monoclonal antibodies having specificity for the IgG2 or IgG4 subclasses. One protein expresses IgG2 and IgG4 epitopes within the C gamma 2 domain. A second protein is deficient in the expression of IgG4 Fc-specific epitopes. These proteins will be of value in defining the structural basis of Fc effector functions and individual epitope expression.

Antibodies, Monoclonal↗
PubMed1984

Mechanism and dynamics of conformational ordering in xanthan polysaccharide.

The thermally induced order-disorder transition of xanthan (extracellular bacterial polysaccharide from Xanthomonas campestris) has been investigated by optical rotation, differential scanning calorimetry, stopped-flow reaction kinetics and low-angle laser light scattering, and the results have been analysed in terms of Zimm -Bragg helix-coil transition theory. The reciprocal of the transition midpoint temperature (Tm) varies linearly with the logarithm of cation (K+) the salt dependence of Tm, is in agreement with Manning polyelectrolyte theory the ordered structure. The associated increase in cation binding, calculated from the salt dependence of tm, is in agreement with the Manning polyelectrolyte theory for one of the candidate structures from X-ray diffraction, a 5(1) single helix stabilized by packing of side-chains along the polymer backbone, but not for the alternative double-helix structure that has also been proposed. At each salt concentration, the two fundamental parameters of the Zimm -Bragg theory, s and sigma, were calculated. The equilibrium constant for growth of the ordered structure (s) is derived directly from calorimetric measurement of transition enthalpy (delta Hcal ), and sigma, which quantifies the relative instability of the helix nucleus, is derived from the ratio of delta Hcal to the apparent transition enthalpy (delta Happ ) obtained by van't Hoff analysis of the optical rotation data. The temperature course of conformational ordering calculated theoretically is in good quantitative agreement with experimental results from both optical rotation and scanning calorimetry. The calculated average length of stable, ordered chain-sequences increases with decreasing temperature, but equals or exceeds the total chain length from light scattering only at temperatures more than approximately equal to 70 K below Tm, suggesting that ordered and disordered regions may co-exist within the same xanthan molecule. Consistent with this interpretation, the observed rate of conformational ordering increases sharply under conditions where the starting solution for dynamic measurements is partially ordered, suggesting that ordered sequences within each chain may act as helix nuclei for adjacent disordered regions, so that helix growth, rather than the slower nucleation process, becomes rate limiting.

Calorimetry, Differential Scanning↗