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Biomedical subjects

L Spencer

Publications and source records attributed to L Spencer.

At least 73 records · Page 4Linked to original sources
PubMed1991

Ethanol euthanasia and its effect on the binding of antibody generated against an immunogenic peptide construct.

Mice were immunised with an immunogenic peptide construct CKNNNSTNSGI coupled to diphtheria toxoid as a carrier. This peptide sequence contains the epitope STNS which is the target of inhibitory monoclonal antibodies directed against the second merozoite surface antigen of Plasmodium falciparum. Antisera raised against the peptide construct were taken using an injection of 70 per cent ethanol or sodium pentobarbitone as methods of euthanasia and these methods compared by determining their effects on the binding specificity of the antibody to the antigen using the immunological criteria of immunofluorescence, immunoblotting criteria of immunofluorescence, immunoblotting and ELISA assays. There was no significant decrease in antibody binding with either sodium pentobarbitone, or ethanol with a final concentration of less than 30 per cent in mouse antisera. Antisera with an added ethanol concentration of 40 to 60 per cent relaxed antibody conformation and this raises the possibility of using the differential effects of ethanol as a tool in mapping antigenic fine structure of a range of antibodies directed against defined epitopes. The cross-reactive response of non-specific antibodies in polyclonal antisera was lowered at the suggested dosage for ethanol euthanasia. Ethanol has immense potential as an alternative method of euthanasia when barbiturate drugs, such as sodium pentobarbitone, are unavailable in specific experimental protocols. This may especially aid research workers in developing countries involved in vaccine development, antibody production and subsequent serological analysis.

Amino Acid Sequence↗
PubMed1991

Comparative immunogenicity of free and carrier-conjugated peptides derived from the constant regions of a polymorphic malarial surface antigen.

One peptide (L7) representing the entire constant N-terminal region, and two peptides (L5 and L6) representing the entire C-terminal constant region of the variable merozoite surface antigen MSA2, were synthesised by solid-state (tBOC) chemistry. Mice were immunised with the peptides alone and conjoined to the carrier protein diphtheria toxoid (DT) using the hetero-bifunctional reagent maleimidocaproyloxysuccinimide (MCS). Immune response was evaluated against the peptide itself by ELISA and against the intact protein MSA2 by immunoblotting and immunofluorescence assay (IFA). Whereas all peptides elicited a strong specific antipeptide response when administered as conjugates only L7 and L6 elicited an anti-peptide response in the absence of carrier. L5 and L7 conjugates elicited sera reactive with the intact MSA2, whereas only L7 elicited such specificity in the absence of carrier.

Amino Acid Sequence↗
PubMed1991

Immunological fine structure of the variable and constant regions of a polymorphic malarial surface antigen from Plasmodium falciparum.

The 51-kDa merozoite surface antigen MSA2 of Plasmodium falciparum shows considerable strain-dependent polymorphism. Although marked sequence variation occurs in the central region of the molecule, the N and C-terminal sequences are highly conserved. A number of monoclonal antibodies directed against MSA2 have been described which inhibit parasite growth in vitro, but these are all directed against variable regions. In an attempt to raise strain independent antibodies we have prepared peptide-diphtheria toxoid (DT) constructs from 36 N-terminal octapeptides spanning the constant region and extending into the variable region of the FCQ/27 PNG variant staggered by one amino acid at either end. Similarly, we prepared 26 C-terminal octapeptides spanning the C-terminal constant region as well as 10 octapeptides from the variable region of the Indochina I variant MSA2. Most of the peptides elicited antipeptide titres in excess of 1/10(4) when administered to mice as peptide-DT adducts emulsified with Freund's complete adjuvant. Only 3 of the 43 N- and C-terminal constant region peptides elicited antibodies which reacted appropriately on immunofluorescence (IFA) or immunoblotting analysis with the intact MSA2 of both strains studied (FCQ/27 and Indochina I), whereas 3 other peptides from the variable region elicited antibodies reactive with the parent MSA2 only. Peptide constructs eliciting antibodies recognising the intact protein corresponded to elements in the cognate sequence of high antigenicity as predicted by the Jameson and Wolf algorithm.

Amino Acid Sequence↗
PubMed1990

Peptide vaccines derived from a malarial surface antigen: effects of dose and adjuvants on immunogenicity.

Peptides P2122 (CKNNNSTNSGI) and P513 (CSQRSTNSAST) containing an epitope of a malarial surface antigen (MSA2) recognised by inhibitory monoclonal antibodies were conjugated to diphtheria toxoid (DT) protein and formulated with various gel-based and water in oil emulsion adjuvants in vaccine trials in mice and rabbits. The P2122-DT construct was effective in raising antibodies reactive with both the immunising peptide and the native antigen. Effective adjuvanticity as measured by the titre of the anti-peptide or anti-protein response in mice varied in the order: Algammulin, Montanide ISA 50 greater than or equal to Freund's adjuvant, Montanide ISA 708, 721, 70 much greater than alum, Squalene Arlacel greater than SAF-1. A similar order of adjuvant efficacy: Freund's greater than alum greater than Squalene Arlacel greater than SAF-1, was observed in rabbits.

Adjuvants, Immunologic↗
PubMed1990

Variable linking region immunogenicity using malarial peptide carrier protein conjugates of defined composition.

Thirty-five overlapping peptides from both conserved and variable parts of the N-terminal region of a malarial merozoite surface antigen (MSA2) were synthesised using solid phase chemistry. All peptides were synthesised with an added N-terminal cysteine and purified by reverse-phase HPLC to facilitate coupling to a carrier protein diphtheria toxoid (DT) using the hetero-bifunctional reagent maleimidocaproloxysuccinimide (MCS). Mice were immunised with these peptide-DT conjugates using Freund's complete adjuvant (FCA). The immune response of these mice was tested against peptide hapten, carrier protein (DT) and the linking region itself, using enzyme-linked immunoassay (ELISA). Although there was great variation in the immune response to each part of the immunogen construct, no significant correlation could be seen between each set of responses.

Amino Acid Sequence↗
PubMed1990

The role of handwashing in prevention of endemic intensive care unit infections.

Handwashing is believed to be the most important means of preventing nosocomial infections. Previous studies of healthcare workers (HCWs) have shown that handwashing practices are poor. No one has shown that handwashing practices can be easily improved and that this prevents endemic (nonepidemic) nosocomial infection. Handwashing and infection rates were studied in two intensive care units (ICUs) of a community teaching hospital. Handwashing rates were monitored secretly throughout the study. After six months of observation, we started interventions to increase handwashing. Handwashing increased gradually, but overall rates before (22.0%) and after (29.9%) interventions were not significantly different (p = .071). Handwashing never occurred before intravenous care, whereas it occurred 67.5% for all other indications (p less than .0001). When questioned, nurses felt they were washing appropriately nearly 90% of the time. Infection rates seemed unrelated to handwashing throughout the study, and no clusters of infection were detected. We conclude that handwashing rates, when measured against arbitrary but reasonable standards, are suboptimal, difficult to change and not closely related to evidence of cross-infection. Further, nurses wash hands selectively, depending on the indication for handwashing, and generally believe they are washing much more frequently than an objective observer believes they are.

Cross Infection↗
PubMed1989

The use of maleimidocaproyloxysuccinimide to prepare malarial peptide carrier immunogens. Immunogenicity of the linking region.

Malarial peptides synthesized with an added N terminal cysteine were conjugated to purified diphtheria toxoid (DT) protein using the bifunctional reagent maleimidocaproyloxysuccinimide (MCS). The molar ratio of peptide to carrier was determined by subtractive sulphydryl titration and confirmed by sodium dodecyl sulphate (SDS) electrophoresis. For enzyme-linked immunoabsorbent (ELISA) analysis of sera from animals immunized with the DT conjugates, peptides were conjugated to bovine serum albumin (BSA) using MCS as well as a glutaraldehyde based coupling procedure. Western blotting analysis shows that both DT and BSA adducts were recognized by monoclonal antibodies (Mabs) directed against the peptide epitopes in the native sequences. Animals immunized with the DT-peptide conjugates produced antibodies to the coupling reagent (MCS) as well as diphtheria toxoid and peptide specific antibodies. This MCS specificity could be largely abolished by pre-incubation of sera with a soluble MCS homologue, a thiosuccinimidocaproylamide (TSC).

Amino Acid Sequence↗
PubMed1989

Clonidine transdermal patches for use in outpatient opiate withdrawal.

We conducted a study to assess the clinical usefulness of transdermal clonidine for heroin detoxification in an outpatient drug treatment clinic. Twenty-two young otherwise healthy heroin addicts participated. Outcome was assessed on the basis of (a) hypotension and other side effects of clonidine; (b) patient retention; (c) concomitant drug use; (d) subjective symptoms of withdrawal; and (e) objective signs of withdrawal. Side effects were present but in no case necessitated discontinuation of treatment. Drop-out rates were equal to conventional treatment offered at the clinic. The availability of clonidine broadens the therapeutic options available to patients and clinicians in treating the opiate-addicted patient. Transdermal clonidine offers several advantages over the oral form: patches can be applied weekly, fewer supplemental medications are required, and patches supply an even blood level of medication. A protocol for use is recommended.

Administration, Cutaneous↗
PubMed1988

Neuropeptide Y, epinephrine and norepinephrine in the paraventricular nucleus: stimulation of feeding and the release of corticosterone, vasopressin and glucose.

The paraventricular nucleus (PVN) is known to have an important function in mediating a variety of behavioral and endocrine responses. In the present study, the responsiveness of the PVN to the effects of the coexisting neurotransmitters, neuropeptide Y (NPY), epinephrine (EPI) and norepinephrine (NE), was examined. Albino rats were each chronically implanted with a swivel brain-cannula that permits chemicals to be infused without disturbing the animals' ongoing behavior. When infused into the PVN, each of these neurotransmitters elicited a reliable feeding response during the first hour after injection. The response to EPI was significantly stronger than that of NE and NPY, while the latency to eat after injection was considerably longer for NPY as compared to the catecholamines. In tests with food absent, each of these substances also increased blood levels of corticosterone (EPI greater than NE = NPY) and vasopressin (NPY greater than EPI greater than NE) and revealed a significant positive correlation between circulating levels of these two hormones. In addition, EPI and NE, in contrast to NPY, caused a simultaneous rise in blood glucose, producing levels that were positively correlated with the hormones. No relationship, however, was detected between these endocrine changes and the rats' feeding-stimulatory responses. Together with other evidence, these results suggest that adrenergic as well as noradrenergic innervation to the PVN has a key role in the behavioral and endocrine systems of this nucleus and, moreover, that NPY generally mimics the effects of these catecholamines in the PVN.

Animals↗
PubMed1987

Ferric chloride-catalyzed activation of hydrogen peroxide for the demethylation of N,N-dimethylaniline, the epoxidation of olefins, and the oxidative cleavage of vicinal diols in acetonitrile: a reaction mimic for cytochrome P-450.

In dry acetonitrile, anhydrous Fe(III)Cl3 catalyzes the demethylation of N,N-dimethylaniline, the epoxidation of olefins, and the oxidative cleavage of 1-phenyl-1,2-ethanediol (and other 1,2-diols) by hydrogen peroxide. For each class of substrate the products closely parallel those that result from their enzymatic oxidation by cytochrome P-450. Because of the close congruence of products, the catalytic nature of the Fe(III)Cl3/H2O2 reaction mimic, and the similarity of the dipolar aprotic solvent (acetonitrile) to the proteinaceous lipid matrix of the biomembrane, the form of the reactive intermediate may be the same in each case. A mechanism is proposed in which an initial Lewis acid-base interaction of Fe(III)Cl3 with H2O2 generates a highly electrophilic Fe(III)-oxene species as the reactive intermediate. This is in contrast to the prevailing view that cytochrome P-450 acts as a redox catalyst to generate an Fe(V)-oxo species or an Fe(IV)-oxo cation radical as the reactive intermediate.

Acetonitriles↗
PubMed1986

Tris(picolinato)manganese(II): a chemical model for the mechanism and function of mitochondrial superoxide dismutase.

The reaction of HO2. with the allylic groups of lipids initiates their peroxidation and auto-oxidation, and probably represents the most serious biological hazard of O2.- -derived species. The presence of tris(picolinato)manganese(II) [MnII(PA)2(PAH)(H2O)], a model complex for mitochondrial superoxide dismutase, (i) efficiently catalyzes the disproportionation of O2.-, (ii) precludes the formation HO2., and thereby (iii) prevents hydrogen abstraction from allylic and thiol groups. Such protection demonstrates that a primary function of superoxide dismutase is to block the formation of HO2., which is the obligatory intermediate for the nonenzymatic proton-induced disproportionation process. This requires that the primary step for the enzyme-O2.- reaction be kinetically favored and dominant relative to the protonation reaction (HA + O2.-).

Benzene↗
PubMed1985

Photosynthetic water oxidation. A new chemical model.

A sequential four-step chemical model for the water oxidation process in photosystem II is presented, based on the observation that a peroxide-linked biquinone complex can be chemically formed as a result of hydroxide ion addition to quinone. In our model, the hydroxide ion intermediate is generated in photosystem II as a result of proton abstraction from water. In the model, the first two flashes of light raise the oxidation state of the bimanganese center, while the third and fourth flashes of light sequentially generate the peroxide-linked biquinone which is then directly oxidized by the bimanganese center to produce oxygen and regenerate quinone.

Benzoquinones↗