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Biomedical subjects

D R Harper

Publications and source records attributed to D R Harper.

At least 37 records · Page 2Linked to original sources
PubMed1991

Antibody avidity following varicella-zoster virus infections.

The avidity of IgG antibodies following varicella-zoster virus (VZV) infections was investigated using urea treatment of antigen-bound serum antibody by indirect radioimmunoassay (RIA) and immunoblotting techniques. Sequential sera from 16 patients with varicella and 17 patients with zoster were tested, as well as sera from 80 seropositive individuals without a recent history of VZV disease. Both types of assay showed that low-avidity antibodies predominate early after primary infection, but that antibody avidity increases markedly during convalescence. Using RIA, all sera taken up to 12 weeks after the onset of varicella showed greater than 50% reduction in antibody titre after treatment with 8 M urea but thereafter the proportion of urea resistant antibody increased with time. In contrast, after recurrent infection, high avidity antibodies were found to predominate at all times. Only 6 of 47 sera tested from zoster cases showed greater than 30% reduction after urea treatment and all these were taken within 2 weeks after onset of rash. Immunoblotting also showed that the highly immunogenic p32/p36 nucleoproteins appear to induce predominantly low avidity antibodies, even after recurrent VZV infection. The results of this study indicate that treatment with 8 M urea in RIA for IgG antibodies may be a simple and reliable method for distinguishing primary and anamnestic antibody responses against VZV.

Antibodies, Viral↗
PubMed1991

Nonuniform variation in band pattern with luminol/horseradish peroxidase western blotting.

When optimized, the peroxidase-catalyzed oxidation of luminol can provide a useful, sensitive detection system for Western blotting. However, while the luminescence from intense bands appears rapidly, faint bands require at least 30 min after removal of the membrane from reaction buffer for maximum luminescence to develop. This can result in the detection of a variant band pattern if films are exposed to the blotted membrane too soon after reaction, while exposure later after reaction can result in the preferential detection of faint bands. As a consequence, in order to detect a range of bands similar to that seen using autoradiographic or chromogenic systems, it is necessary to determine the correct time after the initiation of the luminol reaction for film exposure. These effects are due to enhancement of luminescence as a result of the peroxidase-immunoglobulin conjugate binding to a solid phase.

Autoradiography↗
PubMed1990

Antibody responses in recipients of varicella vaccine assayed by immunoblotting.

Sera taken from 35 children with cancer who had been vaccinated with live varicella vaccine were assayed using immunoblotting for the presence of IgG class antibodies to proteins present in varicella-zoster virus (VZV)-infected cells. Sera from 23 of these patients were also assayed for IgM class antibodies. The patterns of immunoreactivity observed for these patients following vaccination were substantially weaker and more variable than those detected following natural VZV infection in otherwise healthy individuals. The IgG responses detected following vaccination involved up to 10 protein bands between 28 and 188 kDa. Bands were particularly frequent in the 78-96 kDa region. IgM responses involved up to 10 bands between 28 and 114 kDa, with the bands in the 78-96 kDa region and at 32-36 kDa being detected most frequently. Repeated vaccination generally produced a stronger IgG antibody response than did single vaccination, and subsequent exposure of vaccinees to natural VZV infection resulted in an increased level of reactivity for IgG antibodies, but not for IgM. Similar reaction patterns were obtained with sera from vaccinees when the vaccine virus and wild-type VZV were used as antigens. Immunoblotting showed good correlation with indirect radioimmunoassay for the detection of a vaccine-induced IgG response.

Antibodies, Viral↗
PubMed1990

Protein blotting: ten years on.

Protein blotting was originally described in 1979 as an outgrowth of nucleic acid techniques, and received its commonly used designation of 'Western' blotting in 1981. The use of the technique to render electrophoresed proteins accessible for further analysis has found many roles, the most prominent being subsequent reaction with antibodies or antisera, which has many clinical and research applications. Since the initial development of the system there have been many changes to the techniques involved, but the basic principles remain unaltered. This review discusses these changes, and also provides a summary of current techniques.

Antibodies↗
PubMed1990

Lipoproteins of varicella-zoster virus.

Human fibroblast cells infected with varicella-zoster virus (VZV) showed a slight increase in lipoprotein synthesis, with the production of two major viral lipoproteins, as detected by radioimmunoprecipitation (RIP). Three bands of Mr 73,000, 90,000 and 97,000 were identified as forms of the VZV gpI glycoprotein. All three incorporated both palmitic and myristic acid, and were shown by thin-layer chromatography to contain myristic, palmitic and stearic acids. A very strong band corresponding to 7,000 Mr, which may represent the product of VZV gene 49, was detected after RIP and in VZV-infected cells, and was shown to contain almost entirely myristic acid. Several minor bands were also detected. The possible functions of the lipoproteins are discussed.

Cell Line↗
PubMed1989

A novel plaque assay system for paramyxoviruses.

A combination of 5% allantoic fluid and 200 micrograms/ml DEAE-dextran with 30 mM MgCl2 used as a supplement to normal overlay medium was found to give large, clear plaques on monolayers of secondary chick embryo fibroblasts infected with avirulent strains of Newcastle disease virus which did not produce plaques without these additions. This modified overlay also allowed plaque assay of Sendai virus strain Z, and avoided any requirement for special cell lines or the use of serum-free medium.

Allantoin↗
PubMed1989

A comparative study of the microbiological contamination of postmortem blood and vitreous humour samples taken for ethanol determination.

Postmortem blood and vitreous humour samples were taken from each of 51 subjects. None of the vitreous humour samples contained large numbers of bacteria or fungi, whereas many micro-organisms were detected in 32 of the blood samples. The results of the microbiological examinations provided useful information for the interpretation of some ethanol levels that might otherwise have been misleading.

Adolescent↗
PubMed1989

IgM and IgG responses to varicella-zoster virus p32/p36 complex after chickenpox and zoster, congenital and subclinical infections, and vaccination.

Varicella-zoster virus (VZV) directs the synthesis of a nonglycosylated polypeptide complex with two prominent species with relative molecular masses of 32,000 and 36,000; the p32/p36 complex is present in both the viral nucleocapsid and the nuclear matrix of the infected cell. We have now further characterized the humoral immune response of VZV-infected humans to this complex and established that it is a major viral antigen. Antibodies to VZV p32/p36 were detected in sera from patients with both primary VZV infection (chickenpox) and VZV reactivation (zoster); the response after zoster was more pronounced. When the IgM and IgG components of the immune response were distinguished, IgM to the viral nucleoproteins was observed following chickenpox and zoster and, to a lesser extent, in recipients of VZV vaccine. An IgM response to VZV p32 also was detected during intrauterine VZV infection and subclinical VZV infection.

Antibodies, Viral↗
PubMed1988

Serological responses in varicella and zoster assayed by immunoblotting.

Acute and convalescent zoster sera taken from 11 patients with varicella and 12 patients with zoster were assayed using immunoblotting for the presence of IgG- and IgM-class antibodies to proteins present in varicella-zoster virus-infected cells. All patients exhibited a detectable virus-specific response with both antibody classes. The IgG responses involved up to 28 protein bands between 28 and 255 kilodaltons (kDa). The reactivity was particularly strong in the 78-114-kDa region, with additional bands observed with all patients at 32, 35, 66, and 220 kDa. This pattern of reactivity typically developed more slowly and was weaker and more variable in patients with varicella compared to those with zoster. The reactivity of IgM antibodies in immunoblotting was similar after varicella and after zoster. Individual sera showed up to 25 bands, with the major reactivity being directed against the 78-96-kDa region and two bands at 32 and 35 kDa. Some differences were apparent between the primary and anamnestic responses with both IgG and IgM antibodies, but this did not allow reliable discrimination of the two types of infection.

Adult↗
PubMed1988

Reduction in immunoreactivity of varicella-zoster virus proteins induced by mycoplasma contamination.

During the study of protein differences between several strains of varicella-zoster virus (VZV), two of the strains were found to be contaminated with Mycoplasma hyorhinis. Polyacrylamide gel electrophoresis showed fourteen extra bands present in MRC5 fibroblasts infected with these strains compared to other strains of VZV. A more striking difference was observed when infected cultures were used as antigens in immunoblotting. Certain viral proteins, corresponding in molecular weight to the viral glycoproteins, showed greatly reduced immunoreactivity. Experimental contamination of a mycoplasma-free strain of VZV with the glucose fermenting M. hyorhinis produced similar effects on immunoreactivity, while contamination with the arginine-hydrolysing M. orale produced no detectable effects. Given these data, it appears likely that the glucose-fermenting species induced significant changes in the VZV glycoproteins, possibly by depletion of sugars or interference in glycosylation pathways. The implications of this are discussed.

Cells, Cultured↗
PubMed1988

Chronic leg ulceration: socio-economic aspects.

Six hundred patients with chronic leg ulcers were assessed and classified with regard to social class, mobility and employment. It has been shown that there is no increased incidence of chronic leg ulceration in the more disadvantaged socio-economic groups, but when leg ulceration occurs it is more likely to be recurrent and take longer to heal. Although in only 11% of patients was mobility affected by leg ulceration alone, the condition interfered with work and or leisure activities to a moderate or severe degree in 42%.

Activities of Daily Living↗
PubMed1987

A controlled trial of weekly ultrasound therapy in chronic leg ulceration.

In a controlled trial to ascertain whether ultrasound given weekly in conjunction with a standard treatment for chronic leg ulcers improves the rate of healing 56 patients were randomised to standard treatment (paste impregnated bandage and a self-adhesive elastic bandage) and 52 to standard treatment plus pulsed ultrasound given weekly. After 12 weeks the proportion of ulcers healed was 20% greater in the ultrasound than in the control group.

Aged↗
PubMed1987

Chronic ulcer of the leg: clinical history.

Six hundred patients with chronic leg ulcers were studied by detailed history and examination as part of a population survey. In 22% ulceration began before the age of 40, and in this group the sex incidence was equal. Over age 40 there was an increasing preponderance of women. Ulcers were significantly more common in the left leg in women but not in men. The site of 26% of ulcers did not include the classical medial goiter area. The median duration of the ulceration at the time of the survey was nine months and 20% had not healed in over two years. The great majority of patients had had recurrence, 66% having had episodes of ulceration for more than five years. Healing of ulcers is a serious problem, but preventing recurrence is the greater challenge.

Adult↗
PubMed1987

Arterial disease in chronic leg ulceration: an underestimated hazard? Lothian and Forth Valley leg ulcer study.

Six hundred patients with chronic leg ulcers were interviewed and examined for evidence of arterial impairment. There were 827 ulcerated legs. Pedal pulses could not be felt in 94 (11%). A Doppler resting pressure index of 0.9 or less was found in 176 legs (21%). Risk factors for arterial impairment included age, ulceration affecting the foot, and a history of claudication, ischaemic heart disease, or cerebrovascular disease. Roughly half the patients with arterial impairment also showed the clinical features of chronic venous insufficiency. Careful assessment for arterial disease is mandatory before patients with chronic leg ulcers are treated with elastic compression.

Age Factors↗
PubMed1986

The effect of staining on the immunoreactivity of nitrocellulose bound proteins.

It was found that Coomassie blue staining of proteins electroblotted onto nitrocellulose resulted in a significant decrease in subsequent immunoreactivity, relative to unstained proteins. This was not uniform, and resulted in distortion of the pattern detected by immunoreaction as well as an overall decrease in sensitivity. Staining with amido black resulted in no such alteration. Additionally, it was observed that amido black is completely leached from the nitrocellulose strip during immunoreaction, whereas no leaching is apparent with Coomassie blue. This may contribute to the observed effect of staining on immunoreactivity.

Antigen-Antibody Reactions↗
PubMed1985

Chronic ulceration of the leg: extent of the problem and provision of care.

A postal survey in two health board areas in Scotland, encompassing a population of about one million, identified 1477 patients with chronic ulcers of the leg. Women outnumbered men by a ratio of 2.8:1. The median age of the women was 74 and of the men 67. Seventy two (5%) were hospital inpatients, 174 (12%) were managed jointly by the primary care team and outpatient departments, and 1201 (83%) were managed entirely in the community. Efforts to improve the management of chronic ulcers of the leg should focus on primary health care.

Adult↗