Polypeptide analysis of individual immunoprecipitates from crossed immunoelectrophoresis.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
B Norrild
Publications and source records attributed to B Norrild.
Polyacrylamide gel electrophoretic analysis of herpes simplex virus type 1 immunoprecipitates obtained by quantitative immunoelectrophoresis in antibody-containing agarose gel.
Crossed immunoelectrophoresis was used to characterize herpes simplex virus type 1 (HSV-1) antigens produced by infected HEp-2 cells. We report on a method for analyzing the polypeptide content in individual antigen-antibody precipitates eluted from the second-dimensional agarose gel. Four glycoprotein antigens of HSV-1, Ag-8, Ag-11, Ag-6, and Ag-3, were isolated and analyzed for polypeptide content. The molecular weights of the polypeptides are presented.
Antigens, Viral↗
Crossed immunoelectrophoretic studies of the solubility immunogenicity of herpes simplex virus antigens.
The nonionic detergent Triton X-100 was capable of solubilizing 90% of the protein content in herpes simplex virus (HSV)-infected rabbit cornea cells. The solubilized HSV antigens formed well-characterized precipitates by crossed immunoelectrophoresis in Triton X-100-containing agarose gel, allowing both identification and relative quantitation. Water-soluble and detergent-requiring HSV antigens were identified by different solubilization procedures in buffer with and without detergent. Five glycoprotein antigens were solubilized only in the presence of detergent, indicating their membrane-bound state. One non-glycosylated antigen was present in both a water-soluble and a membrane-bound form. Based upon the crossed immunoelectrophoretic precipitating patterns of Triton X-100-solubilized HSV antigens, it has been estimated that infected cells yield an amount of virus-specific protein equivalent to 2,000 enveloped virions per cell. Rabbits inoculated intracutaneously with Triton X-100-solubilized HSV antigens developed neutralizing antibodies against HSV almost as effectively as rabbits with an active HSV infection. Precipitins against individual HSV antigens in sera from rabbits infected with HSV and immunized with the Triton X-100-solubilized HSV antigens were assayed by the crossed immunoelectrophoretic technique. Sera from infected rabbits reacted more strongly and with a higher number of HSV antigens than sera from immunized rabbits.
Animals↗
Prevalence of human papillomavirus types 11, 16 and 18 in cervical swabs. A study of 1362 pregnant women.
Cervical smears from 1362 pregnant women were examined by filter in situ hybridisation for human papillomavirus (HPV) types 11, 16 and 18. 119 women (8.7%) had HPV-positive smears, HPV 16 being the most common type (61% of all infections). There was a correlation with age (r = 0.63, p = 0.004), the highest incidence found in women less than 22 years old with a decline after the age of 30. The incidence of cervical HPV infection was significantly higher (20.3%, p less than 0.01) in the subgroup of women with past or present vulvar condyloma, but not in women with previous pelvic inflammatory disease or genital herpes. In 18 women with current dysplasia the smears harboured HPV 16, 18, or both in eight cases (40%). The incidence of HPV infection in 71 women with earlier dysplasia did not differ from that of the women who never had dysplasia.
Adolescent↗
Filter in situ hybridisation: an evaluation of the FISH technique for HPV detection in cervical swabs.
The filter in situ hybridisation (FISH) method for detection of HPV in cervical swabs was evaluated against the Southern blot technique on concomitant cervical biopsies. Of 73 biopsies, HPV 16 DNA sequences were found in 26 biopsies and HPV 18 sequences in 2 biopsies. Analysis by FISH of the corresponding smears detected 58 and 100% of these, respectively. Of the smears corresponding to the HPV-negative biopsies, 17% were HPV 16-positive and 3% were HPV-18 positive by FISH. Re-hybridisation with cold plasmid added for competition did not change these results. To estimate the risk of spurious hybridisation between vector remnants in the probe and bacterial DNA sequences present in smears, we have hybridised by FISH to preparations of the 19 most common vaginal microorganisms. Of these, E. coli, which is present in about 10% of cervical smears, hybridised strongly with a probe of the plasmid vector pBR322 and may be a significant cause of false positive FISH results. None of the bacteria hybridised with probes of purified HPV when cold, denatured plasmid was added for competition. Analysis by FISH with probes of purified pBR322 to 167 smears of a patient control group resulted in 6% positive reactions. In hybridisations with probes of HPV 16 and 18 to 2 or 3 different filter preparations of the same smear, identical results were obtained in 18 of 19 smears, indicating a good reproducibility by the FISH method. The high percentage of HPV negative smears is equivalent to the rates known from cytology and may reflect sampling errors.
Biopsy↗
Prevalence of HPV in premalignant and malignant cervical lesions in Greenland and Denmark: PCR and in situ hybridization analysis on archival material.
Archival formalin-fixed paraffin-embedded cervical specimens from 125 women in Greenland and 139 women in Denmark who had CIN I-III or cervical cancer diagnosed between 1983 and 1987 were analysed for human papillomavirus type 16 (HPV-16) by in situ hybridization and for HPV-16, 18, 31, 33, 35 and 45 by PCR. In situ hybridization analysis showed an HPV-16 prevalence of 17% (16/95) and 23% (24/105) in the premalignant lesions from Greenland and Denmark, respectively. The HPV-16 prevalence rate in the cancer specimens was 10% (3/30) in the samples from Greenland and 29% (10/34) in the Danish specimens. A total of 82 Greenlandic and 107 Danish specimens were beta-globin-positive by PCR reaction. HPV-16-specific PCR on these samples showed 63% (34/54) of the Greenlandic and 68% (50/74) of the Danish preinvasive lesions to be positive. The corresponding HPV-16-positive rates for the invasive cancers were 82% (23/28) for Greenland and 70% (23/33) for Denmark. This study of patient samples supports our previous population-based studies in which similar HPV-detection rates were found among random samples of women from Greenland and Denmark, although Greenland has a 4-5-fold higher cervical cancer incidence.
Adult↗
A time-related study of Brefeldin A effects in HSV-1 infected cultured human fibroblasts.
Glycoprotein D (gD-1) is an essential virion envelope component of herpes simplex virus type 1 (HSV-1) normally transported to the plasma membrane of the infected cells. In the present study, the intracellular transport of gD-1 was inhibited in cultured HSV-1 infected human fibroblasts by Brefeldin A (BFA) 1 microgram/ml medium added for 12 h after virus adsorption. Immunofluorescence light- and confocal microscopy revealed abolished transport of gD-1 to the plasma membrane, juxtanuclear accumulation of gD-1, and a disorderly arrangement of the tubulin fibres. Withdrawal of BFA influence for more than 60 min resulted in incomplete transport but increasing accumulation of gD-1 in the plasma membrane and in Golgi-like areas close to the nuclei. The tubulin pattern was almost normalized 6 h after removal of BFA. The egress of infectious HSV-1 particles released 9 h post-BFA treatment was not fully reestablished. The results indicate that BFA effects were not completely reversible and caused a sort of cytotoxic influence involving the structure of tubulin.
Antiviral Agents↗
Fused rocket and rocket immunoelectrophoresis adapted for demonstration and quantitation of the A antigen of Marek's disease virus-infected cell cultures.
Preliminary results have shown that the rocket immunoelectrophoresis technique can be used as a method for quantitation of the A antigen of Marek's Disease virus-infected cell cultures.
Animals↗