Search PubMed⌕ Search

Biomedical subjects

B Norrild

Publications and source records attributed to B Norrild.

At least 73 records · Page 4Linked to original sources

Chicken ovalbumin gene fused to a herpes simplex virus alpha promoter and linked to a thymidine kinase gene is regulated like a viral gene.

We are describing a system for the introduction, selection, and expression of eucaryotic genes in higher eucaryotic cells. The carrier consisted of the herpes simplex virus 1 (HSV-1) tk gene covalently linked to an HSV-1 alpha promoter directed away from the tk gene. In this study we fused to the alpha promoter the 5' transcribed noncoding sequences and the coding sequences of the chicken oviduct ovalbumin gene. Cells converted to the TK+ phenotype with this chimeric fragment produced an ovalbumin precursor which was processed and secreted into the extracellular fluid. The ovalbumin gene utilized the HSV-1 alpha promoter and was regulated as a viral gene inasmuch as inversion of the genomic DNA relative to the alpha promoter resulted in no ovalbumin synthesis, and production of ovalbumin was enhanced after superinfection with HSV-1. Synthesis of ovalbumin was not detected when cDNA was linked to the HSV-1 alpha promoter. The carrier system described in this study is suitable for introduction, selection, and expression of eucaryotic genes whose natural promoter is either weak or requires the presence of regulatory elements which may be absent from undifferentiated cells in culture.

Animals↗

Secretory IgA specific for herpes simplex virus in lacrimal fluid from patients with herpes keratitis--a possible diagnostic parameter.

In the present study a solid-phase radioimmune assay was used for the demonstration of herpes simplex virus-specific IgG and secretory IgA antibodies in the lacrimal fluid from patients with active recurrent herpes keratitis. The method was quantitative and made it possible to test specifically for the production of secretory IgA antibodies produced during an active herpes simplex virus infection. The production of secretory IgA was followed in 2 patients with fresh recurrent lesions. The HSV-specific secretory IgA could be demonstrated during the first 10 days of infection, where the maximal concentration was reached 3-5 days after the first symptoms occurred. The secretory antibodies were locally produced, and it is shown for the first time that herpes virus-specific secretory antibodies were of diagnostic value.

Adult↗

Herpes simplex virus specific secretory IgA in lacrimal fluid during herpes keratitis.

The correct diagnosis of herpes keratitis is very important, as the clinical manifestation can be similar to that caused by immunologic defects. The presence of herpes simplex virus (HSV) specified IgG in lacrimal fluid is not of diagnostic value in cases where the eyes are severely inflamed, because the inflammation leads to impairment of the blood vessels and allows a passive diffusion of IgG from serum into the lacrimal fluid. In the present study we have developed a method for the detection of locally produced HSV-specific antibodies of the secretory IgA class present in lacrimal fluid from patients with active herpes keratitis.

Antibodies, Viral↗

Application of molecular genetics to the design of live herpes simplex virus vaccines.

In principle, several avenues for attenuation of herpes simplex viruses (HSV) are now available. These include intermixing of HSV-1 and HSV-2 genes by recombination, altering the regulation of gene expression and the deletion of viral genes not required for replication of the virus in permissive cells in culture. Results of analyses of HSV-1 x HSV-2 recombinants and of mutants containing a deletion in a gene expressed early in infection showed a loss of virulence when infected by intracerebral route into adult Balb/c mice. In addition, immunization of the mice by intracerebral route with relatively low doses of virus protected the mice from challenge with high doses (3000 LD50) of virulent virus. The application of genetic engineering to the construction of live vaccines is discussed.

Chromosome Deletion↗

Differential immunologic reactivity and processing of glycoproteins gA and gB of herpes simplex virus types 1 and 2 made in Vero and HEp-2 cells.

Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2, respectively) specify five major glycoproteins designated as gA, gB, gC, gD, and gE. Previous studies have shown that gA and gB differ in electrophoretic mobility but not in reactivity with antisera prepared to each of these glycoproteins. Moreover, gA and gB of HSV-1 crossreact in serologic tests with the corresponding glycoproteins of HSV-2. In this paper, we report on the reactivities of gA and gB of HSV-1 and HSV-2 with 24 independently derived monoclonal antibodies reactive with these antigens. Our results show the following: (i) Electrophoretic mobilities of HSV-1 and HSV-2 glycoproteins gA and gB made in HEp-2 cells are significantly less than those made in Vero cells. (ii) All monoclonal antibodies precipitated both gA and gB made in infected HEp-2 cells. These include 12 monoclonal antibodies that neutralized virus and 12 that did not. (iii) HSV-2 glycoproteins gA and gB made in HEp-2 cells contain type-specific domains. However, monoclonal antibodies produced by one clone directed to these domains did not react with glycoproteins made in Vero cells. (iv) Lysates of infected Vero cells contain three lower molecular weight polypeptides that also reacted with monoclonal antibodies directed to glycoproteins gA and gB. These polypeptides are virus specific inasmuch as those specified by HSV-1 differ in electrophoretic mobilities from those specified by HSV-2. these polypeptides are absent in lysates of HEp-2 cells.

Animals↗

Immunological reactivity of herpes simplex virus 1 and 2 polypeptides electrophoretically separated and transferred to diazobenzyloxymethyl paper.

In this paper we report that viral polypeptides from herpes simplex virus 1 (HSV-1) and 2 (HSV-2)-infected cells electrophoretically separated in sodium dodecyl sulfate-polyacrylamide-agarose gels and transferred to diazobenzyloxymethyl paper can react with rabbit hyperimmune sera, both polyvalent and prepared against specific antigens. The polyvalent hyperimmune sera against HSV-1 reacted with 17 HSV-1 polypeptide bands and 8 HSV-2 polypeptide bands. Concordantly, polyvalent sera against HSV-2 reacted with at least 16 HSV-2 polypeptide bands and 8 HSV-1 polypeptide bands. The antisera prepared against the specific antigens reacted with a smaller number of polypeptide bands. Preimmune sera and immune sera did not react with electrophoretically separated polypeptides from infected and uninfected cells, respectively. The immune localization of separated antigens test provides a powerful technique for identification of immunogenic viral polypeptides, especially those which are normally insoluble and therefore unavailable for immunological reactivity in immune precipitation tests.

Animals↗

Participation of three major glycoprotein antigens of herpes simplex virus type 1 early in the infectious cycle as determined by antibody-dependent cell-mediated cytotoxicity.

Tissue culture cells infected with herpes simplex virus type 1 synthesize three major glycoprotein antigens (Ag-11, Ag-8, and Ag-6), which have been characterized by crossed immunoelectrophoresis. The three viral antigens have been identified as a mixture of gA and gB (Ag-11), gD (Ag-8), and gC (Ag-6). Recent findings have shown that antibodies directed to each of the three antigens individually are able to mediate antibody-dependent cell-mediated cytotoxicity when tissue culture cells late in the infectious cycle (18 h postinfection) are used. In this work, antibody-dependent cell-mediated cytotoxicity was applied to study the time postinfection at which the individual viral antigens first made their appearance at the cell surface. All three viral antigens (Ag-11, Ag-8, and Ag-6) could be demonstrated as newly synthesized from 3 to 4 h postinfection, and the quantities of the antigens at the surfaces of the infected cells increased with time postinfection. The use of cycloheximide and ultraviolet-inactivated virus demonstrated that input virus could be detected by antibody-dependent cell-mediated cytotoxicity during the first 2 h postinfection, but the cytotoxicity caused by input virus remained constant with time postinfection. In conclusion, these observations demonstrate the participation of individual herpes simplex virus surface antigens in antibody-dependent cell-mediated cytotoxicity attack on cells early in infection.

Antibodies, Viral↗

Herpes simplex virus glycoproteins: participation of individual herpes simplex virus type 1 glycoprotein antigens in immunocytolysis and their correlation with previously identified glycopolypeptides.

Tissue culture cells infected with herpes simplex type 1 virus express virus-specified glycoprotein antigens on the plasma membrane. Three of these have been previously identified and have been designated as Ag-11, Ag-8, and Ag-6. In the present study, immunoglobulins to each of the antigens were shown to be capable of mediating immunocytolysis in the presence of either complement (antibody-dependent complement-mediated cytotoxicity) or peripheral blood mononuclear cells (antibody-dependent cell-mediated cytotoxicity [ADCC]). Two herpes simplex virus type 1 strains, VR-3 and F, reacted similarly in the ADCC test in the presence of immunoglobulins to Ag-11, Ag-8, and Ag-6 in both infected Chang liver cells and HEp-2 cells. Anti-Ag-6, however, produced a lower ADCC reaction in HEp-2 cells than in Chang liver cells, suggesting differences in the Ag-6 surface expression in, or release from, these cells. Chang liver and HEp-2 cells infected with the MP mutant strain of herpes simplex virus type 1 showed reduced ADCC in the presence of anti-Ag-11 and anti-Ag-8, but no reactivity at all with anti-Ag-6. Crossed immunoelectrophoretic analysis showed that MP-infected cell extracts contain Ag-11 and Ag-8, but lack Ag-6. Polypeptide analysis of herpes simplex virus type 1 strains F, VR-3, and MP showed that Ag-11 consists of the glycoproteins gA and gB, that Ag-8 consists of gD, and that Ag-6 consists of gC. In conclusion, the present study demonstrates that either one of the glycoproteins (gC, gD, and a mixture of gA and gB) can function as a target for immunocytolysis and that the antibody preparation to gC (Ag-6) does not cross-react with any of the other glycoproteins.

Antibody-Dependent Cell Cytotoxicity↗

Immunoelectrophoretic identification and purification of herpes simplex virus antigens released from infected cells in tissue culture.

Proteins released from herpes simplex virus type 1 (HSV-1)-and type 2 (HSV-2)-infected HEp-2 cells have been characterized by the crossed immunoelectrophoretic technique. Both HSV type-common and type-specific antigens were found in the tissue culture medium 24 h after infection. Antigen Ag-6, an HSV-1-specific antigen, was found in high concentration in the medium as compared to other HSV antigens released from HSV-1-infected cells. The HSV-2-specific antigens, Ag-4 and Ag-1, were released in molecular modifications with altered electrophoretic mobility as compared to their cellular counterparts. Purification of HSV antigens was performed by ion-exchange chromatography, and an HSV type-common antigen, Ag-11, and an HSV-2-specific antigen, Ag-4A, were isolated.

Antigens, Viral↗

Production of rabbit antibodies against the viral capsid antigen (VCA) of the Epstein-Barr virus (EBV).

Rabbits were immunized with nuclear or cytoplasmic extracts of the Epstein-Barr virus (EBV)-producing marmoset cell line B95-8. Following extensive absorption with human EBV-negative cells (HeP-2, Ramos and BJAB), sera were obtained that no longer reacted with cellular or serum proteins of human origin, but gave a single precipitin band with extracts of the human EBV-producing line, P3HR-1. Immunofluorescence tests performed with appropriate parallel human serum controls showed that the rabbit serum contained no activity against EBNA + EA - VCA - Raji cells, or against P3HR-1 virus superinfected, cytosine arabinoside-treated Raji cells that contained EBNA and EA, but not VCA. The sera gave a brilliant indirect immunofluorescence reaction with the virus-producing (EBNA+ EA+ VCA+) P3HR-1 lines. Two-color fluorescence tests, performed with a direct TRITC-labelled VCA conjugate and indirect FITC-staining with the rabbit serum, showed that the same cells reacted in both red and green fluorescence, confirming that the rabbit serum was directed specifically against some antigen formed in the virus-producer cells. Since the synthesis of the relevant antigen was prevented by cytosine arabinoside it cannot be EA and must be a late antigen. The morphology and localization of the antigen support the conclusion that the antigen is VCA or some part of the VCA complex.

Animals↗

Crossed immunoelectrophoresis of a herpes simplex virus type 1-specific antigen: immunological and biochemical characterization.

Crossed immunoelectrophoresis of herpes simplex virus (HSV) antigens obtained from infected cells solubilized with the nonionic detergent Triton X-100 has shown that antigen number 6 (Ag-6) is found in preparations from HSV type 1-infected cells but not in HSV type 2 preparations. A monospecific antiserum was produced against Ag-6 by intracutaneous inoculation of the corresponding immunoprecipitate into rabbits. This antiserum neutralized HSV type 1 but not HSV type 2. The polypeptide composition of Ag-6 (molecular weight, 131,000 daltons) was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the immunoprecipitate, using radiolabeled antigen. The multitude of information obtained demonstrates that crossed immunoelectrophoresis can be used preparatively, since specific immunoprecipitates represent a short cut for the attainment of immunoreactive viral protein entities useful for further immunological and biochemical analyses.

Antigens, Viral↗

Identification of a common antigen of herpes simplex virus bovine herpes mammillitis virus, and B virus.

In immunoelectrophoretic analyses one common antigen was demonstrated in antigen preparations from herpes simplex virus types 1- and 2- (HSV-1 and HSV-2), bovine herpes mammillitis (BHM) virus-, and B virus-infected cells solubilized by Triton X-100. The antigen was also demonstrated in solubilized purified HSV-1 and BHM virus. The common antigen was identified as antigen 11 of HSV-1 or HSV-2. Differences were found in the polypeptide composition of the related antigens when isolated from the four different herpesviruses, but a glycopolypeptide with a molecular weight of 125,000 was present in each of the four different antigen preparations, indicating that this polypeptide carried the common antigenic determinants.

Antigens, Viral↗

Crossed immunoelectrophoretic analysis and viral neutralizing activity of five monospecific antisera against five different herpes simplex virus glycoproteins.

Crossed immunoelectrophoresis of Triton X-100 solubilized HSV-infected cells has identified several major HSV glycoprotein antigens in the precipitating profile. Immunization of rabbits with the corresponding precipitates resulted in the production of monospecific antisera against the antigen part of the precipitate. The immunological importance of five different HSV glycoproteins in respect of their representation on the outer virion envelope as targets for neutralizing antibodies was evaluated by comparison of immunoprecipitating activity and neutralizing potency of the corresponding monospecific antisera.

Antibodies, Viral↗