Comparison of adenoviral hexon polypeptides (monomers) and of native hexons (trimers) by SDS-polyacrylamide gel electrophoresis.

Purified hexons of 27 serotypes of human, simian, bovine and avian adenoviruses were analysed by SDS-PAGE. The apparent molecular weights of hexon polypeptides calculated by comparison with 5 non-hexon and 3 sequenced hexon polypeptide markers ranged from 98 kDa (for bovine adenovirus Ad bos7) to 118 kDa (for simian adenovirus Ad sim13; SV36). A stability of native hexon capsomers (trimers) in SDS at room temperature permitted us to resolve native (trimeric) hexon by SDS-PAGE and to distinguish them from denatured (monomeric) hexon polypeptides by electrophoretic mobilities. Hexon trimer bands with slow mobility in SDS-PAGE (unlike hexon monomer polypeptide bands) retained native hexon antigenicity as revealed by immunoblot analyses. Possible applications of simultaneous analyses of hexon trimers and monomers by SDS-PAGE are discussed.

Characterization of a series of monoclonal antibodies specific for bovine adenovirus serotype BAV3 hexon antigen and their use in an ELISA for detection of adenoviral hexons.

After immunization of mice with purified hexon (the main capsid antigen) of bovine adenovirus serotype BAV3 we have obtained a set of 16 individual hybridoma clones producing MAb's against BAV3 hexon. All MAb's were shown to belong to immunoglobulin G class. Specificity of the most avid MAb marked B3Hx-1 was tested on a panel of representative hexon antigens from 16 adenovirus serotypes of human and animal origin using several immunoassays. In Western blot analysis the MAb B3Hx-1 reacted only with native (trimeric) form of hexon protein and not with denaturated hexon polypeptide chains. The epitope defined by B3Hx-1 appeared stable against SDS at ambient temperature and against chloramine-promoted iodination. The specificity of the epitope was characterized as almost genus-crossreactive: it was absent from hexons of avian and of bovine subgroup 2 adenovirus serotypes and present in most hexons of bovine, canine, simian and human adenoviruses tested. Within the latter group its expression was weak or absent only for human subgenus C serotypes. Several variants of sandwich-type ELISA were developed using MAb B3Hx-1 and different polyclonal antibodies against hexons of mammalian adenoviruses. The level of hexon detection for different adenovirus serotypes varied in range 10(-9) to 10(-8) g per ml.

[Immunoenzyme analysis of adenovirus hexon. Determination of antibodies from hyperimmunized chickens]. / Immunfermentnyi analiz geksona adenovirusov. Opredelenie antitel ot giperimmunizirovannykh kur.

We have elaborated three systems of enzyme-linked immunosorbent assay (ELISA) for detection of chicken IgG antibodies specific for hexon antigens of three immunologically distinct adenovirus groups: those of mammalian adenoviruses (Mastadenovira), typical avian adenoviruses (Aviadenovira) and of egg-drop syndrome-76 (EDS-76) virus. In each system the antibodies against respective hexons were specifically detected. In mammalian adenovirus hexons the ELISA detects primarily the type-specific (epsilon) and genus-specific (alpha) antigenic determinants. The time course of anti-hexon antibodies content was followed during immunization. The level of anti-hexon antibodies in egg yolk reflects adequately their content in blood serum. The technique is suitable for serological diagnosis of chicken adenoviral infections as well as for characterization of egg-yolk antibodies obtained by preparative hyperimmunization of hens.

[Immunoenzyme analysis of adenovirus hexons. Indication and characteristics of rabbit and mouse antibodies against hexons]. / Immunofermentnyi analiz geksona adenovirusov. Indikatsiia i kharakteristika krolich'ikh i myshinykh antitel protiv geksonov.

An enzyme-linked immunosorbent assay (ELISA) was developed for analysis of rabbit and mouse IgG antibodies specific to adenoviral hexon. The anti-hexon antibodies were detected by capture with purified hexon coated onto polystyrene microtiter plates and visualizing them by respective anti-IgG horseradish peroxidase conjugates. In the sera from hyperimmunized rabbits and mice as well as in the mouse ascite fluids the ELISA procedure revealed primarily type-specific (epsilon) and genus-specific (alpha) antigenic determinants in hexon but not those of intermediate specificities.

[Immunoenzyme analysis of adenovirus hexon: serodiagnosis in human eye diseases]. / Immunofermentnyi analiz geksona adenovirusov: serodiagnostika pri glaznykh zabolevaniiakh cheloveka.

The enzyme-linked immunosorbent assay (ELISA) was adapted for detection of IgG antibodies against adenovirus hexon in the sera and tears of patients with eye diseases. The ELISA procedure is more sensitive than complement fixation test for the detection of antihexon antibodies; it allows the seroconversion to be seen in the course of infection and convalescent period. The ELISA procedure has been used for testing specimens from a group of 66 patients with keratoconjunctivitis of adenoviral etiology.