RESUMEN
We optimized the procedure for the formation of Langmuir films of antibodies based on amphiphilic polyelectrolytes and studied the physicochemical and immunochemical properties of the films obtained. Their immunochemical properties were compared with the immunochemical activity of antibodies in Langmuir films without amphiphilic polyelectrolytes and with antibodies adsorbed on the surface of polystyrene and graphite. The efficiency of immune adsorption by the films based on amphiphilic polyelectrolytes was shown to be greater; the affinity of antibodies and surface concentration of their active conformation depended on the type of amphiphilic polyelectrolytes used to obtain the films. We investigated the structure of these films at the surface of highly oriented pyrolytic graphite using the method of atomic force microscopy. Changes in the structure of the films under study caused by the increase of surface pressure were demonstrated.
Asunto(s)
Anticuerpos/inmunología , Anticuerpos/metabolismo , Electrólitos/metabolismo , Técnicas de Inmunoadsorción , Membranas Artificiales , Polímeros/metabolismo , Técnicas Biosensibles/métodosRESUMEN
A technique for forming Langmuir films from antibodies based on an amphiphilic polyelectrolyte was developed. The physicochemical and immunochemical properties of the Langmuir films obtained were studied. The interaction of HBsAg with the films was found to be described by a model with one binding site, whereas that of HBsAg with antibodies adsorbed on a polystyrene plate, by a model with a positive cooperativity. The use of the novel Langmuir films from antibodies increases the sensitivity of the immunoenzyme assay.
Asunto(s)
Electrólitos/química , Fragmentos Fab de Inmunoglobulinas/química , Membranas Artificiales , Polímeros/química , Tensoactivos/química , Animales , Electrólitos/metabolismo , Cabras , Antígenos de Superficie de la Hepatitis B/química , Antígenos de Superficie de la Hepatitis B/inmunología , Concentración de Iones de Hidrógeno , Fragmentos Fab de Inmunoglobulinas/metabolismo , Metilmetacrilato , Metilmetacrilatos/química , Concentración Osmolar , Polímeros/metabolismo , Propiedades de Superficie , Tensoactivos/metabolismo , TermodinámicaRESUMEN
Atomic force microscopy (AFM) was used for the analysis of rickettsiae and viruses. The specificity of interaction was evaluated on the basis of the adsorption of the analyzed antigen on the polymer-antibody film. The film was formed and transferred onto highly oriented pyrolytic graphite (HOPG) by the method of Langmuir-Schaefer with the use of amphiphilic polymers, alkylated polyethyleneimines. According to the data of AFM, polymer-antibody film was 10-30 nm thick and had ruptures, uneven surface. AFM images of Coxiella burnetii, rotavirus and Venezuelan equine encephalitis virus, immunoadsorbed on the antibody film, were obtained. C.burnetii, due to their size equal to (700-900) x (300-500) x Z = 200 nm, were clearly visible on the underlying surface and could be directly counted. Individual virus particles (60-80 nm) cold not be identified on such surface. To analyze such preparations, the program of image analysis was developed. The program classified the registered image with a certain standard. This program determined the presence of virus antigen on the underlying affinity surface with a high degree of precision.