[Fusion proteins encoded by orf 129L of ectromelia and orf A30L of smallpox viruses cross-react with neutralizing monoclonal antibodies].

Open reading frame (orf) 129L of ectromelia (EV) and orf A30L of smallpox viruses (SPV) encoding fusion proteins were cloned and expressed in E. coli cells. The recombinant polypeptides (prA30L H pr129L) were purified from cell lysates by Ni-NTA chromatography. Recombinant polypeptides were able to form trimers in buffered saline and they destroyed under treatment with SDS and 2-mercaptoethanol. Reactivity of prA30L, pr129L and orthopoxvirus proteins was analyzed by ELISA and Western blotting with panel of 22 monoclonal antibodies (MAbs) against orthopoxviruses (19 against EV, 2 MAbs against vaccinia virus and 1 Mabs against cowpox virus). This data allowed us to conclude that there are 12 EV-specific epitopes of pr129L and EV fusion proteins, ten orthopox-specific epitopes of EV, VV, CPV fusion proteins, from them 9 orthopox-specific epitopes of prA30L and SPV fusion proteins. Five Mabs, which cross-reacted with orthopox-specific epitopes, were able to neutralize the VV on Vero cells and from them two MAbs has neutralizing activity against smallpox virus. Our findings demonstrate that 129L fusion protein have EV-specific epitopes, that EV 129L and SPV A30L fusion proteins have a several orthopox-specific epitopes to induce a neutralizing antibodies against human pathogenic orthopoxviruses.

[A study of neutralizing activity and cross-reactivity of monoclonal antibodies to ectromelia virus with orthopoxviruses pathogenic for man]. / Neitralizuiushchaia aktivnost' i perekrestnoe reagirovanie monoklonal'nykh antitel k virusu éktromelii s patogennymi dlia cheloveka ortopoksvirusami.

An extensive collection of 125 rat hybridomas secreting monoclonal antibodies (Mabs) to ectromelia virus (EV) polypeptide (Poxviridae family, Orhtopoxvirus genus) was set up. A significant portion of Mabs (37 types) recognized epitopes of the 14 kDa polypeptide as well as the 37 and 35 kDa polypeptides. However, a majority of Mabs interacted with conformation-dependent epitopes, which were destroyed in immunoprecipitation. One hundred and thirteen of Mabs cross-interacted with antigenic determinants of vaccinia viruses (VV), cowpox virus (CPV) and smallpox virus (SPV); only 12 of them were found to be specific to EV. The Mabs antigenic activity was tested for 46 types of cross-reactivity Mabs in VV neutralization on Vero cells. Only the 112H12, 113D5, 113F8, 122H9 and 125G9 Mabs, which were specific to the kDa 14 polypeptide (gene A30L EV), had the neutralizing activity. The 122H9 and 125G9 Mabs were able to neutralize SPV. Therefore, it can be assumed that the 14 kDa polypeptide carries, on its surface, cross-reactivity neutralizing epitopes typical of orthopoxviruses.

[The detection and quantitative determination of antigens to the human immunodeficiency virus types 1 and 2]. / Vyiavlenie i kolichestvennoe opredelenie antigenov virusa immunodefitsita cheloveka tipov 1 i 2.

Three modifications of ELISA test system for HIV antigen detection are described. They are based on IgG from HIV-1 and HIV-2-infected human sera and monoclonal antibodies against HIV-1 p24 used as immunosorbents. The peroxidase/anti-HIV-IgG conjugate was used in all the test systems. A possibility of quantitative detection of viral antigen in native culture fluids, lysates, and purified virus preparations was demonstrated. The test system for HIV-1 antigen detection cannot be used for HIV-2 antigen detection and vice versa. The diagnostic value of HIV-1 p24 antigen detection consists in the possibility of earlier AIDS identification and monitoring of the disease at various stages. The sensitivity of "p24" assay is 0.5 ng/ml.

[Study of antigenic structure of HIV-1 protein p24 using monoclonal antibodies]. / Izuchenie antigennoi struktury belka p24 VICh-1 s pomoshch'iu monoklonal'nykh antitel.

During the experiments 4 murine and 3 rat hybridomas producing monoclonal antibodies (MAb) against the protein p24 of human immunodeficiency virus type 1 (HIV-1) have been obtained. Using the immunoblotting technique, it was established that all the species of MAb reacted with the same viral proteins which are derivatives of gag gene--p24 and p55. The properties of MAb have been studied in competitive binding. Their ability of binding to different fragments of the gag protein produced by the recombinant plasmids in E. coli cells have been investigated in ELISA. The analysis of the findings suggests that the HIV-1 protein p24 contains at least 3 antigenic epitopes. All species of MAb reacted with 3 different HIV-1 strains and 2 HIV-1 isolates, but failed with 2 different HIV-2 strains. The only MAb NS5E4 can be used as an immunosorbent in the antigenic capture reaction.