title: Barnase-barstar high affinity interaction phenomenon as the base for the heterogenous bioluminescence pseudorabies virus' immunoassay.
abstract: The effective new variant of "sandwich" bioluminescent enzyme immunoassay (BEIA) for the sensitive detection of glycoprotein B (gB) of pseudorabies virus (PrV) was presently developed. The high affinity interaction of barnase-barstar protein pair and photoprotein obelin as bioluminescent marker were for the first time successfully applied to BEIA development. Preliminary the two monoclonal antibodies, 11/5 and 34/2, were raised against gB for ELISA PrV detection. Presently we used the same immuno-"sandwich" principle for BEIA. To do this the two different bioconjugates were elaborated. Recombinant barnase was chemically conjugated with monoclonal anti-PrV's gB IgG, and also barstar was fused in frame to obelin. The characteristics of BEIA method have been compared to ELISA PrV detection. We have shown the proposed here gB-BEIA was 40-fold more sensitive as opposed to gB-ELISA test. The construction might have a broad promise in multiple potential immunological applications.
authors.affiliation: Branch of Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, RAS, 142290, Pushchino, Russia.
authors.forename: Natalia V
authors.initials: NV
authors.lastname: Rudenko
authors.forename: Lilia L
authors.initials: LL
authors.lastname: Sinegina
authors.forename: Maxim A
authors.initials: MA
authors.lastname: Arzhanov
authors.forename: Vladimir N
authors.initials: VN
authors.lastname: Ksenzenko
authors.forename: Tatiana V
authors.initials: TV
authors.lastname: Ivashina
authors.forename: Oleg S
authors.initials: OS
authors.lastname: Morenkov
authors.forename: Lyubov A
authors.initials: LA
authors.lastname: Shaloiko
authors.forename: Leonid M
authors.initials: LM
authors.lastname: Vinokurov
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002460:Cell Line; D006224:Cricetinae; D011558:Herpesvirus 1, Suid / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D007118:Immunoassay / Q000379:methods; D049449:Luminescence; D008164:Luminescent Proteins / Q000235:genetics; D016366:Open Reading Frames; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D008164:Luminescent Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D011994:Recombinant Proteins; D014764:Viral Proteins; C010497:obelin; C057438:barstar protein, Bacillus amyloliquefaciens