title: Flow cytometry based identification of simian immunodeficiency virus Env-specific B lymphocytes.
abstract: SIV infection of macaques is the most widely employed model for preclinical AIDS vaccine and pathogenesis research. In macaques, high-titer virus-specific antibodies are induced by infection, and antibody responses can drive evolution of viral escape variants. However, neutralizing antibodies (Nabs) induced in response to SIVmac239 and SIVmac251 infection or immunization are generally undetectable or of low titer, and the identification and cloning of potent Nabs from SIVmac-infected macaques remains elusive. Based on recent advances in labeling HIV-specific B lymphocytes [1-3], we have generated recombinant, secreted, soluble SIVmac envelope (Env) proteins (gp120 and gp140) for detection and quantification of SIVmac Env-specific B lymphocytes. In contrast to HIV-1, we found that soluble SIVmac239 gp140 retains the ability to form stable oligomers without the necessity for introducing additional, stabilizing modifications. Soluble oligomeric gp140 reacted with rhesus anti-SIV Env-specific monoclonal antibodies (MAbs), and was used to deplete Env-specific antibodies with SIV neutralization capability from plasma taken from a rhesus macaque immunized with live attenuated SIVmac239∆nef. Soluble gp120 and gp140 bound to SIV-specific immortalized B cells, and to SIV Env-specific B lymphocytes in peripheral blood of immunized animals. These reagents will be useful for analyzing development of Env-specific B cell responses in preclinical studies using SIV-infected or vaccinated rhesus macaques.
authors.affiliation: Microbiology Division, New England Primate Research Center, Harvard Medical School, One Pine Hill Drive, Southborough, MA 01772, USA.
authors.forename: Ismael Ben F
authors.initials: IB
authors.lastname: Fofana
authors.forename: Arnaud D
authors.initials: AD
authors.lastname: Colantonio
authors.forename: R Keith
authors.initials: RK
authors.lastname: Reeves
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Connole
authors.forename: Jacqueline M
authors.initials: JM
authors.lastname: Gillis
authors.forename: Laura R
authors.initials: LR
authors.lastname: Hall
authors.forename: Shuji
authors.initials: S
authors.lastname: Sato
authors.forename: Craig R
authors.initials: CR
authors.lastname: Audin
authors.forename: David T
authors.initials: DT
authors.lastname: Evans
authors.forename: Hisatoshi
authors.initials: H
authors.lastname: Shida
authors.forename: R Paul
authors.initials: RP
authors.lastname: Johnson
authors.forename: Welkin E
authors.initials: WE
authors.lastname: Johnson
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology / Q000821:virology*; D002460:Cell Line; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D008253:Macaca mulatta; D008562:Membrane Glycoproteins / Q000032:analysis* / Q000276:immunology; D015302:Simian Immunodeficiency Virus / Q000276:immunology*; D014759:Viral Envelope Proteins / Q000032:analysis* / Q000276:immunology
chemical_list: D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins; C069623:gp120 protein, Simian immunodeficiency virus