Enriching for HIV-infected cells using anti-gp41 antibodies indirectly conjugated to magnetic microbeads.
The isolation of a pure population of human immunodeficiency virus (HIV)-infected cells is highly desirable for evaluating the impact of HIV on cellular gene expression. Given that HIV gp41 transmembrane protein is anchored on the surface of HIV-infected cells, we evaluated the use of pooled anti-gp41 monoclonal antibodies (MAbs) and HIV immunoglobulins (HIV-Igs) indirectly conjugated to magnetic microbeads to positively select for infected cells. We demonstrate that pooled anti-gp41 monoclonal antibodies enriched for H9 cells infected with HIV IIIB by approximately 98%. Peripheral blood mononuclear cells (PBMCs) infected with a primary (HIV strain 302151) or laboratory-adapted (IIIB) strain were enriched by 54%-62%, depending on the initial viral inoculum. Using HIV-Ig in this magnetic positive-selection approach was also highly efficient for enriching for H9 cells infected with IIIB but less efficient for infected PBMCs. Both types of antibodies used in the selection process resulted in > 80 viability of selected HIV-infected cells. Analysis of interleukin 2 (IL-2) mRNA expression using real-time reverse transcription PCR (RT-PCR) of the HIV-enriched population demonstrated a higher level of IL-2 mRNA, by approximately four cycles, and an 8-fold increase in IL-2 expression, as evaluated by intracellular staining and flow cytometric analysis, in comparison to gp41-negative cells. Collectively, these data illustrate that antibodies targeting gp41 can be used to enrich for HIV-positive populations. This represents a novel approach for studying the impact of HIV on infected cells and on bystander/uninfected cells.
Bahbouhi
Bouchaib
B
Rush University Medical Center, Chicago, IL, USA.
al-Harthi
Lena
L
D005434:Flow Cytometry / Q000379:methods*; D006678:HIV / Q000276:immunology / Q000502:physiology*; D015700:HIV Envelope Protein gp41 / Q000276:immunology*; D018189:Immunomagnetic Separation / Q000379:methods*; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000473:pathology* / Q000821:virology*
D015700:HIV Envelope Protein gp41