High-Dose Mannose-Binding Lectin Therapy for Ebola Virus Infection

Michelow, Ian C.; Lear, Calli; Scully, Corinne; Prugar, Laura I.; Longley, Clifford B.; Yantosca, L. Michael; Ji, Xin; Karpel, Marshall; Brudner, Matthew; Takahashi, Kazue; Spear, Gregory T.; Ezekowitz, R. Alan B.; Schmidt, Emmett V.; Olinger, Gene G.

Mice were assessed daily for changes in physical appearance and weight. Viremia was assessed by reverse transcription-polymerase chain reaction (RT-PCR) and plaque assays as described elsewhere [11] , and anti-Ebola virus antibodies were measured using standard enzymelinked immunosorbent assays (ELISAs) [12] . Standard blood counts were evaluated with a Coulter A C ÁT diff (Beckman Coulter). For analysis with flow cytometry, spleens were ground into single cell suspensions with the BD Medimachine tissue grinder. After incubation with Fc Block (BD), cells were washed and incubated with antibody (CD3 FITC BD no. 555274, CD8 V450 BD no. 560469, CD14 PerCP eBio no. 45-0141, CD4 PE eBio no. 12-0041-82, CD11b APC BD no. 553312, and CD19 PE-Cy7 BD no. 557655). Cells were washed with PBS and fixed in BD cytofix. Data were immediately acquired with a BD FACSCantoII and analyzed with FlowJo (version 7). The Bio-Plex Mouse Cytokine 23-Plex Panel assay (Bio-Rad 171-F11241) was used to measure multiple cytokines, chemokines, and growth factors in serum and tissue supernatants according to the manufacturer's instructions. Mice that survived the initial infection were tested for Ebola-specific serological response on day 21 and rechallenged with the same virus dose without further treatment, and antibody titers were retested 28 days later.