Journal: PLoS ONE

Article Title: Heterogeneity in Macrophage Phagocytosis of Staphylococcus aureus Strains: High-Throughput Scanning Cytometry-Based Analysis

doi: 10.1371/journal.pone.0006209

Figure Lengend Snippet: Adherent GM-MФs were either treated with cytochalasin D (to block uptake) or control prior to and during incubation with GFP- S. aureus RN6390 and incubated with lysostaphin (to degrade external S. aureus , eliminating external bacteria) or control following incubation with S. aureus . External S. aureus were labeled with an IgG 3 monoclonal mouse anti- S. aureus primary antibody and Texas-RedX-conjugated goat anti-mouse secondary antibody. (A) No treatment. (B) Treatment with cytochalasin D. (C) Treatment with lysostaphin. (D) Treatment with both cytochalasin D and lysostaphin. (Original magnification, 200X).

Article Snippet: To achieve red fluorescent labeling of external bacteria cells were incubated for 15 minutes with PBS/4% FBS (blocking buffer), incubated for 40 minutes with 10 µg/ml mouse monoclonal IgG 3 anti- S. aureus antibody in blocking buffer, washed three times with blocking buffer, incubated for 20 minutes with 40 µg/ml Texas-RedX-conjugated goat anti-mouse IgG (Life Technologies) in blocking buffer, and washed twice with blocking buffer.

Techniques: Blocking Assay, Incubation, Labeling

Journal: PLoS ONE

Article Title: Heterogeneity in Macrophage Phagocytosis of Staphylococcus aureus Strains: High-Throughput Scanning Cytometry-Based Analysis

doi: 10.1371/journal.pone.0006209

Figure Lengend Snippet: Adherent GM-MФs were incubated with unopsonized GFP- S. aureus RN6390. External S. aureus were labeled with an IgG 3 monoclonal mouse anti- S. aureus primary antibody and Texas-RedX-conjugated goat anti-mouse secondary antibody. Collapsed confocal stack images were acquired by scanning cytometry. (A) CellTracker Blue and Hoechst channel (cells). (B) GFP channel (all bacteria). (C) Texas Red channel (external bacteria). (D) Composite image. (Original magnification, 200X).

Article Snippet: To achieve red fluorescent labeling of external bacteria cells were incubated for 15 minutes with PBS/4% FBS (blocking buffer), incubated for 40 minutes with 10 µg/ml mouse monoclonal IgG 3 anti- S. aureus antibody in blocking buffer, washed three times with blocking buffer, incubated for 20 minutes with 40 µg/ml Texas-RedX-conjugated goat anti-mouse IgG (Life Technologies) in blocking buffer, and washed twice with blocking buffer.

Techniques: Incubation, Labeling, Cytometry