Monocytes were isolated and incubated with GM-CSF and IL-4 to induce dendritic cell differentiation. Pseudotyped viruses and virus-like particles were produced by transient transfection of 293FT cells using TransIT-293 (Mirus). Infections were performed by incubating 105 MDDCs in 96 well U bottom plates in the presence of 8 µg/ml polybrene. Cell surface staining of activation markers was performed 48h after infection. shRNA vectors carrying GFP were transduced into fresh monocytes together with SIVVLP(G) and dendritic cell differentiation was induced. More than 90% of cells were routinely transduced and cells were challenged at day 4 with HDVIRESRFP(G) or other control PAMPs.
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Monocyte-Derived Dendritic Cell Infection
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Cell differentiation
Cells
Dendritic
Dendritic cell
Gm csf
Infection
Mddcs
Monocytes
Pamps
Polybrene
Pseudotyped viruses
Shrna
Transfection
Transient
Vectors
Virus particles
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Variable analysis
independent variables
- Incubation of monocytes with GM-CSF and IL-4 to induce dendritic cell differentiation
- Transient transfection of 293FT cells with pseudotyped viruses and virus-like particles
- Transduction of shRNA vectors carrying GFP into monocytes together with SIVVLP(G) and dendritic cell differentiation induction
- Cell surface staining of activation markers 48h after infection
- Transduction efficiency of cells (more than 90% routinely transduced)
- Cell response to challenge with HDVIRESRFP(G) or other control PAMPs on day 4
- Incubation of 10^5 MDDCs in 96 well U bottom plates in the presence of 8 µg/ml polybrene
- Positive control: Not specified
- Negative control: Not specified
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