Protocol detail

Cytokine Production Assay with TA

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Cytokine production was determined using 96-well cell culture supernatants and commercial ELISA kits for TNF-a (R&D Systems) and IL-6 (Sanquin) following the instructions of the manufacturer. One hour before LPS treatment on day 6 of Mo differentiation^{43 (link)}, the medium was refreshed and 1 nM to 1 μM TA and optionally 1 μM RU486 of the GR antagonist (Sigma, M8046) were added to the medium. LPS (100 ng ml⁻¹) treatment lasted 24 h at which point the supernatant was collected for ELISA.

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Wang C., Nanni L., Novakovic B., Megchelenbrink W., Kuznetsova T., Stunnenberg H.G., Ceri S, & Logie C. (2019). Extensive epigenomic integration of the glucocorticoid response in primary human monocytes and in vitro derived macrophages. Scientific Reports, 9, 2772.

Publication 2019

M8046

Cell culture Cytokine Elisa Ru486 Tnf a

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Variable analysis

independent variables

TA (1 nM to 1 μM)
RU486 (1 μM) of the GR antagonist

dependent variables

Cytokine production (TNF-α and IL-6)

control variables

Medium refreshed one hour before LPS treatment on day 6 of Mo differentiation
LPS (100 ng ml^-1) treatment lasting 24 h

controls

Positive control: LPS (100 ng ml^-1) treatment
Negative control: Not explicitly mentioned

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