Detectability of cytokine and chemokine using ELISA, following sample-inactivation using Triton X-100 or heat.

Clinical samples are routinely inactivated before molecular assays to prevent pathogen transmission. Antibody-based assays are sensitive to changes in analyte conformation, but the impact of inactivation on the analyte detectability has been overlooked. This study assessed the effects of commonly used inactivation-methods, Triton X-100 (0.

Promoter chromatin remodeling of immediate-early genes is mediated through H3 phosphorylation at either serine 28 or 10 by the MSK1 multi-protein complex.

Upon activation of the ERK and p38 MAPK pathways, the MSK1/2-mediated nucleosomal response, including H3 phosphorylation at serine 28 or 10, is coupled with the induction of immediate-early (IE) gene transcription. The outcome of this response, varying with the stimuli and cellular contexts, ranges from neoplastic transformation to neuronal synaptic plasticity. Here, we used sequential co-immunoprecipitation assays and sequential chromatin immunoprecipitation (ChIP) assays on mouse fibroblast 10T1/2 and MSK1 knockdown 10T1/2 cells to show that H3 serine 28 and 10 phosphorylation leads to promoter remodeling.

Functional analysis of the quantitative expression of a costimulatory molecule on dendritic cells using lentiviral vector-mediated RNA interference.

The increasing number of co-stimulatory molecules identified on dendritic cells (DC) to date highlights the complex regulation of co-stimulatory signals in T cell activation. We previously established a single lentiviral vector system to stably express short hairpin RNA (shRNA) to induce RNA interference (RNAi) in cell lines and primary T cells. We reasoned that the choice of shRNA target sequences in the lentiviral vector system would also allow us to regulate different levels of surface expression of a co-stimulatory molecule stably and reproducibly.