Bio plex magpix multiple reader system

R&D system Luminex Kit was used to analyze cytokine's secretion both in sera obtained from BM aspirates of young and aged donors and in conditioned media (CM) derived from young and aged MSC. Sera from 6 BM of young adults and 8 BM of aged subjects were collected from BM aspirates and analyzed prior MSC derivation. For CM collection, MSC from young and aged donors were plated the day before at 4 × 10⁵ cells/well in a 6‐well plate in StemSpan (Stem Cell Technologies) without addition of other factors for 48 hr. Media were then collected, filtered with a 0.22µm PVDF filter (Millipore), and stored at −80°C to use for Luminex or CM experiments.
Customized Luminex plates were obtained to screen for: IL1α, IL8, IL6, MCP1, IL1β, TNF‐α, GROβ, and CCL4. Assays were run as per manufacturers’ instructions with standards and samples in technical duplicates. Data were acquired on a calibrated Bio‐Plex MAGPIX multiple reader system (Bio‐Rad) and visualized with Bio‐Plex manager Software.

R&D system Luminex Kit was used to analyze cytokine’s secretion both in sera obtained from peripheral blood of mice transplanted with wtBRAF and BRAF^V600E-expressing HSPCs and in supernatants derived from wtBRAF and BRAF^V600E-expressing HSPCs in liquid culture. Customized Luminex plates were obtained to screen for: IL1α, IL8, IL6, MCP1, IL1β, TNFα, IFNγ, IL12, GM-CSF, and CCL4. Assays were run as per manufacturers’ instructions with standards and samples in technical duplicates. Data were acquired on a calibrated Bio‐Plex MAGPIX multiple reader system (Bio‐Rad) and visualized with Bio‐Plex manager Software.