ioMicroglia™ -人iPSC derived cells (ab317488)

Flow cytometry analysis of male donor-derived ioMicroglia shows key phenotypic marker expression.

Flow cytometry analysis of day 10 male donor-derived ioMicroglia shows key microglia marker expression of TMEM119, P2RY12, CD14, CD45 and CD11b with a purity of above 95% for CD45, CD11b and CD14, >80% for TMEM119 and CD45, and >70% for TMEM119 and P2RY12. 

Male donor-derived ioMicroglia show key microglia marker expression.

Immunofluorescent staining of day 10 male donor-derived ioMicroglia shows homogenous expression of P2RY12, IBA1 and TREM2, and a typical ramified morphology. DAPI counterstain (blue). Image taken at 10x magnification.

Male donor-derived ioMicroglia show ramified morphology by day 10.

Rapid morphological changes in the cells upon reprogramming, with key ramified morphology first identified by day 4 and continuing through to day 10. Day 1 to 10 post-thawing; 100x magnification; scale bar; 400 µm.

Whole transcriptome analysis demonstrates high lot-to-lot consistency of male donor-derived ioMicroglia.

Bulk RNA sequencing analysis was performed on three independent lots of male donor-derived ioMicroglia at three different time points throughout the reprogramming protocol. Principal component analysis represents the variance in gene expression between the lots and shows the high consistency across each lot at each given time point. Populations of male donor-derived ioMicroglia with equivalent expression profiles can be generated consistently from every vial, allowing confidence in experimental reproducibility.

Day 10 male donor-derived ioMicroglia from three independent lots were incubated with 1 µg/0.33 cm² pHrodo RED labelled E. coli particles for 24 hours +/- cytochalasin D control. Images were acquired every 30 mins on the Incucyte^® looking at red fluorescence and phase contrast. The graph displays the proportion of cells phagocytosing E. coli particles over 24 hours. Three technical replicates were performed per lot. 

Day 10 male donor-derived ioMicroglia from three independent lots were incubated with 1 µg/0.33 cm² pHrodo RED labelled E. coli particles for 24 hours +/- cytochalasin D control. Images were acquired every 30 mins on the Incucyte^® looking at red fluorescence and phase contrast. The graph displays the fluorescence intensity per cell displaying degree of phagocytosis per cell, data from three independent lots. Three technical replicates were performed per lot. 

Day 10 male donor-derived ioMicroglia were incubated with 500 nM AF488 labelled Aβ42 +/- cytochalasin D for 20 hours with images acquired every 30 mins on the Incucyte^® and degree of phagocytosis calculated based on fluorescence. Three technical replicates were performed.

Day 10 male donor-derived ioMicroglia were stimulated with LPS 100 ng/ml and IFNɣ 20 ng/ml for 24 hours or pHrodo RED labelled E. coli particles. Supernatants were harvested and analysed using MSD V-plex Proinflammatory Kit. These cells secrete TNF⍺, IL-6, IL-8, IL-1b, IL-12p70 and IL-10 in response to stimuli. Predominantly producing a pro-inflammatory response. This is consistent across two independent lots. Three technical replicates were performed per lot.