MOESM1 of Putative stem cells in the hemolymph and in the intestinal submucosa of the solitary ascidian Styela plicata

Additional file 1: Figure S1. ALDEFLUORTM analyses optimization procedures for excluding doublets and debris, and to evaluate autofluorescence of Styela plicata hemocytes. (A) Gating strategy used to exclude doublets and debris from the analysis. The scatterplots present all data points available. The gate ‘single cells’ (Red) was selected based on area and aspect ratio features (width in relation to total area) and using direct observations of single celled images for further analysis. (B) Scatterplot of the ‘single cells’ gate showing one possible gating strategy to separate ‘small’ and ‘large cells’. (C) Histograms showing the normal frequencies of cells at different intensities of fluorescence in the green channel (Ch02) of the ‘small cells’ and ‘large cells’ gates in the blank control above (i.e., no BAAA), and in cells treated with BAAA (i.e., ALDEFLUORTM); the dotted line indicates the recommended threshold of intensity to consider a positive result. (D) Randomly selected brightfield and fluorescent images of cells of the ‘small cells’ and ‘large cells’ gates of the blank control show complete absence of autofluorescence. (C, D) demonstrate a clear absence of autofluorescence in S. plicata hemocytes.