Fig. 3: Analyzing cell state dynamics with a 206-plexed SeqIS dataset.

a IMR90 ER:Ras cells were treated with 100 nM 4OHT to induce senescence were picked on days 0, 3, and 6, then used for SeqIS. b PECAbs used for the SeqIS. Blue fill shows relevant features to the indicated antibody. Experiments were performed for 2 times and representative images are shown in (c), analyzed data are shown in (d–j). c SeqIS images of IMR90 ER:Ras cells. Representative 56-plexed images from day 0 and 6 samples are shown. Scale bars, 20 µm. d Left panel, heatmap showing average protein levels. Right panel, UMAP of quantified cells. e Silhouette analysis of the subsampled dataset. Upper panel, silhouette coefficients at each number of antibodies. 25, 50, or 100 antibodies were randomly extracted. Lower panel, box plots indicate the average silhouette coefficient using 25, 50, 100, and 206 antibodies. The analysis was repeated 100 times using different random numbers for each antibody set. Boxes, 25th–75th percentiles; center, median whiskers, 1.5 × interquartile range. f PHATE embedding using senescence-associated proteins and inferred pseudotime. g Histogram of cells in pseudotime. h Dynamics of senescence-associated protein levels in pseudotime. i Dynamics of signal transduction during pseudotime. j Spatial expression patterns of NOTCH1, pATM, and p21 on day 3. The stitched image was divided onto a 50 × 50 grid. Protein levels were averaged for each grid. Source data are provided as a Source Data file.