Fig. 4

Serum antibodies are required for predicting chronological age from peptide array binding data. Furthermore, serum small molecules do not contribute to prediction of chronological age. a Schematic of column size filter. The 30 kDa filter columns can be used to separate serum molecules into flow-through fraction that contains < 30 kDa molecules and filtered fraction that contains > 30 kDa molecules. b Size filter columns are effective at depleting IgG using a 30 kDa filter, as quantified by Coomassie Blue staining. Filtrate (> 30 kDa) produces bright bands for both light and heavy chains. Flow-through (< 30 kDa) is depleted for heavy and light chain; however lower concentrations of > 30 kDa molecules can still be seen. Ladder standard and heavy/light chain weights are annotated. Image is crop edited and rotated, unedited image can be found in Figure S8. c-e Antibody purification through column filter shows that IgG is required for prediction of chronological age. Sixteen donor samples were selected to obtain coverage of chronological age regression dynamic range (Methods). These 16 samples were processed in 4 ways: (1) no processing (sample source), (2) filtered through 30 kDa column and only the filtrate (> ~ 15 kDa molecules retained; filtrate), (3) filtered through 30 kDa column and only the flow-through retained (<~ 75 kDa molecules retained; flow through), and (4) the filtrate and flow through were recombined after running through column. c Correlation between log10 peptide intensities show sample source, filtrate + flow-through, and filtrate all recapitulate original signal. In contrast, the flow-through alone, which is IgG depleted, has no correlation with original peptide-antibody binding. d In addition to raw signal being recapitulated, the machine learning regression model is recapitulated only when IgG is present. The 16 samples are plotted as machine learning regression values from the original (x-axis) and filter column-processed (y-axis). e Same as (d), but axes’ values are the di-serine peptide score rather than chronological age regression model