Suspended particulate organic matter (POM) plays a critical role in the planktonic ecology of Lake Michigan during seasonal thermal stratification. We show, based on stable isotope and ribosomal RNA (rRNA) data, that the relationship between suspended POM and active biomass changes as thermal stratification persists. Stable isotope data indicated that sources of suspended POM change between July and October, moving from primary production at a deep chlorophyll layer to recycling-based production in surface waters. Concomitant change in the distribution of active bacterial and eukaryotic biomass was observed as indicated by rRNA abundances. Active bacterial and eukaryotic biomass were highly correlated throughout the year. However, the correlation between suspended POM and active bacterial biomass varied seasonally and reflected the transitions in planktonic ecology. Suspended POM from depths >60 m was primarily of sedimentary origin. The combined application of stable isotope and rRNA analysis of suspended POM indicated a dynamic relationship between the bulk POM reservoir and living planktonic biomass.