Thin layers of phytoplankton are well documented, common features in coastal areas globally, but little is known about the relationships of these layers to higher trophic levels. We deployed the In Situ Ichthyoplankton Imaging System (ISIIS) to simultaneously quantify the three trophic levels of plankton, including phytoplankton, primary consumers (copepods and appendicularians) and secondary consumers (gelatinous zooplankton). Over a 2-week sampling period, phytoplankton thin layers, primarily composed of Pseudo-nitzschia spp., were common on two of the five sampling days. Imagery showed copepods aggregating in zones of lower chlorophyll-a fluorescence, while appendicularians were more common at greater depths and higher chlorophyll-a levels. All gelatinous zooplankton generally increased in abundance with depth. Bolinopsis spp. ctenophores underwent a ‘bloom,’ and they were the only species observed to aggregate within phytoplankton thin layers. The vertical separation between copepods, phytoplankton and gelatinous zooplankton suggests that copepods may use the surface waters as a predation refuge, only performing short migrations into favorable feeding zones where gelatinous predators are much more abundant. Thin layers containing dense diatom aggregates obstruct light reaching deeper waters (>10 m), which may allow gelatinous zooplankton to avoid visual predation as well as improve the effectiveness of contact predation with copepod prey.