This repository contains flow cytometry (FCS) and imaging flow cytometry (ImageStream) datasets generated from Nematostella vectensis larval-derived cells to characterize RLRb-expressing immune cells and their functional responses to immune stimulation. The dataset includes experiments designed to profile and functionally characterize RLRb-positive cells, including intracellular RLRb immunostaining, RLRb::mCherry reporter expression, poly(I:C)-induced immune activation, phagocytosis of pHrodo-labeled Staphylococcus aureus, pHrodo-labeled Escherichia coli, and DQ-ovalbumin, apoptosis (Apotracker green), proliferation (Edu incorporation), and detailed morphological analysis using imaging flow cytometry. Together, these data capture multiple aspects of RLRb-mediated antiviral and phagocytic immune responses at the single-cell level. Data were acquired using an Amnis ImageStreamX Mk II imaging flow cytometer and BD FACSAria III. Fluorescent labels and probes include RLRb::mCherry, DRAQ5, pHrodo-conjugated Staphylococcus aureus and Escherichia coli, DQ-ovalbumin, Calcein Violet-AM, Zombie NIR viability dye, Apotracker-green, and SYTOX Blue. The repository contains raw FCS files, raw ImageStream (.rif) files, and associated analysis outputs (e.g., compensation matrices, IDEAS exports, and reports), allowing full reanalysis and reuse of the data. Gating strategies and experimental design are described in the associated manuscript. These datasets support the analyses presented in the preprint: Kozlovski et al., “Functional characterization of specialized immune cells in a cnidarian reveals an ancestral antiviral program.”bioRxiv (2025). https://doi.org/10.1101/2025.01.24.634691 This Zenodo record provides the raw and processed single-cell cytometry data underlying the figures and conclusions of that study, enabling independent validation, re-analysis, and comparative immunology research in early-branching metazoans.