The PaviaATN data consists of 62 4-channel fluorescence microscopy images of size 2720 × 2720. The four channels, in order, label Nuclei (first two), Actin and Tubulin. It was imaged in the Synthetic Physiology Laboratory(https://www.syntheticphysiologylab.com/) of the University of Pavia, and introduced in μSplit: image decomposition for fluorescence microscopy(https://arxiv.org/abs/2211.12872), published at ICCV 2023. Detailed Description The PaviaATN dataset comprises static lambda-stacks from a human keratinocytes cell line (HaCaT) expressing GFP-tubulin, RFP-LifeAct, and a customized version of the cell cycle indicator FastFUCCI that uses various combinations of a yellow fluorescent protein (YPF, mTurquoise2) and a far-red fluorescent protein (iRFP, miRFP670) to indicate multiple phases of the cell cycle. When a cell is in the G1 phase, increasing intensities of YFP fluorescence are detected in the nucleus. As a cell moves from G1 to S phase (G1/S), both YFP and iRFP fluorescence are detected in the nucleus of the cell. Finally, the sole iRFP fluorescence is detected in the nucleus during the S-G2-M phase. At the onset of the G1 phase, the nucleus shows no visible fluorescence intensity. In [μSplit: image decomposition for fluorescence microscopy](https://arxiv.org/abs/2211.12872), this dataset was used to resolve overlapping structures. All Images were acquired in a Nikon Ti2 microscope (100x silicon oil objective) equipped with an Okolab environmental control chamber and a Crest V3 spinning disk confocal in widefield mode.