Continuous live cell imaging using dark field microscopy

The optical observation of live cell behavior is critical for biological and biomedical studies. The development of techniques for long-term cell and tissue imaging is, however, hindered by phototoxicity induced by excited fluorophores. We, herein, propose a methodology to capture live cell behavior using dark field microscopy (DM). Since the light intensity of DM is merely ∼0.1% of bright-field microscopy (BM) and ∼0.5% of fluorescence microscopy (FM), it allows super long and frequent live cell imaging. Our results demonstrate that continuous exposure to DM light for 48 h brings about no observable effect on the growth rate of 3T3 fibroblasts and HepG2 hepatoma cells, indicating minimum photo-toxicity. Moreover, DM images show contrast comparable to FM, which does not depend on the probes and staining efficiency. We, therefore, conclude that the proposed approach is suitable for long-term live cell imaging with super-high temporal resolution.