Near Zero Photon Bioimaging

Enhancing the reliability and reproducibility of optical microscopy by reducing specimen irradiance continues to be an important biotechnology target. As irradiance levels are reduced, however, the particle nature of light is heightened, giving rise to Poisson noise, or photon sparsity that restricts only a few (0.5%) image pixels to comprise a photon. Photon-sparsity can be addressed by collecting more than 200 photons per pixel; this, however, requires extended acquisition durations and, thus, suboptimal imaging rates. Here, we introduce near-zero photon imaging, a method that operates at kHz rates and 10,000-fold lower irradiance than modern microscopy. To achieve this performance, we deployed a judiciously designed epi-fluorescence microscope enabling ultralow background and artificial intelligence that learns to reconstruct biological images from as low as 0.01 photons per pixel. We demonstrate that near-zero photon imaging captures the structure of both multicellular and subcellular targets with high fidelity, including features represented by nearly zero photons. Beyond optical microscopy, the near-zero photon imaging paradigm can be applied in remote sensing, covert applications, and biological or biomedical imaging that utilize damaging or quantum light.