Synthetic holographic phase imaging in confocal microscopy and applications

We present synthetic optical holography (SOH) as a new phase imaging modality in confocal microscopy, where amplitude and phase information are encoded holographically by a reference wave with a linear-in-time phase function. The implementation of SOH is technically simple, only requiring the addition of a linearly moving reference mirror. We demonstrate reliable confocal optical topography mapping with sub-nanometer depth resolution. Furthermore, we show numerical refocusing of out-of-focus confocal data, restoring a spatial resolution near the diffraction limit for situations where the sample is located up to 60 microns away from the system’s focal plane. We envision that quantitative phase imaging as provided by SOH will allow for a host of new possibilities in monochromatic confocal microscopy, ranging from stain- and dye-free confocal bio-imaging and phase-resolved second harmonic microscopy to three-dimensional imaging and computational aberration correction.