High-contrast epi-fluorescence wide-field imaging of biological cells using integrating-bucket method

Abstract

We present an optical lock-in detection scheme, called the integrating-bucket technique, as a signal-to-background ratio enhancement method for wide-field fluorescence imaging of biological cells. The proposed method uses sinusoidally modulated illumination light and captures four frames of fluorescence images, one per each quarter of the modulation period, by integrating the fluorescence intensity signal. The capability of this technique is demonstrated by imaging fluorescent bead solutions as well as labeled cells. The results show that the method yields a 4–10dB higher signal contrast than conventional fluorescence microscopy, and a background-free fluorescence image can be extracted within a sub-second time scale. Our findings indicate that the proposed method could be advantageous for the long-term study of live cells.