Dynamic speckle illumination wide-field fluorescence microscopy with actively optical manipulation of rotational angles.

We present a dynamic speckle illumination wide-field fluorescence microscopy (DSIWFM) combined with a line optical tweezers (LOTs) for rotational fluorescence sectioning imaging. In this method, large polystyrene fluorescent microspheres are stably trapped with LOTs, and precisely manipulated to rotate around a specific rotation axis. During the rotation process, multiple raw fluorescence images of trapped microspheres are obtained with dynamic speckle illumination. The root-mean-square (RMS) algorithm is used to extract the drastically changing fluorescent signals in the focal plane to obtain the fluorescence sectioning images of the samples at various angles. The influence of speckle granularity on the image quality of fluorescence sectioning images is experimentally analyzed. The rotational fluorescence sectioning images obtained by DSIWFM with LOTs could provide an alternative technique for applications of biomedical imaging.