Image subtraction method for improving lateral resolution and SNR in confocal microscopy

A method based on the subtraction of images taken under different pinhole sizes is provided to improve the lateral resolution and SNR in confocal microscopy. Theoretical analyses have been performed and a notable enhancement in lateral resolution and SNR is shown by experimental results. Subtraction parameter and pinhole size have been discussed in detail. Compared with other resolution enhancement concepts, this method is much simpler and cheaper without using strong lasers. And it well suits for real time applications. It can be easily applied to a general confocal microscope and suitable for both fluorescence and non-fluorescence sample.

► Using dual-axis confocal structure.
► Theoretical analysis of PSFs and OTFs in detail.
► An improved subtraction form to avoid signal loss.
► Greatly improved lateral resolution and SNR compared with the confocal result.
► Analyses of the subtraction parameter and pinhole size.