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Improved multiphoton imaging in biological samples by using variable pulse compression and wavefront assessment


A variable prism-pair-based pulse compressor with wavefront aberration sensing was used to enhance multiphoton imaging in biomedical samples. This was incorporated into a custom made microscope to reduce pulse temporal length, improve the quality of images of different layers of thick tissues and increase penetration depth. The laser beam aberrations were found to hardly change with the different experimental configurations of the pulse compressor. The optimum pulse compression state was maintained with depth within the tissue, independently of its thickness. This suggests that for each sample, a single experimental configuration is able to provide the best possible image at any depth location; although this needs to be experimentally obtained. Furthermore, a simple method based on laser average power reduction is presented to minimize the risk of photo-damage in biological samples. The use of pulse compression in multiphoton microscopy might have a potential for accurate and improved biomedical imaging.


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