Image Stabilization for In Vivo Microscopy by High-Speed Visual Feedback Control

Abstract

This paper presents image stabilization for microscopy using horizontal visual feedback control of the objective lens through a five-bar linkage and piezoelectric actuators, and its application to in vivo imaging. Even very small in vivo motion due to heartbeat and breathing makes microscopic observation difficult by blurring the microscope image or impossible by sending a region of interest out of view. In order to remove those unwanted effects of the motion, we have introduced motion-canceling robotic technologies into microscopy. Our image stabilization system through motion-canceling provides users with stabilized image sequences with respect to trembling of in vivo subjects. The developed image stabilization system, in term of robotics, corresponds to a visual feedback control system that consists of a robotic mechanism and a high-speed vision. A high-speed camera installed in the microscope detects the motion of the in vivo subject having topically applied fiducials. To virtually cancel this motion, we move the objective lens, synchronizing the motions of the subject and the lens to remove the relative motion between the two. As a result, we observe motion-free images to ±10 µm. This technology is one of the very demanding technologies in biological research for in vivo observation with high resolution. In this paper, we verify the effectiveness of the developed system through in vivo experiments.Index Terms-Image matching, in vivo imaging, motioncanceling technique, robotic mechanism with piezoelectric actuators, visual feedback control.

Fast Measurement of Sarcomere Length and Cell Orientation in Langendorff-Perfused Hearts Using Remote Focusing Microscopy

“…36 The use of the geometric cell model could be avoided if tissue motion is minimized or accounted for. There are several published methods for compensating for tissue motion in whole-organ preparations, [37][38][39][40][41] which we intend to evaluate and potentially implement in future studies.…”

“…This will facilitate translation of single-cell experiments to the intact heart and, in turn, can be used to obtain SL information to drive subsequent single-cell workloop studies. 53 Further refinements of the technique, in conjunction with emerging methods for tissue immobilization, registration, and tracking, [37][38][39] hold the promise of allowing SL measurements in the unarrested beating heart. Experimental measurement of SL during the full cardiac cycle could be used further to refine biophysically accurate electromechanic simulations, which presently rely on a combination of MRI and histological data sets.…”

Image Sabilization for In Vivo Microscopic Imaging

“…The first solution is a vision based compensation system (Lee et al 2008a). We use a highspeed camera for detecting the in vivo motion, and move the objective lens to follow the detected motion.…”

Improved intravital microscopy via synchronization of respiration and holder stabilization

“…1 Tissue movement, however, is unavoidable as it is the result of vital physiological processes such as respiration and cardiovascular activity. Movement arising from respiration is particularly extensive and has effects on each and every organ of the body and every organ of the body such as tibila anterior, 2 lung, 3 kidney, 4 spinal cord, 5 and brain. 6,7 Because the time taken to acquire an image is typically the same as or longer than a breathing cycle (approx.…”

“…Recently, considerable research has been directed towards developing strategies to overcome motion artifacts. [2][3][4][5][6][7][8][9][10][11] Unlike previous attempts, the method presented here does not require the construction of an expensive mechanical system 4,5 nor does it sacrifice image quality in order to achieve faster image acquisition. 8,9 Rather, the only equipment required is a custom made mechanical holder and a commercial ventilator; this setup can be easily applied to all commercial laser scanning microscopes (LSM) without any specific hardware modification.…”