Position referencing in optical microscopy thanks to sample holders with out‐of‐focus encoded patterns

Abstract

SummaryThis article introduces smart sample holders for optical microscopy. Their purpose is to allow the absolute determination of the position of the observed zone with respect to the sample holder itself and with a high accuracy. It becomes then straightforward to find a given zone of interest by positioning coarsely the microscope slide to the same position coordinates. Furthermore images recorded during different observation sessions; i.e. for slightly different positions; can be processed numerically in order to superimpose them with a high accuracy. Thus the slight deviations of the microscope slide position and orientation due to the different observations are compensated numerically and a perfect superimposition of the recorded images is performed. Then accurate site-by-site image comparisons become possible even for images recorded during different observation sessions and over a long period of time. The subpixel capability of the proposed method is demonstrated and those smart microscope slides constitute a new tool for live cell experiment.In practise, an encoded geometrical pattern used as position reference is inserted in a plane parallel to the surface receiving the tissue section or sample. Then the transition of the focus position from the tissue section to the position reference requires only a vertical adjustment and does not affect the lateral coordinates of observation. The numeric processing of the image of the position reference pattern allows the retrieval of the lateral coordinates that are also used for the tissue section image. Thus each image is recorded with a set of position coordinates that defines accurately the position of the observed area with respect to the sample holder itself.

Characterization and compensation of XY micropositioning robots using vision and pseudo-periodic encoded patterns

“…The PPP algorithm can be extended to measure the angular position and also the out-of-plane motion (z direction) using depth-from-focus approach [17].…”

“…Those methods are feature dependent so that the pattern has to remain inside the region of interest to be analyzed thus limiting the range of measurement. To overcome such a drawback, a technique based on pseudoperiodic patterns has been proposed by different authors [15], [16], [17].…”

Twin-scale Vernier Micro-pattern for Visual Measurement of 1-D in-plane Absolute Displacements with Increased Range-to-Resolution Ratio

“…Those methods are feature-dependent so that salient objects have to remain inside the region of interest to be analyzed thus limiting the allowed range of displacement. To overcome such a drawback, a technique based on pseudo-periodic patterns has been proposed by different authors (Masa et al 2008;Boyton 2003;Galeano-Zea et al 2010;Sandoz et al 2007). In these methods, the pseudo-periodic pattern is attached to the mobile object of interest.…”

Twin-scale vernier micro-pattern for visual measurement of 1D in-plane absolute displacements with increased range-to-resolution ratio

“…Those methods are feature dependent so that salient objects have to remain inside the region of interest to be analyzed limiting thus the allowed range of displacement. To overcome such drawback, a technique based on pseudo-periodic patterns has been proposed by different authors [6], [7], [8], [9]. In these methods, the pseudoperiodic pattern is attached to the mobile object of interest.…”