Autofocusing in computer microscopy: Selecting the optimal focus algorithm

Autofocusing is a fundamental technology for automated biological and biomedical analyses and is indispensable for routine use of microscopes on a large scale. This article presents a comprehensive comparison study of 18 focus algorithms in which a total of 139,000 microscope images were analyzed. Six samples were used with three observation methods (brightfield, phase contrast, and differential interference contrast (DIC)) under two magnifications (100× and 400×). A ranking methodology is proposed, based on which the 18 focus algorithms are ranked. Image preprocessing was also conducted to extensively reveal the performance and robustness of the focus algorithms. The presented guidelines allow for the selection of the optimal focus algorithm for different microscopy applications. Microsc. Res. Tech. 65:139–149, 2004. © 2004 Wiley‐Liss, Inc.

[1]  Sim Heng Ong,et al.  Autofocusing for tissue microscopy , 1993, Image Vis. Comput..

[2]  I T Young,et al.  A comparison of different focus functions for use in autofocus algorithms. , 1985, Cytometry.

[3]  C. Ortiz de Solórzano,et al.  Evaluation of autofocus functions in molecular cytogenetic analysis , 1997, Journal of microscopy.

[4]  J. F. Brenner,et al.  An automated microscope for cytologic research a preliminary evaluation. , 1976, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[5]  D. Vollath.  Automatic focusing by correlative methods , 1987 .

[6]  Bradley J. Nelson,et al.  Wavelet-based autofocusing and unsupervised segmentation of microscopic images , 2003, Proceedings 2003 IEEE/RSJ International Conference on Intelligent Robots and Systems (IROS 2003) (Cat. No.03CH37453).

[7]  K Cook,et al.  Comparison of autofocus methods for automated microscopy. , 1991, Cytometry.

[8]  Shree K. Nayar,et al.  Shape from Focus , 1994, IEEE Trans. Pattern Anal. Mach. Intell..

[9]  D. Vollath.  The influence of the scene parameters and of noise on the behaviour of automatic focusing algorithms , 1988 .

[10]  Bradley J. Nelson,et al.  Micromanipulation contact transition control by selective focusing and microforce control , 2003, 2003 IEEE International Conference on Robotics and Automation (Cat. No.03CH37422).

[11]  M. Mendelsohn,et al.  Computer-oriented analysis of human chromosomes. 3. Focus. , 1972, Computers in biology and medicine.