BACKGROUND: A critical step in automatic microscopy is focusing. This report describes a robust and fast autofocus approach useful for a wide range of microscopic modalities and preparations. METHODS: The focus curve is measured over the complete focal range, reducing the chance that the best focus position is determined by dust or optical artifacts. Convolution with the derivative of a Gaussian smoothing function reduces the effect of noise on the focus curve. The influence of mechanical tolerance is accounted for. RESULTS: The method is shown to be robust in fluorescence, bright-field and phase contrast microscopy, in fixed and living cells, as well as in fixed tissue. The algorithm was able to focus accurately within 2 or 3 s, even under extremely noisy and low contrast imaging conditions. CONCLUSIONS: The proposed method is generally applicable in light microscopy, whenever the image information content is sufficient. The reliability of the autofocus method allows for unattended operation on a large scale. Copyright 2000 Wiley-Liss, Inc.
BACKGROUND: A critical step in automatic microscopy is focusing. This report describes a robust and fast autofocus approach useful for a wide range of microscopic modalities and preparations. METHODS: The focus curve is measured over the complete focal range, reducing the chance that the best focus position is determined by dust or optical artifacts. Convolution with the derivative of a Gaussian smoothing function reduces the effect of noise on the focus curve. The influence of mechanical tolerance is accounted for. RESULTS: The method is shown to be robust in fluorescence, bright-field and phase contrast microscopy, in fixed and living cells, as well as in fixed tissue. The algorithm was able to focus accurately within 2 or 3 s, even under extremely noisy and low contrast imaging conditions. CONCLUSIONS: The proposed method is generally applicable in light microscopy, whenever the image information content is sufficient. The reliability of the autofocus method allows for unattended operation on a large scale. Copyright 2000 Wiley-Liss, Inc.
Authors: Guy Garty; Alan W Bigelow; Mikhail Repin; Helen C Turner; Dakai Bian; Adayabalam S Balajee; Oleksandra V Lyulko; Maria Taveras; Y Lawrence Yao; David J Brenner Journal: Microsc Res Tech Date: 2015-05-04 Impact factor: 2.769
Authors: J M Castillo-Secilla; M Saval-Calvo; L Medina-Valdès; S Cuenca-Asensi; A Martínez-Álvarez; C Sánchez; G Cristóbal Journal: Biomed Opt Express Date: 2017-02-22 Impact factor: 3.732