Filtering out contrast reversals for microscopy autofocus

Autofocus functions based on measurement of image resolution appear to be precise and robust for biological microscopy. However, the through-focus response of these functions previously exhibited unwanted local maxima, or side peaks. Here we report theoretical and experimental studies showing that s...

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Bibliographic Details
Published in:Applied optics (2004) 1999-02, Vol.38 (4), p.638-646
Main Authors: Oliva, M A, Bravo-Zanoguera, M, Price, J H
Format: Article
Language:English
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Summary:Autofocus functions based on measurement of image resolution appear to be precise and robust for biological microscopy. However, the through-focus response of these functions previously exhibited unwanted local maxima, or side peaks. Here we report theoretical and experimental studies showing that side peaks are mainly a result of contrast reversals inherent in optical systems at mid-range frequencies. These contrast reversals are not present in frequencies near optical cutoff. Contrast reversals thus limit the lower cutoff for resolution measurement filters, whereas signal-to-noise limits the upper cutoff. These improved bandpass design criteria led to sharp, unimodal autofocus responses for all tested microscopy specimens.
ISSN:1559-128X
DOI:10.1364/AO.38.000638