Methods, Software, and Apparatus for Focusing an Optical System Using Computer Image Analysis

Abstract

Methods, software, and apparatus for focusing an image in biological instrument are disclosed. Focusing elements are moved to various focus positions within a focus element travel range, and sample images are captured at the various focus positions. The sample images are resolved into subregions and an optimal focus position is determined based on the image intensity statistical dispersions within the identified subregions.

Description

CROSS-REFERENCE TO RELATED APPLICATION[0001]This application is a continuation of U.S. application Ser. No. 10 / 881,868 filed Jun. 30, 2004, now U.S. Pat. No. 7,394,943, which is incorporated herein by reference.INTRODUCTION[0002]The present teachings generally relate to methods, software, and apparatus useful in machine-vision-based focusing in imaging instrumentation, for example in biological instruments.[0003]During imaging operations such as biological analysis involving nucleotide sequencing or microarray processing, photo-detectors are used to detect signals arising from labeled samples or probe features responsive to selected target analytes. These signals can take the form of electromagnetic emissions that are desirably analyzed to quantify signal intensities arising from labeled samples or probe features and are subsequently resolved to quantitatively or qualitatively evaluate the presence of a target analyte within a sample. Frequently, images associated with such biologic...

AI Technical Summary

Benefits of technology

[0008]In the case of a very high-density microarray, for example, lack of sharpness or blur in an image increases cross-talk between individual microarray element signals, and reduces the instrument's ability to differentiate among individual microarray elements. However, for a particular biological instrument, it is typically a requirement that the instrument be able to “read” (which is to say differentiate and measure the intensity of) substantially every region in the target sample. Accordingly, adequate focus is typically required in substantially every region of the target sample's recorded image.

[0010]Images of a target sample are recorded while the target sample and / or an optical system associated with the target sample are moved through a series of positions bounding an ideal focus position. An analysis of the recorded images produces a map of the target sample's surface relative to the optical system's focus plane. In various embodiments, the target sample's surface height distribution is analyzed to select a substantially optimal focus position for the target sample. Additionally, the present teachings can provide information regarding target sample physical defects, thereby providing beneficial sample quality control procedures in addition to automatically focusing optics for imaging a target sample.

Problems solved by technology

However, optical systems of desirable cost and complexity, suffer from various types of optical aberrations that complicate the focusing process.

A common optical aberration is field curvature aberration, which results in the situation where, for example, at one focal position, the center of an image will be clear and in sharp focus, but the edges of the image will be blurry and out of focus.

There are also higher-order aberrations in optical systems which may be non-symmetrical with regard to focus.

Further, typical detectors used to image target samples are not one-dimensional point detectors but rather are two-dimensional and therefore subject to tip and tilt.

However, in real systems, there typically is some tip and / or tilt between the plane of the image detector and the surface of the target sample.

In the case of a very high-density microarray, for example, lack of sharpness or blur in an image increases cross-talk between individual microarray element signals, and reduces the instrument's ability to differentiate among individual microarray elements.

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