Focusing of microscopes and reading of microarrays

• US 7,095,032 B2
• Filed: 06/03/2005
• Issued: 08/22/2006
• Est. Priority Date: 03/20/1998
• Status: Expired due to Fees

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: James W. Overbeck, Jean I. Montagu
• Inventors: Montagu, Jean I., Overbeck, James W.
• Primary Examiner(s): Hannaher, Constantine
• Application Number:
  US11/145,529
• Publication Number:
  US 20050285049A1
• Time in Patent Office: 445 Days
• Field of Search: 250/458.1, 250/459.1, 250/461.1, 250/461.2
• US Class Current: 250/458.1
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G02B 21/16 adapted for ultraviolet illumination ; Fluorescence microscopes G02B21/0076 takes precedence

Abstract

Microscopes, including viewing and other microscopic systems, employ a hinged, tiltable plate to adjust focus on a flat object such as a microscope slide or biochip by motion, achieved by tilting, which is substantially normal to the focus point on the plane of the object. By employing two such tiltable arrangements, relatively long scan lines of e.g., flying objective, single pixel on-axis scanning can be accommodated. The tilting support plate is specifically constructed to provide tailored locations for different objects in series along the Y axis of the plate. The plate can accommodate beaters and cooled plates and/or the flat object being examined. In a fluorescence scanning microscope, locations are specifically adapted to receive microscope slides and biochip cartridges such as Affymetrix'"'"'s “Gene Chip®”. A scanning microscope under computer control, employing such a focusing action, enables unattended scanning of biochips with a simple and economical instrument. Also shown are flexure-mounting of a support plate to define the hinge axis, techniques for automatically determining position and focus, and a rotatably oscillating flying micro-objective scanner combined with the tilting plane focus system. Construction and control techniques are shown that realize simple and accurate focusing. Methods of examination of biological materials are disclosed. Simple and efficient focused scanning with a flying micro-objective of ordered arrays of nucleotides and nucleic acid fragments carried upon a microscope slide or other substrate is disclosed. Quantified fluorescence imaging is economically achieved by combined use of the described scanning and focusing arrangement and use of simple and accurate calibration modules respectively for example for Affymetrix'"'"'s “Gene Chip” microarray modules and for microscope slides.

Claims

1. An optical scanner, comprising:

a scanning structure including an oscillating rotary arm constructed to support an objective lens arranged to provide an optical light path to and from an examined object being scanned;

said light path extending partially along a rotation axis of said rotary arm;

a translation mechanism constructed to produce movement between said examined object and said objective lens;

an automatic focusing system constructed to bring a portion of said object in focus with respect to said objective lens;

a data collection control and processing unit arranged to collect optical scan data during said scanning and process the collected data.

2. The optical scanner of claim 1 including a support constructed to receive said examined object.

3. The optical scanner of claim 2, wherein said translation mechanism is constructed to displace linearly said support including said examined object.

4. The optical scanner of claim 3, wherein said object is associated with a microscope slide.

5. The optical scanner of claim 4, wherein said object includes an ordered array of nucleotides that may fluoresce.

6. The optical scanner of claim 4, wherein said object includes an ordered array of oligonucleotides.

7. The optical scanner of claim 4, wherein said object includes an ordered array of deposits of nucleic acid fragments.

8. The optical scanner of claim 3, wherein said object includes a biochip.

9. An optical scanner, comprising:

a scanning structure including an oscillating rotary arm constructed to support an objective lens arranged to provide an optical light path to and from a biochip mounted on a module and having a scanned surface located inside a cavity and through a transparent support;

said light path extending partially along a rotation axis of said rotary arm;

a translation mechanism constructed to produce movement between said examined object and said objective lens;

an automatic focusing system constructed to bring a portion of said scanned surface in focus with respect to said objective lens; and

a data collection control and processing unit arranged to collect optical scan data during scanning of said scanned surface of said biochip and process the collected data.

10. The optical scanner of claim 9, wherein said scanned surface includes an ordered array of nucleotides that may fluoresce.

11. The optical scanner of claim 9, wherein said scanned surface includes an ordered array of oligonucleotides.

12. The optical scanner of claim 9, wherein said scanned surface includes an ordered array of deposits of nucleic acid fragments.

13. The optical scanner of claim 9, wherein said automatic focusing system is constructed to perform dynamic focusing.

14. The optical scanner of claim 9 wherein said scanned surface includes a calibrating tool having a surface layer of effective fluorophores.

15. The optical scanner of claim 9, wherein said processing unit arranged to create an image of the optical scan data.

16. The optical scanner of claim 9, wherein said automatic focusing system includes a light emitting diode (LED).

17. The optical scanner of claim 9, wherein said automatic focusing system further includes a split photocell detector.

18. The optical scanner of claim 9, wherein said automatic focusing system includes a prescan analyzer constructed to determine a height of focus for each location used during an examination scan.

19. The optical scanner of claim 9 further including a thermoelectric plate.