Methods and apparatus for measuring optical properties of biological and chemical substances

• US 5,721,435 A
• Filed: 04/09/1996
• Issued: 02/24/1998
• Est. Priority Date: 04/09/1996
• Status: Expired due to Fees

• Current Assignee: Agilent Technologies, Inc.
• Original Assignee: Hewlett Packard Company (HP Inc.)
• Inventors: Troll, Mark A.
• Primary Examiner(s): Westin, Edward P.
• Assistant Examiner(s): Lee, John R.
• Application Number:
  US08/630,054
• Time in Patent Office: 686 Days
• Field of Search: 250/559.29, 250/559.3, 250/559.4, 250/559.06, 250/559.44, 250/216, 250/237 R, 250/237 G, 356/320, 356/372, 356/375, 356/397, 356/398, 356/400, 356/401, 356/244, 435/289, 435/291, 422/50, 422/55, 422/82.05, 422/82.08
• US Class Current: 250/559.29
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: G01N 21/253 for batch operation, i.e. multisample apparatus analytical automats G01N35/00

Abstract

An optical measurement system includes a test surface having disposed thereon a plurality reference markings and a plurality of test spots containing chemical and biological substances. Molecules of interest contained in the chemical and biological substances are tagged with optically detectable labels. A light source generates a beam of light which is then directed by a deflecting sub-system across the test surface, thereby individually impinging the test spots and the reference markings. Optical signals resulting from the impingement are detected by a detection means, which records such detection and the time of the detection. The reference markings have optically unique signatures to distinguish the markings from the optical signatures of the signals emitted from the chemical and biological substances in the test spots. The reference markings are spaced apart at known distances and serve to provide a constant calibration of the scan speed of the beam of light. In one embodiment, a Winston concentrator directs the light emanating from the test surface to the detector. In another embodiment, an off-axis parabolic mirror arrangement directs the light to the detector.

Claims

1. A method of measuring optical properties of optically detectable substances, including the steps of:

providing a surface;

forming reference marks on said surface, including selecting areas on said surface and interspersing said reference marks among said selected areas, said step of forming reference marks further including selecting known spacings therebetween;

forming a plurality of test spots by disposing said optically detectable substances upon said selected areas;

selectively illuminating portions of said surface with light, forming a light spot thereon, said light spot illuminating at least some of said test spots and said reference marks;

detecting an optical property of each of a plurality of said test spots and said reference marks, including for each detection the substeps of recording said optical property and recording the time of said detection; and

computing positions of said test spots on said surface based upon said recorded times of detection.

2. The method of claim 1 further including a step of performing a test on some of said test spots, said step of performing a test being taken prior to said step of selectively illuminating portions of said surface.

3. The method of claim 2 further including repeating said steps of performing a test, selectively illuminating portions of said surface, and detecting an optical property.

4. The method of claim 1 wherein said step of forming reference marks further includes selecting varying spacings between said reference marks.

5. The method of claim 1 wherein said step of computing positions of said test spots includes substeps of:

plotting data points representing relative positions of said reference marks versus recorded times of detection of said reference marks;

fitting a curve to said data points; and

interpolating between said data points for said recorded times of detection of said test spots, basing said interpolating on said curve.

6. The method of claim 1 wherein said step of computing positions of said test spots includes substeps of:

computing a rate of motion of said light spot between two of said reference marks, including dividing a measure of spacing between said two reference marks by the difference between said recorded times of detection of said two reference marks;

computing a time difference between said recorded time of detection of one test spot positioned between said two reference marks and said recorded time of detection of a first of said two reference marks; and

multiplying said rate of motion of said light spot by said time difference, thereby determining a position of said one test spot relative to said first of said two reference marks.

7. An optical detection system for detecting a plurality of optically tagged biological and chemical materials, including:

a source of collimated light;

a surface onto which said optically tagged biological and chemical materials are disposed;

a plurality of reference marks disposed on said surface, said reference marks being spaced apart by known amounts, said reference marks being optically distinguishable from said optically tagged biological and chemical materials;

raster means for directing said collimated light across said surface in raster fashion so that said collimated light impinges upon said optically tagged biological and chemical materials and said reference marks, wherein impingement by said collimated light gives rise to optical signals;

means for detecting said optical signals, including means for storing a parameter of said optical signals and means for storing the time of each detection of said optical signals; and

computing means, in communication with said means for storing the time of detection, for determining positions of said optically tagged biological and chemical materials.

8. The optical detection system of claim 7 wherein said optical signals result in the transmission of electromagnetic (EM) energy from said surface, the optical detection system further including collector means for directing said transmission of EM energy to said means for detecting.

9. The optical detection system of claim 8 wherein said collector means is a portion of a parabolic mirror arranged between said surface and said means for detecting such that light emissions of said optical signals from said surface are directed to said means for detecting.

10. The optical detection system of claim 8 wherein said collector means is a compound parabolic concentrator having two parabolic mirrors symmetrically arranged about an axis, said means for detecting being positioned proximate to and in alignment with foci of said two parabolic mirrors, said surface being positioned opposite to and facing said means for detecting so that light emissions of said optical signals from said surface are reflected by said two parabolic mirrors toward said means for detecting.

11. The optical detection system of claim 10 wherein said two parabolic mirrors are curved about said axis to form a parabolic surface of rotation about said axis.

12. The optical detection system of claim 7 wherein said surface is a front face of a transparent substrate, said collimated light being directed by said raster means upon a rear face of said transparent substrate, wherein said collimated light impinges upon said optically tagged biological and chemical materials and said reference marks by traversing through a thickness of said transparent substrate toward said front face.

13. The optical detection system of claim 7 wherein a first pair of said reference marks is spaced apart by a first measure of separation and a second pair of said reference marks is spaced apart by a second measure of separation.

14. The optical detection system of claim 7 wherein reference marks disposed on a central portion of said surface are more closely spaced than reference marks disposed on a periphery of said surface.

15. The optical detection system of claim 7 wherein said reference marks have optical signatures unique thereto so as to be distinguishable from optical signatures of said optically tagged biological and chemical materials.

16. The optical detection system of claim 7 wherein said means for detecting includes a first detector for detecting optical signals originating from said optically tagged biological and chemical materials, and a second detector for detecting optical signals originating from said reference marks.

17. An optical detection system for measuring optical tags attached to selected molecules in chemical and biological specimens, including:

a laser beam source for generating a beam of light;

means for deflecting said beam of light;

an optically transparent plate having a front surface onto which said chemical and biological specimens are disposed, said transparent plate having a rear surface opposite said front surface, said front surface having optically identifiable reference marks formed thereon, said reference marks having known measures of separation therebetween;

electromagnetic energy (EM) detection means for generating output signals in response to detection of electromagnetic energy;

a parabolic concentrator having a central axis and further having at least two parabolic reflective surfaces symmetrically arranged about said central axis, said at least two parabolic reflective surfaces defining an entry aperture formed at a first end of said concentrator and further defining an exit aperture formed at an end opposite said first end, foci of said two parabolic reflective surfaces being proximate said exit aperture; and

computation means for computing positions of said chemical and biological specimens disposed upon said front surface, including memory means for storing said output signals generated by said EM detection means and for storing a time value indicating the time at which said EM detection means generates said output;

said optically transparent plate being received in said entry aperture of said concentrator such that said front surface is directed toward said exit aperture;

said EM detection means being received in said exit aperture of said concentrator;

said beam of light being scanned across said rear surface of said optically transparent plate by said means for deflecting, said beam of light passing through said optically transparent plate to impinge upon said chemical and biological specimens and said reference marks;

wherein impingement of said beam of light results in electromagnetic energy emissions from each of said chemical and biological specimens and said reference marks, said emissions being directed by said concentrator to said EM detection means, thereby generating output signals from said EM detection means.

18. The optical detection system of claim 17 wherein said at least two parabolic reflective surfaces are curved about said axis of said concentrator to form a parabolic surface of rotation about said axis.

19. The optical detection system of claim 17 further including a prism coupled to said rear surface so that said means for deflecting directs said beam of light upon a surface of said prism, said beam of light passing through said prism and across a prism-rear surface interface with minimum refraction.

20. The optical detection system of claim 17 wherein said front surface of said transparent plate is non-planar.

21. The optical detection system of claim 17 wherein at least two of said measures of separation between said reference marks are unequal.