Molecular interferometric imaging process and apparatus

• US 7,522,282 B2
• Filed: 05/04/2007
• Issued: 04/21/2009
• Est. Priority Date: 11/30/2006
• Status: Active Grant

• Current Assignee: Purdue Research Foundation (Purdue University)
• Original Assignee: Purdue Research Foundation (Purdue University)
• Inventors: Nolte, David D., Zhao, Ming
• Primary Examiner(s): Lee; Hwa (Andrew) S
• Application Number:
  US11/744,726
• Publication Number:
  US 20080129981A1
• Time in Patent Office: 718 Days
• Field of Search: 356/450, 356/451, 356/456
• US Class Current: 356/450
• CPC Class Codes: G01N 21/253 for batch operation, i.e. multisample apparatus analytical automats G01N35/00
• CPC Class Codes: G01N 21/45 using interferometric methods; using Schlieren methods

Abstract

A molecular interferometric imaging system for detecting an analyte in a sample, that includes an illumination source providing a beam of radiation; a pixel array for detecting radiation in an image plane; a biolayer designed to react to the analyte when it comes in contact with the sample; a substrate designed to convert phase modulation into intensity modulation which can be detected and imaged directly by the pixel array, the biolayer being on the substrate; a reference surface; an image switching means for switching between a first position for collecting a sample image of the biolayer, and a second position for collecting a reference image of the reference surface; and a processing means for producing a composite image using the sample image and the reference image for illumination normalization.

Claims

1. A molecular interferometric imaging system for detecting an analyte in a sample, the molecular interferometric imaging system comprising:

an illumination source providing a beam of radiation;

a pixel array comprising a plurality of pixels for detecting radiation at a plurality of locations in an image plane to produce an image comprising a plurality of pixel readings;

a biolayer designed to react to the analyte when the biolayer comes in contact with the sample;

a substrate, the biolayer being located on the substrate, the substrate being designed to convert phase modulation into intensity modulation which can be detected and imaged directly by the pixel array;

a reference surface;

an image switching means for switching between a first position in which the beam provided by the illumination source is incident on the biolayer and a reflected sample beam is produced which is directed to the pixel array to generate a sample image, and a second position in which the beam provided by the illumination source is incident on the reference surface and a reflected reference beam is produced which is directed to the pixel array to generate a reference image; and

a processing means for producing a composite image using the sample image and the reference image for illumination normalization.

2. The system of claim 1, wherein the substrate comprises a support layer and a spacer, the thickness of the spacer being selected to produce an in-line quadrature condition.

3. The system of claim 2, wherein the spacer acts as the reference surface.

4. The system of claim 2, wherein the support layer is silicon and the spacer is a thermal oxide layer.

5. The system of claim 4, wherein the thickness of the oxide layer is 120 nm.

6. The system of claim 1, wherein the substrate acts as the reference surface, and the image switching means is a stage for moving the substrate and the biolayer between the first position and the second position.

7. The system of claim 1, wherein the biolayer comprises a plurality of separate spots deposited on the substrate with the substrate being exposed to separate the plurality of separate spots.

8. The system of claim 7, wherein the substrate acts as the reference surface.

9. The system of claim 1, wherein the processing means computes a difference between the plurality of pixel readings of the reference image and the plurality of pixel readings of the sample image on a pixel by pixel basis.

10. The system of claim 1, wherein the illumination source is a laser tuned to a desired frequency, the desired frequency being selected such that the reflected radiation from the substrate and the radiation from the biolayer are in an in-line quadrature condition.

11. The system of claim 1, further comprising an illumination filter.

12. The system of claim 11, wherein the illumination source produces a multi-frequency beam and the illumination filter filters the beam such that only a desired frequency passes through the illumination filter, the desired frequency being selected such that the reflected radiation from the substrate and the radiation from the biolayer are in an in-line quadrature condition.

13. The system of claim 11, wherein the illumination source produces a multi-frequency beam and the illumination filter filters the beam such that only a first wavelength and a second wavelength pass through the illumination filter, the first and second wavelengths being selected such that the reflected radiation from the substrate and the radiation from the biolayer are in an in-line quadrature condition, the first wavelength being different from the second wavelength.

14. The system of claim 11, further comprising a detection filter, the detection filter being matched to the illumination filter to produce a desired property in the beam incident on the pixel array.

15. The system of claim 1, further comprising a beam splitter, the beam splitter directing the beam from the illumination source to the image plane and directing the reflected beam from the image plane to the pixel array.

16. The system of claim 1, further comprising a trigger to synchronize image collection with switching by the image switching means.

17. The system of claim 16, wherein the image switching means is a mirror that moves between the first position to view the biolayer and the second position to view the reference surface.

18. The system of claim 16, wherein the image switching means is a spinning disc that includes an aperture, the spinning disc being in the first position when the biolayer is visible through the aperture and the spinning disc being in the second position when the biolayer is not visible through the aperture.

19. The system of claim 16, wherein the image switching means is a polarizing beam splitter that switches back and forth between orthogonal polarizations, the polarizing beam splitter being in the first position when the polarization setting directs the beam to the biolayer and the polarizing beam splitter being in the second position when the orthogonal polarization setting directs the beam to the reference surface.

20. The system of claim 1, further comprising a trigger to synchronize image collection with changing of properties of the beam incident on the pixel array.

21. The system of claim 1, wherein the biolayer is immersed in a fluid during collection of the sample image.

22. The system of claim 21, wherein the fluid is actively flowing during collection of the sample image.

23. The system of claim 1, wherein the image switching means is a stage, the stage moving between the first position in which a first set of pixels of the plurality of pixels is exposed to the biolayer, and the second position in which a second set of pixels of the plurality of pixels is exposed to the biolayer, there being no intersection between the first set of pixels and the second set of pixels.

24. A method of molecular interferometric imaging for detecting an analyte in a sample using a pixel array comprising a plurality of pixels for detecting radiation at a plurality of locations in an image plane, and a substrate designed to convert phase modulation into intensity modulation which can be detected and imaged directly by the pixel array, the method comprising:

exposing a biolayer that is deposited on the substrate to the sample, the biolayer being designed to react to the analyte when the biolayer comes in contact with the sample;

positioning an image switching means in a first position;

illuminating the biolayer with a radiation source to produce a sample beam;

capturing the sample beam in a sample image using the pixel array;

positioning the image switching means in a second position;

illuminating a reference surface with the radiation source to produce a reference beam;

capturing the reference beam in a reference image using the pixel array;

computing a composite image using the sample image and the reference image for illumination normalization.

25. The method of claim 24 wherein the sample image exposes a first set of pixels to the biolayer and exposes a second set of pixels to the reference surface, and the reference image exposes the first set of pixels to the reference surface and exposes the second set of pixels to the biolayer, and the computing a composite image step comprises:

subtracting the pixel readings from the sample image from the pixel readings of the reference image.

26. The method of claim 24 wherein the sample image exposes a first set of pixels to the biolayer and exposes a second set of pixels to the reference surface, and the reference image exposes the first set of pixels to the reference surface and exposes the second set of pixels to the biolayer, and the computing a composite image step comprises:

computing, on a pixel by pixel basis,

$\begin{array}{cc}{I}_{\mathrm{Diff}}=2<br><br>\frac{\left(I_A-I_B\right)}{\left(I_A+I_B\right)}& \left(4\right)\end{array}$

where I_B are the pixel readings from the sample image and I_A are the pixel readings from the reference image.

27. The method of claim 24, further comprising filtering the radiation source such that only a desired set of wavelengths are captured in the sample image and the reference image.

28. The method of claim 24, further comprising rotating the image switching means to move between the first position and the second position.

29. The method of claim 24, further comprising synchronizing the capturing of the sample image and the reference image with movement of the image switching means.

30. The method of claim 24, wherein the sample image is collected while the biolayer is immersed in a liquid.

31. The method of claim 30, wherein the sample beam passes through the liquid.

32. The method of claim 30, wherein the sample beam does not pass through the liquid.

33. The method of claim 24, wherein

capturing the sample image comprises capturing a multiple exposure image of the biolayer as it traverses the field of view of the pixel array, and

capturing the reference image comprises capturing a multiple exposure image of the reference surface as it traverses the field of view of the pixel array.

34. The method of claim 24, wherein

capturing the sample image comprises capturing a time lapse image of the biolayer as it traverses the field of view of the pixel array, and

capturing the reference image comprises capturing a time lapse image of the reference surface as it traverses the field of view of the pixel array.