Methods for improved selectivity in photo-activation and detection of molecular diagnostic agents

• US 7,346,387 B1
• Filed: 05/05/1998
• Issued: 03/18/2008
• Est. Priority Date: 10/30/1996
• Status: Expired due to Fees

• Current Assignee: Provectus Devicetech, Inc.
• Original Assignee: Provectus Devicetech, Inc.
• Inventors: Wachter, Eric A., Dees, H. Craig, Fisher, Walter G.
• Primary Examiner(s): Winakur; Eric F.
• Assistant Examiner(s): ROZANSKI, MICHAEL T
• Application Number:
  US09/072,963
• Time in Patent Office: 3,605 Days
• Field of Search: 600/473, 600/310, 600/407, 600/476, 356/317, 356/300, 356/318, 250/458.1, 250/459.1, 372/4, 372/9
• US Class Current: 600/476
• CPC Class Codes: A61B 2017/00057 Light
• CPC Class Codes: A61K 41/008 Two-Photon or Multi-Photon PDT, e.g. with upconverting dyes or photosensitisers
• CPC Class Codes: A61K 49/0017 Fluorescence in vivo
• CPC Class Codes: A61K 49/0039 Coumarin dyes
• CPC Class Codes: A61K 49/0043 Fluorescein, used in vivo
• CPC Class Codes: A61N 5/062 Photodynamic therapy, i.e. excitation of an agent
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors

Abstract

A method for the imaging of a particular volume of plant or animal tissue, wherein the plant or animal tissue contains at least one photo-active molecular agent. The method comprises the steps of treating the particular volume of the plant or animal tissue with light sufficient to promote a simultaneous two-photon excitation of the photo-active molecular agent contained in the particular volume of the plant or animal tissue, photo-activating at least one of the at least one photo-active molecular agent in the particular volume of the plant or animal tissue, thereby producing at least one photo-activated molecular agent, wherein the at least one photo-activated molecular agent emits energy, detecting the energy emitted by the at least one photo-activated molecular agent, and producing a detected energy signal which is characteristic of the particular volume of plant or animal tissue. The present invention also provides a method for the imaging of a particular volume of material, wherein the material contains at least one photo-active molecular agent.

Claims

1. Imaging apparatus adapted for in vivo imaging a particular volume of plant or animal tissue containing at least one photo-active molecular agent, the apparatus comprising:

a source of collimated light, said light having a frequency effective to penetrate substantially into the tissue, said light being adapted to promote simultaneous two-photon excitation (TPE) of the molecular agent contained within the tissue;

beam expanding apparatus for expanding said collimated light into an expanded beam of collimated light;

focusing apparatus for focusing the expanded beam of collimated light throughout a range of focal lengths extending from a surface of said tissue to a depth substantially beyond said surface, said light source and focusing apparatus cooperating to promote TPE of the molecular agent,

wherein a focal point or focal plane is adjustable over a distance of up to 100 mm with respect to said tissue;

a detector located proximate to the tissue and positioned to detect said light emitted by the molecular agent and which travels a path that does not retrace an optical path of the light incident on the tissue, said detector configured to produce a detected signal characteristic of the particular volume at which the light source has been focused;

a processor coupled to said detector; and

a modulation system associated with said light source, said processor being coupled to said modulation system and adapted to produce a demodulated energy signal which is characteristic of the particular photo-activated molecular agent.

2. The apparatus of claim 1, wherein said light source produces a pulsed output having a pulse repetition frequency above about 75 megahertz and a sub-nanosecond pulse duration.

3. The apparatus of claim 2 wherein said light source produces near-infrared light.

4. The apparatus of claim 3 wherein said light source produces pulse energies of about 20 nanojoules.

5. The apparatus of claim 3 wherein said light source comprises a laser.

6. Apparatus adapted for in vivo medical diagnostic imaging comprising:

light source means for directing a confined light at and into tissue to be imaged, said light being selected in frequency and energy to penetrate below a tissue surface and to promote TPE substantially only at a confocal region at a distance of up to 100 mm below said tissue surface, said light source means including beam expanding means and focusing means cooperating together to shape said confocal region;

means for varying a position of a confocal region of the light within a range of depths in the tissue to be imaged;

detector means positioned to receive and detect isotropic radiation emitted by a photo-activated molecular agent within the tissue after said agent has been excited using two-photon excitation;

a modulator means for modulating said light with a type of modulation and producing a modulated light; and

a processor coupled to said detector, said processor being coupled to said modulation system and adapted to produce a demodulated energy signal which is characteristic of said photo-activated molecular agent.

7. The apparatus of claim 6:

wherein the light source means includes means for producing a collimated light beam; and

wherein the light source means includes focusing means for focusing the collimated light beam to a confocal region located with tissue at a point below the tissue surface.

8. The apparatus of claim 7 wherein the means for producing a collimated light beam comprises a pulsed laser operating the near infra-red spectrum.

9. The apparatus of claim 6 wherein said imaging apparatus is a microscope.

10. An apparatus adapted for in vivo microscopic imaging to a distance of up to 100 mm within tissue comprising:

a light source, said light source producing a beam of light directed to or into a material to be imaged, wherein said light source includes beam expanding means and beam focusing means cooperating together to shape said beam of light into a confocal region, said light being selected in frequency and energy to penetrate into or below a surface of the material and to promote two-photon excitation substantially only in a confocal region to be imaged;

a modulator, said modulator cooperating with said light source to modulate said light with a type of modulation and produce a modulated light;

a detector positioned to receive and detect radiation emitted by a photo-activated molecular agent within the material after said agent has been excited using two-photon excitation; and

a demodulator, said demodulator coupled to said detector and producing a demodulated energy signal which is characteristic of the particular photo-activated molecular agent.

11. An apparatus according to claim 10 wherein said demodulator is a processor.

12. Apparatus adapted to in vivo medical diagnostic imaging comprising:

a light source, said light source producing light directed to or into deep tissue to be imaged, said light being selected in frequency and energy to penetrate into or below a surface of the tissue and to promote two-photon excitation substantially only in a region to be imaged;

a beam expanding apparatus adapted for expanding said light into an expanded beam of light;

a focusing apparatus adapted for focusing said expanded beam of light, said focusing apparatus being able to vary the position of the light within a range of depths in the region of tissue to be imaged, said range of depths extending up to 100 mm into said tissue to be imaged; and

a detector positioned to receive and detect radiation emitted by a photo-activated molecular agent within the material after said agent has been excited using two-photon excitation,

wherein said light source means includes means for producing a collimated light beam and focusing means for focusing the collimated light beam to a confocal region located with tissue at a point below the tissue surface.

13. An apparatus according to claim 12 wherein said light source produces near-infrared light.

14. An apparatus according to claim 13 wherein said light source is a laser.

15. An apparatus according to claim 12 further comprising a modulator, said modulator cooperating with said light to modulate said light with a type of modulation and produce a modulated light.

16. An apparatus according to claim 15 further comprising a demodulator, said demodulator coupled to said detector and producing a demodulated energy signal which is characteristic of the particular photo-activated molecular agent.

17. An apparatus according to claim 16, wherein said demodulator is a processor.

18. An apparatus according to claim 12 wherein said light source produces a pulsed output having a pulse repetition frequency above about 75 megahertz and a subnanosecond pulse duration.

19. An apparatus according to claim 12 wherein said light source produces pulse energies of about 20 nanojoules.