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

A method for identifying a particular volume of material, wherein the material contains at least one photo-active molecular agent, the method comprising the steps of:

• (a) treating the particular volume of the material with light sufficient to promote a simultaneous two-photon excitation of at least one of the at least one photo-active molecular agent contained in the particular volume of the material;
• (b) photo-activating the at least one photo-active molecular agent in the particular volume of the material, thereby producing at least one photo-activated molecular agent, wherein the at least one photo-activated molecular agent emits energy;
• (c) detecting the energy emitted by the at least one photo-activated molecular agent;
• (d) producing a detected energy signal which is characteristic of the particular volume of the material;
• and
• (e) identifying the particular volume of the material based on said detected energy signal.

A method for medical diagnostic identification of tissue comprising the steps of:

• introducing a selected photo-active molecular agent into a tissue, said agent being selected for a specific feature of interest, said agent being susceptible of two-photon excitation;
• allowing said agent to accumulate in said specific feature of interest;
• directing light to regions of interest within the tissue, including regions substantially below a tissue surface and including at least a portion of said specific feature of interest, said light being selected in frequency and energy to penetrate the tissue and to promote two-photon excitation substantially only at locations within confocal region of said light;
• controlling the locations of said confocal region over a range of depths within said tissue;
• photo-activating, using two-photon excitation, any of said photo-active agent present at said confocal region over said range of depths, wherein the photo-activated agent emits energy;
• detecting the emitted energy;
• producing a detected energy signal which is characteristic tissue at the confocal region;
• and
• identifying the tissue based on said detected energy signal.

A method for a material, the material including at least one photo-active molecular agent, the method comprising:

• encoding light from a light source with a modulation pattern to produce a modulated light;
• treating the material with said modulated light to promote simultaneous two-photon excitation of at least one photo-active molecular agent so that said at least one excited molecular agent becomes photo-activated in said material and emits a modulated energy;
• detecting the portion of the modulated emitted energy;
• producing a detected modulated energy signal which is characteristic of the material;
• and
• identifying the material based on said detected modulated energy signal.

A method for identifying a particular volute of tissue, wherein the tissue includes at least one photo-active molecular agent, the method comprising:

• encoding light from a light source with a modulation pattern to produce a modulated light;
• directing light to specific regions of interest within the tissue, including regions substantially below a tissue surface, said light being selected to penetrate the tissue and to promote two-photon excitation substantially only at locations within a confocal region;
• controlling the locations of said confocal region over a range of depths within said tissue;
• using two-photon excitation, photo-activating said agent over said range of depths within said tissue, so that said at least one excited molecular agent becomes photo-activated substantially only at the confocal region, wherein said photo-activated agent emits a modulated energy;
• detecting a potion of the emitted modulated energy;
• producing a detected modulated energy signal which is characteristic of the tissue;
• and
• identifying the tissue based on said detected modulated energy signal.

A method for the diagnostic identification of tissue, the tissue having a surface, the tissue being relatively transparent to light having preselected characteristics, the method comprising the steps of:

• introducing a selected photo-active agent into a tissue, said agent being susceptible to two-photon excitation;
• allowing said agent to accumnlate at features of interest, if any, within said tissue;
• operating a laser to obtain therefrom a beam of light having said preselected characteristics;
• directing said laser beam to specific regions of interest within the tissue, including regions substantially below the tissue surface, including penetrating the tissue with said beam and promoting two-photon excitation of said agent substantially only at locations within a confocal region;
• moving the locations of said confocal region over a cross sectional area located at a range of depths within said tissue thereby to define an examined volume;
• using two-photon excitation, photo-activating any of said agent which has accumulated at any said feature of interest within said examined volume through which said confocal region passes, thereby producing a photo-activated agent at each said feature of interest when said confocal region intersects said feature of interest, wherein said photo-activated agent emits energy;
• detecting the emitted energy;
• producing a detected energy signal that is characteristic of tissue at the confocal region;
• and
• identifying the tissue based on said detected energy signal.