Devices and methods for optical pathology

Abstract

Currently most cancers, including breast cancers, are removed without any intraoperative margin control. Post-operative methods inspect 1-2% of the surgical margin and are prone to sampling errors. The present invention relates to an optical imaging system that will enable evaluation of the surgical margin in vivo and in real-time. The invention provides for simultaneous fluorescence and fluorescence polarization imaging. The contrast of the acquired images will be enhanced using fluorescent agents approved for diagnostic use in patients. As the staining pattern of fluorescence images is similar to that of histology, and the values of fluorescence polarization are significantly higher in cancerous as compared to normal cells, the invention provides for further improvements in diagnostic methods. The systems and methods can be applied to the intra-operative delineation of cancerous tissue.

1 . A method for interpretative histological imaging of a mammalian subject comprising:
 delivering a contrast agent to a region of tissue of a mammalian subject during a surgical procedure;   illuminating the region of tissue with at least first wavelength of light;   detecting an image of the region of tissue, the image having a field of view including a polarization component with a distribution of the contrast agent in the region of tissue identifying a location of cancerous tissue within the region of tissue;   selecting a sub-region within the field of view of the tissue,   illuminating the sub-region of tissue with light that is scanned across the sub-region;   detecting light from the scanned sub-region of tissue to form a confocal image of the sub-region; and   surgically removing the identified cancerous tissue from the imaged sub-region of the mammalian subject.   
     
     
         2 . The method of  claim 1  wherein the contract agent comprises methylene blue. 
     
     
         3 . The method of  claim 1  wherein the detected image comprises a fluorescence image. 
     
     
         4 . The method of  claim 1  further comprising using a first detector to detect the image of the polarization compound and a second detector to detect the confocal image. 
     
     
         5 . The method of  claim 4  wherein the step of using a first detector comprises detecting a first polarization component and a second polarization component. 
     
     
         6 . The method of  claim 1  further comprising detecting a reflectance image. 
     
     
         7 . The method of  claim 1  further comprising using a laser to illuminate the sub-region of tissue. 
     
     
         8 . The method of  claim 6  further comprising detecting the image including the polarization component with a first detector. 
     
     
         9 . The method of  claim 8  further comprising detecting a fluorescence image with the first fluorescence detector. 
     
     
         10 . The method of  claim 9  further comprising detecting a second fluorescence image including a second polarization component with a second fluorescence detector. 
     
     
         11 . The method of  claim 1  further comprising scanning the sub-region of tissue with illuminating light with a scanning mirror. 
     
     
         12 . The method of  claim 1  further comprising processing image data with a data processor. 
     
     
         13 . The method of  claim 1  further comprising a broadband light source. 
     
     
         14 . The method of  claim 1  further comprising coupling light from a light source onto the region of tissue with an optical system. 
     
     
         15 . The method of  claim 14  wherein the optical system comprises a polarization splitter to separate light from the tissue into a first polarization component and a second polarization component. 
     
     
         16 . The method of  claim 1  further comprising displaying a wide field image of the region of tissue on a display. 
     
     
         17 . The method of  claim 1  further comprising displaying an image of the sub-region of tissue on a display. 
     
     
         18 . The method of  claim 1  further comprising controlling scan parameters with a system controller. 
     
     
         19 . The method of  claim 1  further comprising imaging a wide field of view of the region of interest having an area of at least 4 cm 2 . 
     
     
         20 . The method of  claim 1  further comprising detecting abnormal cellular tissue in at least a single cell. 
     
     
         21 . The method of  claim 20  further comprising detecting cancer in the single cell. 
     
     
         22 . The method of  claim 20  wherein the cell is stained with methylene blue, tetracycline or toludine blue. 
     
     
         23 . The method of  claim 20  further comprising detecting a fluorescence polarization image of tumor tissue, fibroblasts or adipose tissue. 
     
     
         24 . The method of  claim 20  further comprising detecting co-polarized images and computing a quantitative value with a data processor. 
     
     
         25 . A method for interpretative histological imaging of a mammalian subject comprising:
 delivering a first stain into a region of tissue of a mammalian subject during a surgical procedure, the first stain having an affinity for an acidic component in the tissue;   delivering a second stain into the region of tissue of the mammalian subject during the surgical procedure, the second stain having an affinity for a basic component of the tissue;   illuminating the region of tissue with a first wavelength of light to induce fluorescence of the first stain;   illuminating the region of tissue with a second wavelength of light to induce fluorescence of the second stain;   detecting fluorescence from the region of tissue to form an image, the image having a distribution of the first stain and the second stain in the region of tissue identifying a location of cancerous tissue within the region of tissue; and   surgically removing the identified cancerous tissue from the mammalian subject.   
     
     
         26 . The method of  claim 25  wherein the first stain comprises TCN. 
     
     
         27 . The method of  claim 25  wherein the second stain comprises eosin. 
     
     
         28 . The method of  claim 25  further comprising using a first detector to detect this first stain and a second detector to detect a second stain. 
     
     
         29 . The method of  claim 28  wherein the step of using a first detector comprises detecting a first polarization component and a second polarization component. 
     
     
         30 . A system for interoperative imaging of a mammalian subject comprising:
 a wide field imaging system that images a region of tissue of a mammalian subject, the imaging system having a light source and a detector that detects a polarization component of light from a first contrast agent and a second contrast agent in the region tissue; and   a confocal imaging system that images a sub-region of tissue within the region of tissue imaged by the wide field imaging system.   
     
     
         31 . The system of  claim 30  further comprising a scanning system that scans illuminating light across the sub-region of tissue. 
     
     
         32 . The system of  claim 30  further comprising a data processor that processes image data. 
     
     
         33 . The system of  claim 30  further comprising a system controller that controls scan parameters of a light scanning system. 
     
     
         34 . The systems of  claim 32  wherein the data processor computes a quantitative fluorescence polarization value. 
     
     
         35 . The system of  claim 30  wherein the wide field imaging system detects one or more polarized fluorescence images. 
     
     
         36 . The system of  claim 30  further comprising a broadband light source. 
     
     
         37 . The system of  claim 36  further comprising an adjustable filter system that filters light from the broadband light source. 
     
     
         38 . The system of  claim 33  wherein the scanning systems is a moving mirror. 
     
     
         39 . The system of  claim 35  wherein the wide field imaging system detects co-polarized and cross-polarized fluorescence images of the first contrast agent and the second contrast agent, the first contrast agent having an acidic affinity characteristic and the second contrast agent having a basic affinity characteristic. 
     
     
         40 - 46 . (canceled) 
     
     
         47 . The method of  claim 1  further comprising imaging ductal carcinoma in breast tissue. 
     
     
         48 . The method of  claim 1  further comprising detecting an autofluorescence image of the tissue. 
     
     
         49 . (canceled) 
     
     
         50 . The method of  claim 1  further comprising registering a first image with a second image. 
     
     
         51 . The method of  claim 1  further comprising processing image data to generate a pseudo color image. 
     
     
         52 . (canceled) 
     
     
         53 . The method of  claim 1  further comprising detecting a plurality of fluorescence spectral bands. 
     
     
         54 . The method of  claim 1  further comprising illuminating the tissue with light that is scanned over a region of the cellular tissue. 
     
     
         55 . The method of  claim 54  further comprising using a scanning system to scan the tissue with light to generate confocal images of the tissue. 
     
     
         56 - 65 . (canceled)