Attofluor coverslip holder

All studies were performed in conjunction with University of South Alabama Institutional Review Board protocol: IRB # 445452-3 and conform to standard practices and guidelines. Colon tissue specimens were obtained from colorectal surgical resections from the University of South Alabama Medical Center Departments of Surgery and Pathology. Specimens were selected as pairs of cancerous and noncancerous tissues from adenocarcinoma and surrounding normal tissue, respectively. A total of 8 patients have been enrolled thus far in the study. However, data from the first 3 patients are not presented here, due to variations in specimen processing and imaging protocol. In addition, one patient was not considered, as surgical exam revealed extreme fibrosis, possibly due to perforation of the bowel, but no evidence of adenocarcinoma. Specimens were flash frozen in liquid nitrogen and pathology was confirmed by cryosection and hematoxylin and eosin (H&E) staining. Tissue specimens were maintained at −80 °C until they were prepared for imaging – usually within 1-2 days, but never more than 1 week. For imaging, tissues were reconstituted and suspended in cold 1X phosphate buffered saline (PBS) and placed onto a 25 mm round coverslip mounted in an AttoFluor coverslip holder (Life Technologies).

All animal procedures were carried out in accordance with institutional guidelines and were approved by the Institutional Animal Care and Use Committee of the University of South Alabama. Sprague-Dawley rat and C57BL/6 mice tissues were harvested from animals immediately following euthanasia, rinsed, placed in cold PBS, and taken directly to the microscope for imaging. Tissue samples were sliced into approximately 2 cm cubes for imaging to allow for sufficient light penetration and visual access to unique tissue morphology. Human colon samples were obtained from colorectal resections from the University of South Alabama Medical Center Departments of Surgery and Pathology. All human tissue specimens were obtained in accordance with protocols approved by the University of South Alabama Institutional Review Board. Tissue samples came as cancerous lesions and noncancerous surrounding normal tissue, later confirmed through pathology. Human tissue samples were placed in ice upon resection and imaged within 8 hours. All imaging was performed using a 25 mm round coverslip mounted in an AttoFluor coverslip holder (Life Technologies).

Human colon samples were obtained from colorectal resections from the University of South Alabama Medical Center Department of Surgery. All human tissue specimens were obtained in accordance with protocols approved by the University of South Alabama Institutional Review Board. Following resection, samples were sent to the University of South Alabama Medical Center Department of Pathology. Tissue samples obtained from pathology were separated into cancerous lesions and surrounding non-lesional tissue, and later confirmed by histology. Samples were placed on ice immediately after resection, rinsed with PBS, cut into approximately 2 cm cubes, and imaged within 8 hours. All imaging was performed using a 25 mm round coverslip mounted in an AttoFluor coverslip holder (Life Technologies). Each tissue sample scan included a minimum of three fields of view, not including the field of view used to generate the background spectrum.