E). For statistical analysis, the student's t-test for independent sam…

E). For statistical analysis, the student's t-test for independent samples was used with a significance level a = 0.05 to compare gene expression between samples. Statistical tests were performed using GraphPad Prism software (La Jolla, CA).Western blot for activated NotchResultsCase report of patient NR with Invasive Right Maxillary SCCFor western blots, 50 g of protein from sonicated whole cell lysates were fractionated in a 10 Tris-glycine polyacrylamide gel, electrotransferred to PDVF membranes, and probed overnight with primary antibody for activated Notch1 (clone Val1744) (Cell Signaling, Danvers, MA). Blots were stripped and reprobed for GAPDH (clone FL-335) (Santa Cruz Biotech), to normalize the amount of sample loaded. Horseradish peroxidase-conjugated secondary antibodies (Caltag, Burlingame, CA) were then applied, followed by signal detection using Immobilon Western Chemiluminescent HRP Substrate (Millipore, Billerica, MA).Microarray gene expression profilingTotal RNA from USC-HN1 was collected and analyzed by microarray as previously described [6]. Briefly, 5 g of total RNA was reverse-transcribed using 5'-aminomodified primers with amino-allyl-dUTP. cDNA synthesized from USC-HN1 cells was labeled with Cy5 dye, and cDNA from universal RNA (uRNA) labelled with Cy3 dye. Labeled and combined cDNA probes were denatured, mixed in SlideHyb #1 hybridization buffer (Ambion, Austin, TX), and placed onto microarray slides. Thirty five thousand oligonucleotide arrays were hybridized at 42 in MAUI hybridization system (BioMicro Systems) for overnight, washed with 1 ?SSC with 0.05 SDS and 0.1 ?SSC buffers, and then were quickly spin-dried. Microarray slides were scanned on a GenePix 4000B scanner (Axon Instruments, Inc., Foster City, CA) with a 5 m resolution. Scanned raw images were analyzed and data files were generated with GenePix Pro 5.1 (Axon) software. For analysis, data files were uploaded into mAdb (microarray database), and analyzed by Capecitabine the software tools provided by the Center for Information Technology (CIT), NIH. A standard global normalization approach was used for each experiment. All of the extracted data was normalized using a 50 th percentile (median) normalization method. Statistical analyses and t-test were performed to identify differentially expressed genes.Patient NR is a 57-year-old Hispanic female with a past medical history of hypothyroidism and dyslipidemia. Of note, she does not have a family history of head and neck cancer nor does she have a history of tobacco or alcohol use. In February, 2009 she presented to her primary care physician with a three-month history of fatigue, weight loss, right-sided facial pain, oral ulcers, loose teeth, and bleeding gingiva. A right anterior maxillary gingival biopsy subsequently revealed a well-differentiated squamous cell carcinoma. A series of CTs, including the sinuses and neck, were performed which showed an extensive mass of the right hard and soft palates extending into the right maxillary sinus with significant bone destruction without noted nodal metastases (Stage IVa, T4aN0M0). In May, 2009 the patient underwent a complex resection and reconstruction procedure including a complete right PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/17139194 maxillectomy and ethmoidectomy with a split-thickness skin graft repair. Intraoperative frozen section margins of the maxillary crest, pterygoid plates, palate, ethmoid air cells, and buccal cavity were all clear and free of tumor. Final pathological diagnosis of the resected rig.