HighlightsPosterior reversible encephalopathy syndrome, syndrome characterized by headache, confusion, visual loss and seizures.Mesenteric leiomyosarcoma causing Posterior reversible encephalopathy syndrome.Symptoms disappearance after resection of the tumor, suggests a renin production cessation.
Background: Formalin fixed paraffin embedded (FFPE) tissues remains the standard method for fixation and storage of tissues for clinical pathology use. Protein extraction from these tissues remains challenging due to the reduced quality and amount of extracted proteins. Despite multiple successful attempts, isolation of proteins from FFPE tissue sections necessitates routine use of xylene, a highly toxic organic solvent. We previously showed that proteins can be efficiently extracted with a novel technique that utilizes hot distilled water as a substitute for xylene with a quality adequate for western blot analysis. However, its major drawback is the need to use an entire or major part of the paraffin-embedded tissue block. To address these issues we developed a new xylene-free method for protein extraction from FFPE tissue sections of around 8 μm thickness.
Methods: A total of 44 different types of FFPE tissues sections of 8 μm thickness were obtained from various archived FFPE specimens which include: 17 colorectal cancer (5-6 years), 7 breast cancer (3 years), 3 thyroid cancer (2 years), 4 ovarian cancer (2 years), 8 uterine cancer (1 year), and 5 prostate cancer (1 year). Deparaffinization was conducted by gentle treatment of each section with hot distilled water (≈90°C) for less than 10 seconds. Deparaffinized tissues were then placed in a cell lysis buffer and Laemli buffer and incubated at 100°C for 5-10 minutes. The extracted proteins were quantified using NanoDrop Spectrophotometer and evaluated using western blot analysis for the presence of AKT and beta-actin.
Results: Using this method, a significant amount of proteins was successfully isolated with an average amount of 2.670 μg/μl ± 0.623 or 1068 μg/8 μm tissue section. Compared to the standard protein extraction method using xylene the amount of proteins extracted in this experiment is at least four times greater. Protein extracts were of good quality and efficiently analyzed by western blot experiment. Moreover, the isolated proteins showed a similar migration pattern compared to the positive control protein on SDS-PAGE but with better bands’ intensity and clarity. Protein kinase B (PKB/AKT) was successfully identified in all specimens, and beta-actin protein was resolved with an efficiency higher than 80%. Hence, this technique has enabled us to efficiently extract and detect selected proteins from archived samples for up to 6 years.
Conclusion: We developed an efficient, safe, cost-effective, and rapid method to isolate proteins from FFPE tissues sections and adequate for western blot analysis. Hence, utilizing this technique can further aid in the identification of tissue protein biomarkers of various diseases, monitor cancer progression, and improve the diagnosis. It remains to be determined if this method of protein extraction form FFPE is adequate for use in proteomic analyses.
Citation Format: Anthony Mansour, Noha Bejjani, Carole Dagher, Rajaa Chatila, Wissam H. Faour. Optimized xylene-free protein extraction method from formalin-fixed paraffin-embedded tissue sections for western blot analysis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-288. doi:10.1158/1538-7445.AM2015-LB-288
BackgroundGestational trophoblastic neoplasms (GTNs) encompass a wide spectrum of diseases, of which choriocarcinoma is one of the most common. Choriocarcinoma occurs mainly in relation to pregnancy and rarely after the menopause. It has the potential to metastasize to organs other than the uterus.Case ReportWe describe a 62-year-old woman who presented with postmenopausal bleeding 11 years after the menopause. Pelvic ultrasound and abdominal/pelvic computerized tomography showed an intrauterine mass. Choriocarcinoma was diagnosed by Pipelle endometrial biopsy with positive staining for beta-human chorionic gonadotropin (hCG) and KI 67 along with an elevated serum beta-hCG level. The tumor was managed with multiple cycles of multidrug chemotherapy and follow-up based on serum beta-hCG levels according to the guidelines of the International Federation of Gynecology and Obstetrics (FIGO).ConclusionThis case report highlights that choriocarcinoma, a tumor normally associated with pregnancy, can present after the menopause.
This a case of a 77 years old male heavy smoker, known to have Combined Pulmonary Fibrosis and Emphysema complicated by a primary invasive adenocarcinoma of the lung with bone metastasis, who presented with a two weeks history of right inguino-scrotal pain and swelling.Imaging studies revealed a right paratesticular formation that appeared to involve the epididymis and the scrotal wall.A biopsy of the mass showed morphological and Immunophenotypic features in favor of metastasis of an adenocarcinoma of the lung.Based on our literature review, there are only few published cases about scrotal wall metastasis of a lung primary.
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