Manual for starch gel electrophoresis: A method for the detection of genetic variation

The procedure to conduct horizontal starch gel electrophoresis on enzymes is described in detail. Areas covered are (I) collection and storage of specimens, (2) preparation of tissues, (3) preparation of a starch gel, (4) application of enzyme extracts to a gel, (5) setting up a gel for electrophoresis, (6) slicing a gel, and (7) staining a gel. Recipes are also included for 47 enzyme stains and 3 selected gel buffers. (PDF file contains 26 pages.

HIF1-alpha overexpression indicates a good prognosis in early stage squamous cell carcinomas of the oral floor

BACKGROUND: Hypoxia-inducible factor 1 (HIF-1) is a transcription factor, which plays a central role in biologic processes under hypoxic conditions, especially concerning tumour angiogenesis. HIF-1α is the relevant, oxygen-dependent subunit and its overexpression has been associated with a poor prognosis in a variety of malignant tumours. Therefore, HIF-1α expression in early stage oral carcinomas was evaluated in relation to established clinico-pathological features in order to determine its value as a prognostic marker. METHODS: 85 patients with histologically proven surgically treated T1/2 squamous cell carcinoma (SCC) of the oral floor were eligible for the study. Tumor specimens were investigated by means of tissue micro arrays (TMAs) and immunohistochemistry for the expression of HIF-1. Correlations between clinical features and the expression of HIF-1 were evaluated by Kaplan-Meier curves, log-rank tests and multivariate Cox regression analysis. RESULTS: HIF-1α was frequently overexpressed in a probably non-hypoxia related fashion. The expression of HIF-1α was related with a significantly improved 5-year survival rate (p < 0.01) and a significantly increased disease free period (p = 0.01) independent from nodal status and tumour size. In primary node negative T1/T2 SCC of the oral floor, absence of HIF-1α expression specified a subgroup of high-risk patients (p < 0.05). CONCLUSION: HIF-1α overexpression is an indicator of favourable prognosis in T1 and T2 SCC of the oral floor. Node negative patients lacking HIF-1α expression may therefore be considered for adjuvant radiotherapy

Long-term risk of adverse outcomes according to atrial fibrillation type

Sustained forms of atrial fibrillation (AF) may be associated with a higher risk of adverse outcomes, but few if any long-term studies took into account changes of AF type and co-morbidities over time. We prospectively followed 3843 AF patients and collected information on AF type and co-morbidities during yearly follow-ups. The primary outcome was a composite of stroke or systemic embolism (SE). Secondary outcomes included myocardial infarction, hospitalization for congestive heart failure (CHF), bleeding and all-cause mortality. Multivariable adjusted Cox proportional hazards models with time-varying covariates were used to compare hazard ratios (HR) according to AF type. At baseline 1895 (49%), 1046 (27%) and 902 (24%) patients had paroxysmal, persistent and permanent AF and 3234 (84%) were anticoagulated. After a median (IQR) follow-up of 3.0 (1.9; 4.2) years, the incidence of stroke/SE was 1.0 per 100 patient-years. The incidence of myocardial infarction, CHF, bleeding and all-cause mortality was 0.7, 3.0, 2.9 and 2.7 per 100 patient-years, respectively. The multivariable adjusted (a) HRs (95% confidence interval) for stroke/SE were 1.13 (0.69; 1.85) and 1.27 (0.83; 1.95) for time-updated persistent and permanent AF, respectively. The corresponding aHRs were 1.23 (0.89, 1.69) and 1.45 (1.12; 1.87) for all-cause mortality, 1.34 (1.00; 1.80) and 1.30 (1.01; 1.67) for CHF, 0.91 (0.48; 1.72) and 0.95 (0.56; 1.59) for myocardial infarction, and 0.89 (0.70; 1.14) and 1.00 (0.81; 1.24) for bleeding. In this large prospective cohort of AF patients, time-updated AF type was not associated with incident stroke/SE

Expression of hypoxia-inducible factor 1 alpha and its downstream targets in fibroepithelial tumors of the breast

INTRODUCTION: Hypoxia-inducible factor 1 (HIF-1) alpha and its downstream targets carbonic anhydrase IX (CAIX) and vascular endothelial growth factor (VEGF) are key factors in the survival of proliferating tumor cells in a hypoxic microenvironment. We studied the expression and prognostic relevance of HIF-1α and its downstream targets in phyllodes tumors and fibroadenomas of the breast. METHODS: The expression of HIF-1α, CAIX, VEGF and p53 was investigated by immunohistochemistry in a group of 37 primary phyllodes tumors and 30 fibroadenomas with known clinical follow-up. The tumor microvasculature was visualized by immunohistochemistry for CD31. Proliferation was assessed by Ki67 immunostaining and mitotic counts. Being biphasic tumors, immunoquantification was performed in the stroma and epithelium. RESULTS: Only two fibroadenomas displayed low-level stromal HIF-1α reactivity in the absence of CAIX expression. Stromal HIF-1α expression was positively correlated with phyllodes tumor grade (P = 0.001), with proliferation as measured by Ki67 expression (P < 0.001) and number of mitoses (P < 0.001), with p53 accumulation (P = 0.003), and with global (P = 0.015) and hot-spot (P = 0.031) microvessel counts, but not with CAIX expression. Interestingly, concerted CAIX and HIF-1α expression was frequently found in morphologically normal epithelium of phyllodes tumors. The distance from the epithelium to the nearest microvessels was higher in phyllodes tumors as compared with in fibroadenomas. Microvessel counts as such did not differ between fibroadenomas and phyllodes tumors, however. High expression of VEGF was regularly found in both tumors, with only a positive relation between stromal VEGF and grade in phyllodes tumors (P = 0.016). Stromal HIF-1α overexpression in phyllodes tumors was predictive of disease-free survival (P = 0.032). CONCLUSION: These results indicate that HIF-1α expression is associated with diminished disease-free survival and may play an important role in stromal progression of breast phyllodes tumors. In view of the absence of stromal CAIX expression in phyllodes tumors, stromal upregulation of HIF-1α most probably arises from hypoxia-independent pathways, with p53 inactivation as one possible cause. In contrast, coexpression of HIF-1α and CAIX in the epithelium in phyllodes tumors points to epithelial hypoxia, most probably caused by relatively distant blood vessels. On the other hand, HIF-1α and CAIX seem to be of minor relevance in breast fibroadenomas

Emerging concepts in biomarker discovery; The US-Japan workshop on immunological molecular markers in oncology

Supported by the Office of International Affairs, National Cancer Institute (NCI), the "US-Japan Workshop on Immunological Biomarkers in Oncology" was held in March 2009. The workshop was related to a task force launched by the International Society for the Biological Therapy of Cancer (iSBTc) and the United States Food and Drug Administration (FDA) to identify strategies for biomarker discovery and validation in the field of biotherapy. The effort will culminate on October 28th 2009 in the "iSBTc-FDA-NCI Workshop on Prognostic and Predictive Immunologic Biomarkers in Cancer", which will be held in Washington DC in association with the Annual Meeting. The purposes of the US-Japan workshop were a) to discuss novel approaches to enhance the discovery of predictive and/or prognostic markers in cancer immunotherapy; b) to define the state of the science in biomarker discovery and validation. The participation of Japanese and US scientists provided the opportunity to identify shared or discordant themes across the distinct immune genetic background and the diverse prevalence of disease between the two Nations

Population Genetic Structure and Life History Variability in \u3ci\u3eOncorhynchus nerka\u3c/i\u3e from the Snake River Basin

We used the variation at 64 allozyme loci to examine genetic relationships among 32 samples of sockeye salmon Oncorhynchus nerka and kokanee (resident sockeye salmon) from the Snake River basin and other North American locations. The genetic differentiation among populations was pronounced: Wright’s FST was higher (0.244) than has been reported in any other study of Pacific salmon. A detailed examination of the O. nerka from lakes in the Sawtooth Valley of Idaho was undertaken to help guide recovery planning for the endangered Redfish Lake population and to help resolve the relationships between the resident and anadromous forms. In Redfish Lake, adult sockeye salmon that returned in 1991–1993 were genetically distinct from local kokanee but similar to a small group of “residual” sockeye salmon discovered in the lake in 1992. This result is consistent with the hypothesis that the original sockeye salmon population was not extirpated by Sunbeam Dam early in this century. Populations of O. nerka that appear to be native to the Snake River were also found in Alturas Lake, Stanley Lake, and Warm Lake, although the latter two lakes also showed evidence of nonnative gene pools. Kokanee sampled from Pettit Lake are clearly the result of an introduction of late-spawning kokanee from northern Idaho, and we found evidence of two O. nerka gene pools in Wallowa Lake, both traceable to introductions of nonnative kokanee

Towards an integrated analytical technology for the generation of multidimensional protein expression maps

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Compensability index for compensation radiotherapy after treatment interruptions

<p>Abstract</p> <p>Background</p> <p>The goal of our work was to develop a simple method to evaluate a compensation treatment after unplanned treatment interruptions with respect to their tumour- and normal tissue effect.</p> <p>Methods</p> <p>We developed a software tool in java programming language based on existing recommendations to compensate for treatment interruptions. In order to express and visualize the deviations from the originally planned tumour and normal tissue effects we defined the compensability index.</p> <p>Results</p> <p>The compensability index represents an evaluation of the suitability of compensatory radiotherapy in a single number based on the number of days used for compensation and the preference of preserving the originally planned tumour effect or not exceeding the originally planned normal tissue effect. An automated tool provides a method for quick evaluation of compensation treatments.</p> <p>Conclusions</p> <p>The compensability index calculation may serve as a decision support system based on existing and established recommendations.</p