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The capacity for removal of ochratoxin A (OTA) during alcoholic fermentation was evaluated in batch systems with one commercial strain and one wild strain of Saccharomyces cerevisiae. Batch alcoholic fermentations were carried out in yeast extract-malt extract broth (YM) medium, with 18.0% glucose and OTA added to final concentrations of 3.48 and 4.95 ng/mL respectively. The removal capacity of each yeast strain was examined after completion of fermentation in batch culture and after extended contact with yeast biomass. The removal capacity of the yeast strains was also examined in stationary phase cultures. Stationary phase yeasts were studied with biomass harvested from the stationary phase of anaerobic fermentation, by incubation in phosphate buffer, with the addition of 5.00 ng/ mL of OTA. Removal studies with stationary phase cells were performed with viable and non-viable cells inactivated with Na-azide. The study showed that in growing phase cultures, OTA removal was significant only after extended contact with yeast biomass; up to 29.7% and 25.4% for wild yeast ZIM 1927 and commercial yeast Lalvin EC-1118 respectively, but not during alcoholic fermentation. In stationary phase cultures, viable and non-viable cells were not significantly different in OTA removal from the medium. This demonstrated that OTA was not metabolised, but possibly adsorbed by the yeast cells. The presence of OTA in synthetic media influenced yeast metabolism, causing the production of higher volatile acidity by 0.08 and 0.13 g/L for Lalvin EC-1118 and ZIM 1927 respectively, and lower concentrations of reducing sugar, by 0.32 g/L, but only for ZIM 1927.

BackgroundDetection of circulating lung cancer cells with cancer-stem like characteristics would represent an improved tool for disease prognosis. However, current antibodies based methods have some disadvantages and therefore cell SELEX (Systematic Evolution of Ligands by Exponential Enrichment) was used to develop DNA aptamers, recognizing cell surface markers of non-small lung carcinoma (NSLC) cells.Materials and methodsThe human adenocarcinoma cell line A549 was used for selection in seven cell SELEX cycles. We used human blood leukocytes for negative selection, and lung stem cell protein marker CD90 antibody binding A549 cells for positive selection.ResultsThe obtained oligonucleotide sequences after the seventh SELEX cycle were subjected to in silico selection analysis based on three independent types of bioinformatics approaches, selecting two closely related aptamer candidates in terms of consensus sequences, structural motifs, binding affinity (Kd) and stability (ΔG). We selected and identified the aptamer A155_18 with very good binding characteristics to A459 cells, selected for CD90 antibody binding. The calculated phylogenetic tree showed that aptamers A155_18 and the known A549 cell aptamer S6 have a close structural relationship. MEME sequence analysis showed that they share two unique motifs, not present in other sequences.ConclusionsThe novel aptamer A155_18 has strong binding affinity for A549 lung carcinoma cell line subpopulation that is expressing stem cell marker CD90, indicating a possible stemness, characteristic for the A459 line, or a subpopulation present within this cell line. This aptamer can be applied as diagnostic tool, identifying NSLC circulating cells.

Abstract:The availability of devices for cancer biomarker detection at early stages of the disease is one of the most critical issues in biomedicine. Towards this goal, to increase the assay sensitivity, device miniaturization strategies empowered by the employment of high affinity protein binders constitute a valuable approach. In this work we propose two different surface-based miniaturized platforms for biomarker detection in body fluids: the first platform is an atomic force microscopy (AFM)-based nanoarray, where AFM is used to generate functional nanoscale areas and to detect biorecognition through careful topographic measurements; the second platform consists of a miniaturized electrochemical cell to detect biomarkers through electrochemical impedance spectroscopy (EIS) analysis. Both devices rely on robust and highly-specific protein binders as aptamers, and were tested for thrombin detection. An active layer of DNA-aptamer conjugates was immobilized via DNA directed immobilization on complementary single-stranded DNA self-assembled monolayers confined on a nano/micro area of a gold surface. Results obtained with these devices were compared with the output of surface plasmon resonance (SPR) assays used as reference. We succeeded in capturing antigens in concentrations as low as a few nM. We put forward ideas to push the sensitivity further to the pM range, assuring low biosample volume (µL range) assay conditions.

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and its value as a prognostic biomarker in patients with metastatic nonesmall cell lung cancer (NSCLC). Method: The clinical chart of consecutive metastatic NSCLC patients treated in Edgardo Rebagliati Martins National Hospital, Lima-Perú, between July 2014 and December 2015, were retrospectively evaluated. Epidemiological, disease and extension data were collected, as well as white cell differential before either, definitive treatment or best supportive care. Survival analysis was performed using log-rank test, Kaplan-Meier method and Cox regression analysis using NLR cutoff point of 5 as previously reported. R language was used for statistical analysis. Results: Ninety clinical charts of advanced NSCLC patients were evaluated, of which 36 cases were considered for final analysis. The mean age was 69 years (SD 11.9). Twenty-three patients were female (63.9%), 28 were nonsmokers (77.8%) and 32 had adenocarcinoma (88.9%), median NLR was 3.4. The median overall survival (OS) was 7.95 months. Median OS for patients with NLR 3 5 was 3.97 months vs. 12.07 months for NLR < 5 (p ¼ 0.0041). Cox analysis for NLR < 5 was performed, adjusting with variables such as age (HR: 0.27, p ¼ 0.008), gender (HR: 0.30, p ¼ 0.012) and systemic treatment (HR: 0.34, p ¼ 0.038). Finally, we performed multivariate analysis adjusting for all variables that potentially can influence in mortality such as age, gender, systemic treatment and metastatic sites and we found HR 0.27 for NLR < 5 (95%CI 0.09-0.84, p ¼ 0.024). Conclusion: NLR < 5 was statistically associated with better overall survival. Multivariate analysis adjusted by age, gender, systemic treatment and metastatic compromise, was able to predict better overall survival, with a hazard ratio of 0.27 for NLR < 5. The retrospective design and limitations of our study only allow us to generate the hypothesis that NLR < 5 could be an easy and inexpensive marker of better survival in metastatic lung cancer patients and support design of larger and prospective trials.