A comprehensive high-throughput FTIR spectroscopy-based method for evaluating the transfection event: estimating the transfection efficiency and extracting associated metabolic responses

Reporter genes are routinely used in every laboratory for molecular and cellular biology for studying heterologous gene expression and general cellular biological mechanisms, such as transfection processes. Although well characterized and broadly implemented, reporter genes present serious limitations, either by involving time-consuming procedures or by presenting possible side effects on the expression of the heterologous gene or even in the general cellular metabolism. Fourier transform mid-infrared (FT-MIR) spectroscopy was evaluated to simultaneously analyze in a rapid (minutes) and high-throughput mode (using 96-wells microplates), the transfection efficiency, and the effect of the transfection process on the host cell biochemical composition and metabolism. Semi-adherent HEK and adherent AGS cell lines, transfected with the plasmid pVAX-GFP using Lipofectamine, were used as model systems. Good partial least squares (PLS) models were built to estimate the transfection efficiency, either considering each cell line independently (R 2 ≥ 0.92; RMSECV ≤ 2 %) or simultaneously considering both cell lines (R 2 = 0.90; RMSECV = 2 %). Additionally, the effect of the transfection process on the HEK cell biochemical and metabolic features could be evaluated directly from the FT-IR spectra. Due to the high sensitivity of the technique, it was also possible to discriminate the effect of the transfection process from the transfection reagent on KEK cells, e.g., by the analysis of spectral biomarkers and biochemical and metabolic features. The present results are far beyond what any reporter gene assay or other specific probe can offer for these purposes.

Multi-strand narrative structures: a filmic game of multiple players

Session 7: Playing with Roles, images and improvising New States of Awareness, 3rd Global Conference, 1st November – 3rd November, 2014, Prague, Czech Republic.

The tricks of the trade (un)exposed

One of the filmic trends which has been neglected by the Academy Awards is the metacinema, which for practical purposes I will consider to be a cross between the complexities of the self-reflexive cinema (highly connoted with modernism) and the Hollywood Film (the classical films about the urge to ‘make it’ in Hollywood). Indeed, these films have always existed and some, as Sunset Boulevard (Billy Wilder, 1950, USA) and Mulholland Dr. (David Lynch, 2001, FRA/USA), have even made it to the ceremony, but were, predictably, defeated in the main categories, by other more ‘serious’ or less self-reflexive products. The United States has always insisted on not revealing the tricks of the trade while, ironically, generating films that deal with this theme, in order to cater to the curiosity of the metacinema-inclined spectator. For this reason such films are usually about the universe of cinema but not its medium, at least not in a way that discloses the operations of the technical apparatus. Why are these films not viewed as serious enough and artistic enough to be awarded Oscars by the Academy in the categories of Best Picture, Best Director, Best Screenplay, and Best Cinematography? Are they being discarded for the same reasons that comedy and musicals usually are? Or are they being punished for being too unveiling? Or is the industry going for commercial products that can easily be pushed on a global scale and make a profit?

Planning smart cities using wireless low energy monitoring systems

Thesis to obtain the Master Degree in Electronics and Telecommunications Engineering

Maintenance of fire sprinkler systems based on the dynamic assessment of its condition

Safety is one of the major concerns of process safety engineers in most industrial facilities all over the world. To this scope, some events play an important role once the effect of their consequences can be assumed as totally undesirable. One of these events refers to the occurrence of a fire. Such event can result in catastrophic consequences for life, equipment, and continuity of activities or even leading to environmental damage. A fire protection equipment with low reliability means that this equipment are often unavailable and thus the risk of a fire increases. Maintenance of fire protection equipment is very important because this kind of systems is mostly in a dormant mode, which gives uncertainty about their operability when demanded in a real situation of fire. This article outlines the importance of tests, inspection, and maintenance operations in the context of a fire sprinkler system and proposes a methodology based on international standards and supported by test/inspection reports to correct the frequency of these actions according to the level of degradation of the components and regarding safety purposes. © 2015 American Institute of Chemical Engineers.