Detection of naphthalene in sea-water by a label-free plasmonic optical fiber biosensor.

In this study we developed an optical fiber biosensor able to detect the presence of naphthalene in sea-water. With this aim, we designed and produced an antibody specific for the naphthalene molecule. The capability of the antibody to bind to naphthalene was characterized by ELISA tests. A surface plasmon resonance (SPR) sensor platform was realized by sputtering a gold layer on a modified plastic optical fiber (POF). The gold surface was derivatizated and functionalized with the produced antibody by using the EDC/NHS amino-coupling immobilization protocol. The obtained results indicated that the POF-biosensor is able to sense the presence of naphthalene in a sea-water solution. The limit of detection (LOD) value was calculated to be 0.76 ng/mL, a value lower than the maximum residue limit value of naphthalene (0.13 µg/mL) referred as the water environmental quality standards (EQS). In addition, to the high sensitivity of the assay, it is remarkable to point out the possibility to monitor the presence of naphthalene in a real sea water solution by exploiting a simple experimental setup with a remote sensing capability offered by the POF-biosensor.

On the possibility of ephedrine detection: time-resolved fluorescence resonance energy transfer (FRET)-based approach.

Ephedrine is one of the main precursor compounds used in the illegal production of amphetamines and related drugs. Actually, conventional analytical methods such as high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), and gas chromatography-mass spectrometry (GC-MS) are used for the detection of ephedrine; sadly, these methods require qualified personnel and are time-consuming and expensive. In order to overcome these problems, in recent years, different methods have been developed based on the surface plasmon resonance (SPR) and electrochemical method. In this work, we present a simple, rapid, and effective method to detect the presence of ephedrine in solution, based on competitive fluorescence resonance energy transfer (FRET) assay. The antibody anti-ephedrine and ephedrine derivative were produced and labeled respectively, with two different fluorescent probes (donor and acceptor). The change in FRET signal intensity between donor and acceptor ephedrine compounds gives the possibility of detecting ephedrine traces of at least 0.81 ± 0.04 ppm (LOD). Graphical abstract A new Time-resolved Fluorescence Resonance Energy Transfer (FRET) assay for ephedrine detection.

Golgi membrane fission requires the CtBP1-S/BARS-induced activation of lysophosphatidic acid acyltransferase δ.

Membrane fission is an essential cellular process by which continuous membranes split into separate parts. We have previously identified CtBP1-S/BARS (BARS) as a key component of a protein complex that is required for fission of several endomembranes, including basolateral post-Golgi transport carriers. Assembly of this complex occurs at the Golgi apparatus, where BARS binds to the phosphoinositide kinase PI4KIIIß through a 14-3-3γ dimer, as well as to ARF and the PKD and PAK kinases. We now report that, when incorporated into this complex, BARS binds to and activates a trans-Golgi lysophosphatidic acid (LPA) acyltransferase type δ (LPAATδ) that converts LPA into phosphatidic acid (PA); and that this reaction is essential for fission of the carriers. LPA and PA have unique biophysical properties, and their interconversion might facilitate the fission process either directly or indirectly (via recruitment of proteins that bind to PA, including BARS itself).

A novel fluorescence polarization assay for determination of penicillin G in milk.

Detection of penicillin G in milk is of interest because of the wide use of this antibiotic in livestock. Current analytical methods used to quantify the penicillin G in milk are based on HPLC, mass spectrometry and electrophoresis. These methodologies are time-consuming and require trained personnel. In addition, it is not easy to envisage the development of a portable device for in situ analyses based on these methods. We present a novel sensing approach for detecting the presence of penicillin G in milk. The proposed method is based penicillin G conjugate labeled with red-emitting dye with properly produced anti-penicillin G antibodies. The results obtained suggest our method could be applied directly in milk without interference from other substances. The limit of detection of the method was 1.0 nmol/L, which is much less than the required MRL in EU regulations (12.0 nmol/L).

A Rapid and Sensitive Assay for the Detection of Benzylpenicillin (PenG) in Milk.

Antibiotics, such as benzyl-penicillin (PenG) and cephalosporin, are the most common compounds used in animal therapy. Their massive and illegal use in animal therapy and prophylaxis inevitably causes the presence of traces in foods of animal origin (milk and meat), which creates several problems for human health. With the aim to prevent the negative impact of ß-lactam and, in particular, PenG residues present in the milk on customer health, many countries have established maximum residue limits (MRLs). To cope with this problem here, we propose an effective alternative, compared to the analytical methods actually employed, to quantify the presence of penicillin G using the surface plasmon resonance (SPR) method. In particular, the PenG molecule was conjugated to a protein carrier to immunize a rabbit and produce polyclonal antibodies (anti-PenG). The produced antibodies were used as molecular recognition elements for the design of a competitive immune-assay for the detection of PenG by SPR experiments. The detection limit of the developed assay was found to be 8.0 pM, a value much lower than the MRL of the EU regulation limit that is fixed at 12 nM. Thus, our results clearly show that this system could be successfully suitable for the accurate and easy determination of PenG.

Antigen delivery by filamentous bacteriophage fd displaying an anti-DEC-205 single-chain variable fragment confers adjuvanticity by triggering a TLR9-mediated immune response.

Filamentous bacteriophage fd particles delivering antigenic determinants via DEC-205 (fdsc-αDEC) represent a powerful delivery system that induces CD8(+) T-cell responses even when administered in the absence of adjuvants or maturation stimuli for dendritic cells. In order to investigate the mechanisms of this activity, RNA-Sequencing of fd-pulsed dendritic cells was performed. A significant differential expression of genes involved in innate immunity, co-stimulation and cytokine production was observed. In agreement with these findings, we demonstrate that induction of proinflammatory cytokines and type I interferon by fdsc-αDEC was MYD88 mediated and TLR9 dependent. We also found that fdsc-αDEC is delivered into LAMP-1-positive compartments and co-localizes with TLR9. Thus, phage particles containing a single-strand DNA genome rich in CpG motifs delivered via DEC-205 are able to intercept and trigger the active TLR9 innate immune receptor into late endosome/lysosomes and to enhance the immunogenicity of the displayed antigenic determinants. These findings make fd bacteriophage a valuable tool for immunization without administering exogenous adjuvants.

Long-distance FRET analysis: a Monte Carlo simulation study.

A new method for extending the utilizable range of Förster resonance energy transfer (FRET) is proposed and tested by the Monte Carlo technique. The obtained results indicate that the efficiency of FRET can be significantly enhanced at a given distance if the energy transfer takes place toward multiple acceptors that are closely located on a macromolecule instead of a single acceptor molecule as it is currently used in FRET analysis. On the other hand, reasonable FRET efficiency can be obtained at significantly longer distances than in the case of a single acceptor. Randomly distributed and parallel orientated acceptor transition moments with respect to the transition moment of the donor molecule have been analyzed as two extreme cases. As expected, a parallel orientation of donor and acceptor transition moments results in a more efficient excitation energy transfer. This finding could be used to directly reveal the assembly/deassembly of large protein complexes in a cell by fluorescence microscopy.