Biosensor immunoassay for traces of hazelnut protein in olive oil

The fraudulent addition of hazelnut oil to more expensive olive oil not only causes economical loss but may also result in problems for allergic individuals as they may inadvertently be exposed to potentially allergenic hazelnut proteins. To improve consumer safety, a rapid and sensitive direct biosensor immunoassay, based on a highly specific monoclonal antibody, was developed to detect the presence of hazelnut proteins in olive oils. The sample preparation was easy (extraction with buffer); the assay time was fast (4.5 min only) and the limit of detection was low (0.08 μg/g of hazelnut proteins in olive oil). Recoveries obtained with an olive oil mixed with different amounts of a hazelnut protein containing hazelnut oil varied between 93% and 109%

Peptidomics characterization of allergenic and non-allergenic tropomyosin orthologs

Background The cleavage and the digestion patterns of allergenic proteins play a key role in allergenicity. The transformation of food proteins starts with their denaturation by the acidic environment of the stomach. However, protein denaturation is not enough to completely remove allergenic properties of a protein, but it is necessary a complete enzymatic digestion. Whether the enzymatic digestion is not efficient, the persistence of bigger peptides can occur and put the basis for the development of the sensitization process.The hypothesis of the current work takes into consideration the probability that shrimp tropomyosin (TM) is not fully digested or presents a digestion pattern that generates some peptides that can be immunogenic.Therefore, the work plan designed, aims to study the cleavage pattern of: purified chicken TM, recombinant chicken TM, purified TM of Penaeus monodon (Pen m 1), recombinant TM of Penaeus monodon (rPen m 1) and recombinant TM of Crangon crangon (rCrac c 1). Methods One mg of each ortholog has been processed through simulated mouth, gastric and intestinal digestion. The sample was frozen to block the digestion and, after this step, concentrated and cleaned through protein precipitation. The protein pellet was processed for peptidomic analysis through 1D Tricine gel electrophoresis, 2D Tricine gel electrophoresis and mass spectrometry. Results Simulated gastric digestion pattern of shrimp TM highlighted the presence of a resistant band at an average MW of 25 kDa that could be involved in the immunogenic process. Conclusions This innovative approach (peptidomics study through 1D-2D Tricine/MS) could represent a milestone for the study of digestion patterns of allergenic proteins or for the study of allergenic potential of novel foods

Sensitivity to inhibition of DNA repair by Olaparib in novel oropharyngeal cancer cell lines infected with Human Papillomavirus

The incidence of Human Papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OPSCC) is increasing rapidly in the UK. Patients with HPV-positive OPSCC generally show superior clinical responses relative to HPV-negative patients. We hypothesised that these superior responses could be associated with defective repair of DNA double strand breaks (DSB). The study aimed to determine whether defective DNA repair could be associated with sensitivity to inhibition of DNA repair using the PARP inhibitor Olaparib. Sensitivity to Olaparib, and induction and repair of DNA damage, were assessed in a panel of 8 OPSCC cell-lines, including 2 novel HPV-positive lines. Effects on cell cycle distribution and levels of PARP1 and p53 were quantified. RNA-sequencing was used to assess differences in activity of DNA repair pathways. Two HPV-positive OPSCC lines were sensitive to Olaparib at potentially therapeutic doses (0.1–0.5 μM). Two HPV-negative lines were sensitive at an intermediate dose. Four other lines, derived from HPV-positive and HPV-negative tumours, were resistant to PARP inhibition. Only one cell-line, UPCISCC90, showed results consistent with the original hypothesis i.e. that in HPV-positive cells, treatment with Olaparib would cause accumulation of DSB, resulting in cell cycle arrest. There was no evidence that HPV-positive tumours exhibit defective repair of DSB. However, the data suggest that a subset of OPSCC may be susceptible to PARP-inhibitor based therap

Standardization of double blind placebo controlled food challenge with soy within a multicentre trial

Background: Multicentre trials investigating food allergies by double blind placebo controlled food challenges (DBPCFC) need standardized procedures, challenge meals and evaluation criteria. We aimed at developing a standardized approach for identifying patients with birch related soy allergy by means of DBPCFC to soy, including determination of threshold levels, in a multicentre setting. Methods: Microbiologically stable soy challenge meals were composed of protein isolate with consistent Gly m 4 levels. Patients sensitized to main birch allergen Bet v 1 and concomitant sensitization to its soy homologue Gly m 4 underwent DBPCFC. Outcome was defined according to presence and/or absence of ten objective signs and intensity of eight subjective symptoms as measured by visual analogue scale (VAS). Results: 138 adult subjects (63.8% female, mean age 38 years) underwent DBPCFC. Challenge meals and defined evaluation criteria showed good applicability in all centres involved. 45.7% presented with objective signs and 65.2% with subjective symptoms at soy challenge. Placebo challenge meals elicited non-cardiovascular objective signs in 11.6%. In 82 (59.4%) subjects DBPCFC was judged as positive. 70.7% of DPBCFC+ showed objective signs and 85.4% subjective symptoms at soy challenge. Subjective symptoms to soy challenge meal in DBPCFC+ subjects started at significantly lower dose levels than objective signs (p < 0.001). Median cumulative eliciting doses for first objective signs in DBPCFC+ subjects were 4.7 g [0.7–24.7] and 0.7 g [0.2–4.7] total soy protein for first subjective symptoms (p = 0.01). Conclusions: We present the hitherto largest group of adults with Bet v 1 and Gly m 4 sensitization being investigated by DBPCFC. In this type of food allergy evaluation of DBPCFC outcome should not only include monitoring of objective signs but also scoring of subjective symptoms. Our data may contribute to standardize DBPCFC in pollen-related food allergy in multicentre settings. Trial registration EudraCT: 2009-011737-27

Analytical criteria for performance characteristics of IgE binding methods for evaluating safety of biotech food products

There is detailed guidance on how to perform bioinformatic analyses and enzymatic degradation studies for genetically modified crops under consideration for approval by regulatory agencies; however, there is no consensus in the scientific community on the details of how to perform IgE serum studies. IgE serum studies are an important safety component to acceptance of genetically modified crops when the introduced protein is novel, the introduced protein is similar to known allergens, or the crop is allergenic. In this manuscript, we describe the characteristics of the reagents, validation of assay performance, and data analysis necessary to optimize the information obtained from serum testing of novel proteins and genetically modified (GM) crops and to make results more accurate and comparable between different investigation

Impact of Leather on the Fire Resistance of Leather-Wood Fibreboard: FT-IR Spectroscopy and Pyrolysis-GC-MS Investigation

Leather-wood fibreboards are innovative composite materials, which combine together the high mechanical properties of wood with the superior fire behaviour properties of leather. This study deals with the understanding of the combustion mechanism of the wet-white leather panel. During burning, an overlay coating-like surface is formed on top of a foamy structure that creates the heat transfer barrier. The FT-IR spectroscopy results of the leather show the rearrangement of the proteins and the formation of an increasing amount of acid groups when the exposure to hot gun at over 530°C was prolonged. These acid moieties can react with amino groups of other peptide chains, building a protective polymer network which hinders the oxygen to reach the core of the panel. Simultaneously, the gases produced during rearrangement cannot easily leave the material, producing a foamy structure which slows down the heat transfer to the core of the material. The Py-GC-MS analysis shows that the gases produced by the wet-white leather-type protein-based boards were amino-aromatic compounds like the diketopiperazine (DKP), which do not burn easily. The combination of the effects of (i) formation of the overlay coating-like surface, (ii) establishment of the foamy structure, and (iii) degassing of DKP explains the outstanding fire properties of leather and wood-leather fibreboards