Surface plasmon resonance biosensors for highly sensitive detection in real samples

In this work we summarize the main results obtained with the portable surface plasmon resonance (SPR) device developed in our group (commercialised by SENSIA, SL, Spain), highlighting its applicability for the real-time detection of extremely low concentrations of toxic pesticides in environmental water samples. In addition, we show applications in clinical diagnosis as, on the one hand, the real-time and label-free detection of DNA hybridization and single point mutations at the gene BRCA-1, related to the predisposition in women to develop an inherited breast cancer and, on the other hand, the analysis of protein biomarkers in biological samples (urine, serum) for early detection of diseases. Despite the large number of applications already proven, the SPR technology has two main drawbacks: (i) not enough sensitivity for some specific applications (where pM-fM or single-molecule detection are needed) (ii) low multiplexing capabilities. In order solve such drawbacks, we work in several alternative configurations as the Magneto-optical Surface Plasmon Resonance sensor (MOSPR) based on a combination of magnetooptical and ferromagnetic materials, to improve the SPR sensitivity, or the Localized Surface Plasmon Resonance (LSPR) based on nanostructures (nanoparticles, nanoholes,...), for higher multiplexing capabilities

African Swine Fever Virus Uses Macropinocytosis to Enter Host Cells

African swine fever (ASF) is caused by a large and highly pathogenic DNA virus, African swine fever virus (ASFV), which provokes severe economic losses and expansion threats. Presently, no specific protection or vaccine against ASF is available, despite the high hazard that the continued occurrence of the disease in sub-Saharan Africa, the recent outbreak in the Caucasus in 2007, and the potential dissemination to neighboring countries, represents. Although virus entry is a remarkable target for the development of protection tools, knowledge of the ASFV entry mechanism is still very limited. Whereas early studies have proposed that the virus enters cells through receptor-mediated endocytosis, the specific mechanism used by ASFV remains uncertain. Here we used the ASFV virulent isolate Ba71, adapted to grow in Vero cells (Ba71V), and the virulent strain E70 to demonstrate that entry and internalization of ASFV includes most of the features of macropinocytosis. By a combination of optical and electron microscopy, we show that the virus causes cytoplasm membrane perturbation, blebbing and ruffles. We have also found that internalization of the virions depends on actin reorganization, activity of Na+/H+ exchangers, and signaling events typical of the macropinocytic mechanism of endocytosis. The entry of virus into cells appears to directly stimulate dextran uptake, actin polarization and EGFR, PI3K-Akt, Pak1 and Rac1 activation. Inhibition of these key regulators of macropinocytosis, as well as treatment with the drug EIPA, results in a considerable decrease in ASFV entry and infection. In conclusion, this study identifies for the first time the whole pathway for ASFV entry, including the key cellular factors required for the uptake of the virus and the cell signaling involved

Lab-on-a-chip platforms based on highly sensitive nanophotonic Si biosensors for single nucleotide DNA testing

In order to solve the drawbacks of sensitivity and portability in optical biosensors we have developed ultrasensitive and miniaturized photonic silicon sensors able to be integrated in a "lab-on-a-chip" microsystem platform. The sensors are integrated Mach-Zehnder interferometers based on TIR optical waveguides (Si/SiO2/Si3N4) of micro/nanodimensions. We have applied this biosensor for DNA testing and for detection of single nucleotide polymorphisms at BRCA-1 gene, involved in breast cancer development, without target labeling. The oligonucleotide probe is immobilized by covalent attachment to the sensor surface through silanization procedures. The hybridization was performed for different DNA target concentrations showing a lowest detection limit at 10 pM. Additionally, we have detected the hybridization of different concentrations of DNA target with two mismatching bases corresponding to a mutation of the BRCA-1 gene. Following the way of the lab-on-a-chip microsystem, integration with the microfluidics has been achieved by using a novel fabrication method of 3-D embedded microchannels using the polymer SU-8 as structural material. The optofluidic chip shows good performances for biosensing

The C-type lectin homologue gene (EP153R) of African swine fever virus inhibits apoptosis both in virus infection and in heterologous expression

AbstractThe open reading frame EP153R of African swine fever virus (ASFV) encodes a nonessential protein that has been involved in the hemadsorption process induced in virus-infected cells. By the use of a virus deletion mutant lacking the EP153R gene, we have detected, in several virus-sensitive cells, increased levels of caspase-3 and cell death as compared with those obtained after infection with the parental BA71V strain. Both transient and stable expression of the EP153R gene in Vero or COS cells resulted in a partial protection of the transfected lines from the apoptosis induced in response to virus infection or external stimuli. The presence of gene EP153R resulted in a reduction of the transactivating activity of the cellular protein p53 in Vero cell cultures in which apoptosis was induced by virus infection or staurosporine treatment. This is to our knowledge the first description of a viral C-type lectin with anti-apoptotic properties

Label-free detection of exosomes using surface plasmon resonance biosensor

The development of a sensitive and specific detection platform for exosomes is highly desirable as they are believed to transmit vital tumour-specific information (mRNAs, microRNAs, and proteins) to remote cells for secondary metastasis. Herein, we report a simple method for the real-time and label-free detection of clinically relevant exosomes using a surface plasmon resonance (SPR) biosensor. Our method shows high specificity in detecting BT474 breast cancer cell-derived exosomes particularly from complex biological samples (e.g. exosome spiked in serum). This approach exhibits high sensitivity by detecting as low as 8280 exosomes/μL which may potentially be suitable for clinical analysis. We believe that this label-free and real-time method along with the high specificity and sensitivity may potentially be useful for clinical settings

Antiviral activity of lauryl gallate against animal viruses

BACKGROUND: Antiviral compounds are needed in the control of many animal and human diseases. METHODS: We analysed the effect of the antitumoural drug lauryl gallate on the infectivity of the African swine fever virus among other DNA (herpes simplex and vaccinia) and RNA (influenza, porcine transmissible gastroenteritis and Sindbis) viruses, paying attention to its effect on the viability of the corresponding host cells. RESULTS: Viral production was strongly inhibited in different cell lines at non-toxic concentrations of the drug (1-10 microM), reducing the titres 3->5 log units depending on the multiplicity of infection. In our model system (African swine fever virus in Vero cells), the addition of the drug 1 h before virus adsorption completely abolished virus productivity in a one-step growth virus cycle. Interestingly, no inhibitory effect was observed when lauryl gallate was added after 5-8 h post-infection. Both cellular and viral DNA synthesis and late viral transcription were inhibited by the drug; however, the early viral protein synthesis and the virus-mediated increase of p53 remained unaffected. Activation of the apoptotic effector caspase-3 was not detected after lauryl gallate treatment of Vero cells. Furthermore, the presence of the drug abrogated the activation of this protease induced by the virus infection. CONCLUSIONS: Lauryl gallate is a powerful antiviral agent against several pathogens of clinical and veterinary importance. The overall results indicate that a cellular factor or function might be the target of the antiviral action of alkyl gallates.This work was supported by grants from Ministerio de Educación y Ciencia, Spain (BFU2004-00298/BMC), Laboratorios del Dr Esteve, Barcelona, Spain and by institutional grants from Fundación Ramón Areces, Madrid, Spain and Banco Central Hispano, Madrid, Spain. CH was a fellow from Fundación Ramón Areces. AGG was funded by Centro de Investigación en Sanidad Animal (CISA), Valdeolmos, Spain.N

Current treatment goals are achieved by the majority of patients with atopic dermatitis treated with tralokinumab: results from a multicentric, multinational, retrospective, cohort study

Background: Tralokinumab is a human monoclonal antibody targeting interleukin-13 that is approved for the treatment of moderate-severe atopic dermatitis. Studies analyzing the efficacy and safety of tralokinumab in a real-world setting are scarce. Research design and methods: A European, multicentric, real-world, retrospective cohort study was defined to assess the effectiveness and safeness profile of tralokinumab, investigating the achievement of pre-specified treatment goals; and to detect potential differences in terms of effectiveness and safeness across some selected patient subcohorts. Results: A total of 194 adult patients were included in this study. A significant improvement in physician-assessed disease severity was detected at each follow-up visit as compared with baseline and similar trend was observed for patient-reported outcomes and quality of life. No meaningful difference in effectiveness was found when considering patient age (<65 versus ≥65 years), neither dissecting patient cohort in dupilumab-naive vs dupilumab-treated subjects. Among tralokinumabtreated patients, 88% achieved at least one currently identified real-world therapeutic goal at week 16. Conclusions: This retrospective multicenter study confirmed the effectiveness and safeness of tralokinumab throughout 32 weeks of observation, showing the achievement of therapeutic goals identified in both trial and real-world settings in a large proportion of tralokinumab-treated patients

A Multi-Actor Literature Review on Alternative and Sustainable Food Systems for the Promotion of Cereal Biodiversity

Organic and low-input food systems are emerging worldwide in answer to the sustainability crisis of the conventional agri-food sector. “Alternative” systems are based on local, decentralized approaches to production and processing, regarding quality and health, and short supply-chains for products with strong local identities. Diversity is deeply embedded in these food systems, from the agrobiodiversity grown in farmers’ fields, which improves resilience and adaptation, to diverse approaches, contexts and actors in food manufacturing and marketing. Diversity thus becomes a cross-sectoral issue which acknowledges consumers’ demand for healthy products. In the framework of the European project “CERERE, CEreal REnaissance in Rural Europe: embedding diversity in organic and low-input food systems”, the paper aims at reviewing recent research on alternative and sustainable food systems by adopting an innovative and participatory multi-actor approach; this has involved ten practitioners and twenty-two researchers from across Europe and a variety of technical backgrounds in the paper and analysis stages. The participatory approach is the main innovation and distinctive feature of this literature review. Partners selected indeed what they perceived as most relevant in order to facilitate a transition towards more sustainable and diversity based cereal systems and food chains. This includes issues related to alternative food networks, formal and informal institutional settings, grass root initiatives, consumer involvement and, finally, knowledge exchange and sustainability. The review provides an overview of recent research that is relevant to CERERE partners as well as to anyone interested in alternative and sustainable food systems. The main objective of this paper was indeed to present a narrative of studies, which can form the foundation for future applied research to promote alternative methods of cereal production in Europe.Peer reviewe