MIP-based protein profiling: A method for interspecies discrimination

Due to recent public concern and interest in the authenticity and origin of meat, for example, the 2013 “horsemeat scandal” in the human food chain, novel sensor strategies for the discrimination between protein species are highly sought after. In this work, molecularly imprinted polymers (MIPs) are utilised for protein discrimination using electrochemical sensor and spectrophotometric techniques. MIP selectivity between two proteins of similar molecular weight (haemoglobin and serum albumin) were compared across three different species, namely pork, beef and human. Bulk MIPs resulted in Kd and Bmax values of 184±23 µM, and 582 µmol g-1 for BHb, 246.3±26 µM, and 673 µmol g-1 for HHb; 276±31 µM, and 467 µmol g-1 for PHb. With the aid of chemometrics, i.e. multivariate analysis and pattern recognition, distinctive protein profiles have been achieved for species discrimination in both spectrophotometric and electrochemical analysis experiments. MIP suitability and selectivity within complex matrices was also assessed using urine, human plasma and human serum. Pattern recognition MIP-based protein profiling demonstrated positive outputs yielding either a ‘bovine’ or ‘not-bovine’ outcome (p = 0.0005) for biological samples spiked with/without bovine using respective bovine haemoglobin MIPs

Genetic Variation in the Familial Mediterranean Fever Gene (MEFV) and Risk for Crohn's Disease and Ulcerative Colitis

BACKGROUND AND AIMS: The familial Mediterranean fever (FMF) gene (MEFV) encodes pyrin, a major regulator of the inflammasome platform controlling caspase-1 activation and IL-1beta processing. Pyrin has been shown to interact with the gene product of NLRP3, NALP3/cryopyrin, also an important active member of the inflammasome. The NLRP3 region was recently reported to be associated with Crohn's disease (CD) susceptibility. We therefore sought to evaluate MEFV as an inflammatory bowel disease (IBD) susceptibility gene. METHODOLOGY AND RESULTS: MEFV colonic mucosal gene expression was significantly increased in experimental colitis mice models (TNBS p<0.0003; DSS p<0.006), in biopsies from CD (p<0.02) and severe ulcerative colitis (UC) patients (p<0.008). Comprehensive genetic screening of the MEFV region in the Belgian exploratory sample set (440 CD trios, 137 UC trios, 239 CD cases, 96 UC cases, and 107 healthy controls) identified SNPs located in the MEFV 5' haplotype block that were significantly associated with UC (rs224217; p = 0.003; A allele frequency: 56% cases, 45% controls), while no CD associations were observed. Sequencing and subsequent genotyping of variants located in this associated haplotype block identified three synonymous variants (D102D/rs224225, G138G/rs224224, A165A/rs224223) and one non-synonymous variant (R202Q/rs224222) located in MEFV exon 2 that were significantly associated with UC (rs224222: p = 0.0005; A allele frequency: 32% in cases, 23% in controls). No consistent associations were observed in additional Canadian (256 CD trios, 91 UC trios) and Scottish (495 UC, 370 controls) sample sets. We note that rs224222 showed marginal association (p = 0.012; G allele frequency: 82% in cases, 70% in controls) in the Canadian sample, but with a different risk allele. None of the NLRP3 common variants were associated with UC in the Belgian-Canadian UC samples and no significant interactions were observed between NLRP3 and MEFV that could explain the observed flip-flop of the rs224222 risk allele. CONCLUSION: The differences in association levels observed between the sample sets may be a consequence of distinct founder effects or of the relative small sample size of the cohorts evaluated in this study. However, the results suggest that common variants in the MEFV region do not contribute to CD and UC susceptibility.Journal ArticleResearch Support, N.I.H. ExtramuralResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Evaluation of epidermal growth factor receptor mutation status in serum DNA as a predictor of response to gefitinib (IRESSA)

The aim of this study was to evaluate the usefulness of EGFR mutation status in serum DNA as a means of predicting a benefit from gefitinib (IRESSA) therapy in Japanese patients with non-small cell lung cancer (NSCLC). We obtained pairs of tumour and serum samples from 42 patients treated with gefitinib. EGFR mutation status was determined by a direct sequencing method and by Scorpion Amplification Refractory Mutation System (ARMS) technology. EGFR mutations were detected in the tumour samples of eight patients and in the serum samples of seven patients. EGFR mutation status in the tumours and serum samples was consistent in 39 (92.9%) of the 42 pairs. EGFR mutations were strong correlations between both EGFR mutation status in the tumour samples and serum samples and objective response to gefitinib (P<0.001). Median progression-free survival time was significantly longer in the patients with EGFR mutations than in the patients without EGFR mutations (194 vs 55 days, P=0.016, in tumour samples; 174 vs 58 days, P=0.044, in serum samples). The results suggest that it is feasible to use serum DNA to detect EGFR mutation, and that it's potential as a predictor of response to, and survival on gefitinib is worthy of further evaluation

Nucleic acid-based fluorescent probes and their analytical potential

It is well known that nucleic acids play an essential role in living organisms because they store and transmit genetic information and use that information to direct the synthesis of proteins. However, less is known about the ability of nucleic acids to bind specific ligands and the application of oligonucleotides as molecular probes or biosensors. Oligonucleotide probes are single-stranded nucleic acid fragments that can be tailored to have high specificity and affinity for different targets including nucleic acids, proteins, small molecules, and ions. One can divide oligonucleotide-based probes into two main categories: hybridization probes that are based on the formation of complementary base-pairs, and aptamer probes that exploit selective recognition of nonnucleic acid analytes and may be compared with immunosensors. Design and construction of hybridization and aptamer probes are similar. Typically, oligonucleotide (DNA, RNA) with predefined base sequence and length is modified by covalent attachment of reporter groups (one or more fluorophores in fluorescence-based probes). The fluorescent labels act as transducers that transform biorecognition (hybridization, ligand binding) into a fluorescence signal. Fluorescent labels have several advantages, for example high sensitivity and multiple transduction approaches (fluorescence quenching or enhancement, fluorescence anisotropy, fluorescence lifetime, fluorescence resonance energy transfer (FRET), and excimer-monomer light switching). These multiple signaling options combined with the design flexibility of the recognition element (DNA, RNA, PNA, LNA) and various labeling strategies contribute to development of numerous selective and sensitive bioassays. This review covers fundamentals of the design and engineering of oligonucleotide probes, describes typical construction approaches, and discusses examples of probes used both in hybridization studies and in aptamer-based assays

Peasants' Choices? Indian Agriculture and the Limits of Commercialization in Nineteenth-Century Bihar

The article attempts to distinguish and locate choices in agricultural production, with special reference to Bihar, India, during the nineteenth century. On the one hand, it considers closely managed and extensively irrigated areas, long involved in trade under the overall control of 'landlords', and, on the other hand, the expanding production of opium, and also of indigo and sugar (so-called 'forced' commercialization), identifying common features and continuities of production and marketing. Particular the importance of advance payments and local intermediaries is stressed. Thus, in contrast with the more usual evolutionary models, based on unitary categories and modes, the essay illustrates ecological, customary, collective, and local political constraints upon agricultural decisions; and this leads to the identification in turn of their different kinds and levels