Evaluation of specific chicken IgY antibody value developing diagnostic capture antibody ELISA kit against Foot and Mouth disease.

The most preferred method for the detection of foot-and-mouth disease (FMD) viral antigen and identification of viral serotype is the enzyme-linked immunosorbent assay (ELISA). Diagnostic tests with high sensitivity are necessary both to distinguish infected vaccinated animals and execute disease control programs for the identification of the carrier animals. The current strategies for the detection of FMD virus are mainly based on the capture antibody (sandwich) ELISA test. The usage of laying pullets as an animal bioreactor for the production of specific egg yolk antibodies (IgY) has increased in recent years due to its high yield, affinity, low price, and quick production turnover. The present study aimed to produce a concentrated and purified IgY polyclonal antibody to design a capture antibody ELISA kit against the FMD virus (FMDV) serotype A. At first, laying hens were immunized with inactivated FMDV serotype virus, and then, on days 14, 21, and 28 following vaccination, the eggs and sera were collected. Afterward, the IgY polyclonal antibodies were extracted and purified from the chicken egg yolk using a polyethylene glycol 6000-ethanol precipitation procedure. Extracts were filtered, purified by ion exchange chromatography, and dialyzed. The purified IgY concentration, estimated by Bradford assay, confirmed its presence by SDS-PAGE and Western blot and also its specific immune reaction by Ouchterlony double immunodiffusion and Dot blot tests. Moreover, for achieving the optimum concentration of antigen/antibody (sera) in sandwich ELISA, a checkerboard titration test was set up based on indirect ELISA results. Eventually, 119 previously confirmed samples (including 80 positive and 39 negative) by both real-time polymerase chain reaction (quantitative PCR, qPCR) and a commercial ELISA kit were used for evaluation of the sensitivity and accuracy of our developed Capture antibody ELISA kit. In this manner, the sensitivity and specificity of our designed kit were 100% and 98%, respectively. Accordingly, the present developed capture ELISA kit based on IgY had high sensitivity and specificity for FMD virus detection and it could be used in the future for both commercial detecting and serotyping applications.

A comparison of conventional sodium fluoride varnish and nano-sodium fluoride varnish regarding enamel microhardness of deciduous teeth: an in-vitro study.

INTRODUCTION: Dental caries is the most common chronic disease worldwide, and various forms of fluoride are considered useful preventive tools. The production of nanoscale materials can significantly improve their mechanical and chemical properties. The present study compared the microhardness of primary tooth enamel after applying sodium fluoride varnish (conventional) and nano-sodium fluoride varnish. MATERIALS AND METHODS: Sixty-eight sound canine teeth were selected in this experimental study. The teeth were mounted so that the buccal surface was exposed. A 3 × 3 mm square was obtained on the buccal surface of the crown of each tooth. Enamel surfaces were polished using sandpaper in the presence of water as a coolant. The samples were randomly divided into four groups (n = 17): G1, conventional 5% NaF; G2, 1% nano-NaF; G3, 5% nano-NaF; G4, control. The initial microhardness was measured. Before surface treatment with different fluoride compounds, the samples were placed in a demineralizing solution for two days, and the microhardness of all the samples was re-measured. Then G1, G2, and G3 were treated with the fluoride type specified for each group, and G4 was treated as a control (without treatment). Finally, pH cycling was applied, and the microhardness was measured again. Data were analyzed with SPSS 20, using Repeated measure ANOVA and post-hoc Tukey tests. P < 0.05 was considered significant. RESULTS: Repeated measure ANOVA showed that microharness of G1, G2, G3, and G4 was statistically significant different. Tukey tests showed that the microhardness of G1, G2, and G3 were not significantly different. However, these three groups exhibited significantly more microhardness than the control group (P = 0.024, P = 0.027, and P = 0.010). CONCLUSION: There was no significant differences in enamel microhardness of deciduous teeth between conventional 5% NaF,1% nano-NaF and 5% nano-NaF.

Microscopic, Molecular and Antigen Detection and Isolation of Cryptosporidium parvumParasites in Diarrheal Disease of Calves in Iran.

Cryptosporidiosis is a parasitic disease caused by the protozoan Cryptosporidium in vertebrates. In livestock, especially ruminants, infants develop diarrheal syndromes. The infection is common worldwide , including Iran, where it is reported in several species. Morphological diagnosis of Cryptosporidium species is associated with many limitations and has no taxonomic value on its own, so the use of molecular methods can overcome these limitations to some extent. The present aims at microscopic, molecular and antigen detection and isolation of Cryptosporidium parvum parasites. Firstly, 300 samples were collected from different parts of Iran. Subsequently oocysts from feces were purified by the method of Casemore et al. using the flotation technique and stained by the modified Ziehl-Neelsen method (Henriksen method) and identified by diagnostic keys. ELISA test was also performed on the samples with results ranging from 1 to 4 positive. The results of our study show that, of the 300 cases tested for Cryptosporidium, 48 cases (16%) and 54 (18%) were positive in ELISA and PCR, respectively. Microscopic evaluation also mainly confirmed the ELISA results. These cases were collected in summer, autumn, and winter, with, more than 50% of the positive cases found among the samples collected in autumn. In addition, 54 positive cases were found by PCR test, which is 6 cases more than ELISA results. Finally, the results of PCR detection and ELISA were subjected to chi-square analysis, where no significant difference was found between the collected data (p=0.0587).

Diagnosis and treatment of malignant eyelid tumors.

BACKGROUND: Malignant tumors of the eyelid are much less frequent than benign eyelid alterations. These are frequently incidental findings without symptoms which are often overlooked or misinterpreted by patients. OBJECTIVE: This article gives an overview of clinical aspects, diagnostics and treatment of the five most common malignant eyelid tumors and exemplarily explains the essential principles of evidence-based treatment of malignant eyelid tumors. METHODS: This narrative review was prepared based on a selective literature search. The depiction of the treatment of eyelid tumors is supported by illustrations of clinical cases. RESULTS: The medical history and inspection provide initial indications of malignancy. Every eyelid change suspected of being malignant should be examined histologically to confirm a diagnosis. By far the most common malignant eyelid tumor in Europe is basal cell carcinoma, which metastasizes only in exceptional cases. Squamous cell carcinomas, sebaceous adenocarcinomas, melanomas and Merkel cell carcinomas occur much less frequently. In these cases, potential metastasis in particular must be considered when making the diagnosis and staging has to be initiated. Surgical excision into healthy tissue with tumor-free margins is the gold standard for malignant eyelid tumors. Non-surgical adjuvant or neoadjuvant forms of evidence-based treatment can be initiated based on the individual case to minimize the risk of recurrence and metastasis. CONCLUSION: It is essential to recognize eyelid changes at an early stage, to classify them correctly and to initiate the appropriate treatment. The interaction between the general condition and the personal needs of a patient as well as state of the art medicine are the keys to a good personalized treatment.

[Diagnosis and treatment of malignant eyelid tumors]. / Diagnose und Therapie der malignen Lidtumoren.

BACKGROUND: Malignant tumors of the eyelid are much less frequent than benign eyelid alterations. These are frequently incidental findings without symptoms which are often overlooked or misinterpreted by patients. OBJECTIVE: This article gives an overview of clinical aspects, diagnostics and treatment of the five most common malignant eyelid tumors and exemplarily explains the essential principles of evidence-based treatment of malignant eyelid tumors. METHODS: This narrative review was prepared based on a selective literature search. The depiction of the treatment of eyelid tumors is supported by illustrations of clinical cases. RESULTS: The medical history and inspection provide initial indications of malignancy. Every eyelid change suspected of being malignant should be examined histologically to confirm a diagnosis. By far the most common malignant eyelid tumor in Europe is basal cell carcinoma, which metastasizes only in exceptional cases. Squamous cell carcinomas, sebaceous adenocarcinomas, melanomas and Merkel cell carcinomas occur much less frequently. In these cases, potential metastasis in particular must be considered when making the diagnosis and staging has to be initiated. Surgical excision into healthy tissue with tumor-free margins is the gold standard for malignant eyelid tumors. Non-surgical adjuvant or neoadjuvant forms of evidence-based treatment can be initiated based on the individual case to minimize the risk of recurrence and metastasis. CONCLUSION: It is essential to recognize eyelid changes at an early stage, to classify them correctly and to initiate the appropriate treatment. The interaction between the general condition and the personal needs of a patient as well as state of the art medicine are the keys to a good personalized treatment.

[Clinical procedures for intraocular lymphomas]. / Klinisches Vorgehen bei intraokulären Lymphomen.

BACKGROUND: The classification of intraocular lymphomas is based on their anatomical location. They are divided into uveal lymphomas with involvement of the choroid, ciliary body or iris and vitreoretinal lymphomas with isolated or combined involvement of the vitreous body and/or retina. Over the last decades it has become increasingly possible to work out the clinical and pathobiological features of the various subtypes, thereby reducing the diagnostic hurdles and creating improved treatment options. OBJECTIVE: A summary of the various types of intraocular lymphoma in terms of clinical features, diagnostics, treatment and prognosis is given as well as recommendations for follow-up care. METHODS: A selective literature search was carried out on the subject of intraocular lymphomas using PubMed and Google Scholar. RESULTS: Intraocular lymphomas affect different structures, so that the symptoms can also be very different. The diagnostic spectrum ranges from typical ocular examination methods to sample biopsies with subsequent cytological, histological and molecular pathological processing. The treatment pillars available are percutaneous irradiation and intravitreal drug administration as local treatment and systemic treatment or a combination of systemic and local treatment. The prognosis depends mainly on the subtype of the lymphoma and the extent of the infestation when the diagnosis is confirmed. Even though some effective treatment options are now available, it has not yet been possible to significantly reduce the mortality rate. CONCLUSION: Many different options are available for the diagnostics and treatment of intraocular lymphomas, which require close interdisciplinary cooperation. The further developments in the field of molecular pathology allow a faster and more accurate diagnosis and could open up new treatment options in the future.

Plasmonic MoO3-x nanosheets by anodic oxidation of molybdenum for colorimetric sensing of hydrogen peroxide.

Hydrogen peroxide sensing is crucial for various medical diagnostics and industrial monitoring. On the other hand, doped metal oxides have recently emerged as cost-effective materials with localized surface plasmon resonance (LSPR) for colorimetric sensing of hydrogen peroxide. In this paper, using a simple anodic oxidation method, plasmonic MoO3-x colloidal nanosheets with deep blue color were fabricated and examined for the colorimetric sensing of hydrogen peroxide. X-ray photoelectron spectroscopy (XPS) revealed the presence of a considerable level of oxygen vacancy in the nanosheets composition. Depending on its concentration, hydrogen peroxide weakens the LSPR and the blue color of colloids with a sigmoidal sensing behavior. The impact of anodizing potential (10, 20, and 30 V) and time on a sensing performance was investigated and a limit of detection (LOD) as low as 0.2-0.9 µM was obtained. Furthermore, it was found that the LSPR undergoes redshift and the optical bandgap increases in a sigmoidal manner with analyte concentration that was explained by the existing theory on plasmonic semiconductors. To make a colorimetric assay, we immobilized MoO3-x nanosheets on felt fibers, which was observed by scanning electron microscope (SEM) images. The assay was examined to detect hydrogen peroxide by the naked eye in the concentration range of 800 µm to 100 mM and was analyzed using digital image analysis. Overall, our study develops a facile approach to produce MoO3-x nanosheets to detect hydrogen peroxide at the human-positive diabetes level (2.8-5.6 mM).

Validation of a CT-based motion model with in-situ fluoroscopy for lung surface deformation estimation.

Many surrogate-based motion models (SMMs), proposed to guide motion management in radiotherapy, are constructed by correlating motion of an external surrogate and internal anatomy during CT-simulation. Changes in this correlation define model break down. We validate a methodology that incorporates fluoroscopic (FL) images acquired during treatment for SMM construction and update. Under a prospective IRB, 4DCT scans, VisionRT (VRT) surfaces, and orthogonal FLs were collected from five lung cancer patients. VRT surfaces and two FL time-series were acquired pre- and post-treatment. A simulated annealing optimization scheme was used to estimate optimal lung deformations by maximizing the mutual information (MI) between digitally reconstructed radiographs (DRRs) of the SMM-estimated 3D images and FLs. Our SMM used partial-least-regression and was trained using the optimal deformations and VRT surfaces from the first breathing-cycle. SMM performance was evaluated using the MI score between reference FLs and the corresponding SMM or phase-assigned 4DCT DRRs. The Hausdorff distance for contoured landmarks was used to evaluate target position estimation error. For four out of five patients, two principal components approximated lung surface deformations with submillimeter accuracy. Analysis of the MI score between more than 4000 pairs of FL and DRR demonstrated that our model led to more similarity between the FL and DRR images compared to 4DCT and DRR images from a model based on an a priori correlation model. Our SMM consistently displayed lower mean and 95th percentile Hausdorff distances. For one patient, 95th percentile Hausdorff distance was reduced by 11 mm. Patient-averaged reductions in mean and 95th percentile Hausdorff distances were 3.6 mm and 7 mm for right-lung, and 3.1 mm and 4 mm for left-lung targets. FL data were used to evaluate model performance and investigate the feasibility of model update. Despite variability in breathing, use of post-treatment FL preserved model fidelity and consistently outperformed 4DCT for position estimation.

Certain haplotypes of the 3'-UTR region of the HLA-G gene are linked to breast cancer.

Background: Human leukocyte antigen G belongs to the family of non-classical HLA class I genes, its expression considered an important immune escape mechanism of cancer cells. The polymorphisms in the 3'-untranslated region (UTR) region of HLA-G influence the magnitude of the protein by modulating HLA-G mRNA stability. We hypothesised links between any of eight (UTR) single nucleotide polymorphisms (SNPs) and their haplotype of the HLA-G gene with breast cancer. Materials and Methods: Peripheral blood DNA from 100 patients affected by breast cancer and 100 controls was PCR sequenced for genotyping of 25 HLA-G 3'-UTR regions, including rs371194629 (+2960), rs1707 (+3003), rs1710 (+3010), rs17179101 (+3027), rs1063320 (+3142), rs9380142 (+3187), rs1610696 (+3196), and rs1233331 (+3227). Results: The 14-bp deletion (p = 0.01), and the +3010 (p = 0.021), +3142 (p = 0.006) and +3187 (p = 0.046) variants were significantly more prevalent in patients than in controls. In combining these data, two haplotypes of all eight SNPs and deletion/insertion (UTR-1 and UTR-4) are associated with breast cancer. Conclusion: Certain variants in the 3-UTR, and their combination as a haplotype, of the HLA-G gene are linked to breast cancer.

Design and expression of polytopic construct of cathepsin-L1, SAP-2 and FhTP16.5 proteins of Fasciola hepatica.

The enzyme-linked immunosorbent assay (ELISA) technique can play an important role in the early detection of fascioliasis. However, they have some diagnostic limitations, including cross-reaction with other helminths. It seems that the combination of recombinant parasite proteins as antigen can reduce these problems. Hence, the present study was aimed to design and confirm the antigenic recombinant multi-epitope (rMEP) construct of three protein epitopes (linear and conformational B-cell epitopes) of the parasite using immunoinformatic tools. For this purpose, the tertiary structures of Fasciola hepatica cathepsin-L1, saposin-like protein 2 and 16.5-kDa tegument-associated protein were predicted using the I-TASSER server. Validation of the modelled structures was performed by Ramachandran plots. The antigenic epitopes of the proteins were achieved by analysing the features of the IEDB server. The synthesized gene was cloned into the pET-22b (+) expression vector and transformed into the Escherichia coli BL21. Sodium dodecyl sulfate polyacrylamide gel electrophoresis was used to verify and analyse the expression of the rMEP protein. Western blotting was utilized to confirm rMEP protein immunogenicity in two forms, one using an anti-His tag antibody and the other with human pooled sera samples (fascioliasis, non-fascioliasis and negative control sera). Our results demonstrated that the rMEP designed for the three proteins of F. hepatica was highly antigenic, and immune-detection techniques confirmed the antigen specificity. In conclusion, the presented antigenic multi-epitope may be very helpful to develop serodiagnostic kits such as indirect ELISA to evaluate the proper diagnosis of fascioliasis in humans and ruminants.

Evaluation of choroidal substructure perfusion in patients affected by systemic sclerosis: an optical coherence tomography angiography study.

Objective: Vasculopathy in systemic sclerosis (SSc) is characterized by the obliteration of arterioles and a reduced capillary density in various tissues. In SSc, atrophic alterations of the choroid have been suggested based on morphological data acquired by optical coherence tomography (OCT). In this study, we aimed to assess the choroid in eyes of patients with SSc from a microcirculatory, dynamic point of view by adding optical coherence tomography angiography (OCTA) to the diagnostic spectrum.Method: SSc patients were enrolled, and age- and gender-matched healthy subjects were used as controls. In addition to basic ophthalmological and rheumatological examinations, individuals underwent enhanced-depth imaging OCT and OCTA. Subfoveal thicknesses of the choroid as well as all three choroidal vascular sublayers were measured and submacular perfusion values were evaluated.Results: In total, 12 patients with SSc and 12 matched controls were included. The median age of participants was 64 years. Submacular perfusion was significantly lower in the choriocapillaris (Δ = 0.72%; p = 0.045), Sattler's layer (Δ = 2.87%; p = 0.001), and Haller's layer (Δ = 2.69%; p = 0.018) of SSc patients compared to controls. Subfoveal thicknesses of Sattler's layer (Δ = 15 µm; p = 0.026) and Haller's layer (Δ = 41 µm; p = 0.045) were also significantly smaller in the SSc group.Conclusion: Choroidal microcirculation is impaired in SSc, even in patients without ophthalmological symptoms. Choroidal OCT and OCTA may offer additional biomarkers for SSc activity.

Novel Applications of Immuno-bioinformatics in Vaccine and Bio-product Developments at Research Institutes.

There are many challenges in the field of public health sciences. Rational decisions are required in order to treat different diseases, gain knowledge and wealth regarding research, and produce biological or synthetic products. Various advances in the basic laboratory science, computer science, and the engineering of biological production processes can help solve the occurring problems. Bioinformatics is defined as a field of science combined of biology, mathematics, physics, chemistry, and computer sciences. Recently, bioinformatics has been extensively used in the designing of the epitope, vaccines, antibodies, adjuvants, diagnostic kits, and therapeutic purposes (e.g., proteins, peptides, or small molecules). Moreover, bioinformatics includes chemoinformatics that has been employed to produce various biological or chemical products to target and combat pathogens. Bioinformatics is involved in other areas of data analysis and prediction, such as structural biology, system biology, phylogeny, population genetics, and next-generation data sequencing. To the best of our knowledge, no published study coherently described the benefits of bioinformatics fields applied for medication development or diagnostic aims in bio-productive and pharmaceutical/vaccine companies. Therefore, in the current review, we attempted to present the available bioinformatics resources, practical experiences, and other findings in the mentioned field along with providing a harmonized and applied model(s). The key points presented in the current review may help to elevate production and reduce the costs for the development of novel vaccines, medicines, and antibodies. In addition, these methods can facilitate the identification of organisms and may guarantee the quality of biological products.

Phylogenetic Analysis of bor Gene in an Escherichia coli Strain 1378 (O78:K80) Isolated from an Avian Colibacillosis Case in Tehran, Iran.

Colibacillosis is known as a fatal bacterial disease resulting in a high level of commercial loss worldwide. This study amid to elucidate the sequence, genetic characteristics, and phylogeny of the bor gene in Escherichia coli (E. coli) strain c1378 (O78:K80) isolated from avian colibacillosis in Iran and develop a rapid and optimal polymerase chain reaction (PCR) molecular-based technique with specific primers to detect this gene in E. coli. A virulent avian E. coli (i.e., laboratory designation E. coli strain c1378) isolated from a chicken with systemic colibacillosis from a broiler farm in Tehran, Iran, in 2004 was used as a source of the bor gene. After DNA extraction, PCR method was used to amplify the bor gene. A 658 bp fragment of the bor gene was amplified, sequenced, blasted, and phylogenetically studied. The most similar sequences to the bor gene in E. coli strain c1378 were E. coli APEC O78, Enterobacteria phage HK630, and Escherichia coli BW2952, respectively. There was a high similarity between the bor gene in E. coli bacteria with their phage and plasmid. Moreover, a high similarity was observed between the bor and iss genes (approximately 92%) showing that they were homologous genes. In addition, the similarity analysis of different bacterial species, as well as their plasmid and bacteriophage, to the bor gene indicated that the highest similarity to O78:K80 was related to Paracoccidioides brasiliensis, Bacillus thuringiensis CT43 plasmid pBMB0558, and Salmonella enterica subsp. enterica serovar Kentucky strain CVM29188 plasmid, respectively. Altogether, the results of the present study confirmed the presence of the bor gene in the studied isolates and clarified its sequence, phylogenetic relationship, and similarities of E. coli strain c1378 (O78:K80) isolated from avian colibacillosis.

Development and prospective in-patient proof-of-concept validation of a surface photogrammetry + CT-based volumetric motion model for lung radiotherapy.

PURPOSE: We develop and validate a motion model that uses real-time surface photogrammetry acquired concurrently with four-dimensional computed tomography (4DCT) to estimate respiration-induced changes within the entire irradiated volume, over arbitrarily many respiratory cycles. METHODS: A research, couch-mounted, VisionRT (VRT) system was used to acquire optical surface data (15 Hz, ROI = 15 × 20 cm2 ) from the thoraco-abdominal surface of a consented lung SBRT patient, concurrently with their standard-of-care 4DCT. The end-exhalation phase from the 4DCT was regarded as reference and for each remaining phase, deformation vector fields (DVFs) with respect to the reference phase were computed. To reduce dimensionality, the first two principal components (PCs) of the matrix of nine DVFs were calculated. In parallel, ten phase-averaged VRT surfaces were created. Surface DVFs and corresponding PCs were computed. A principal least squares regression was used to relate the PCs of surface DVF to those of volume DVFs, establishing a relationship between time-varying surface and the underlying time-varying volume. Proof-of-concept validation was performed during each treatment fraction by concurrently acquiring 30 s time series of real-time surface data and "ground truth" kV fluoroscopic data (FL). A ray-tracing algorithm was used to create a digitally reconstructed fluorograph (DRF), and motion trajectories of high-contrast, soft-tissue, anatomical features in the DRF were compared with those from kV FL. RESULTS: For five of the six fluoroscopic acquisition sessions, the model out-performed 4DCT in predicting contour Dice coefficient with respect to fluoroscopy-derived contours. Similarly, the model exhibited a marked improvement over 4DCT for patch positions on the diaphragm. Model patch position errors varied from 5 to -15 mm while 4DCT errors ranged between 5 and -22.4 mm. For one fluoroscopic acquisition, a marked change in the a priori internal-external correlation resulted in model errors comparable to those of 4DCT. CONCLUSIONS: We described the development and a proof-of-concept validation for a volumetric motion model that uses surface photogrammetry to correlate the time-varying thoraco-abdominal surface to the time-varying internal thoraco-abdominal volume. These early results indicate that the proposed approach can result in a marked improvement over 4DCT. While limited by the duration of the fluoroscopic acquisitions as well as the resolution of the acquired images, the DRF-based proof-of-concept technique developed here is model-agnostic, and therefore, has the potential to be used as an in-patient validation tool for other volumetric motion models.

Designing a polytopic complex vaccine candidate against Gallibacterium anatis: an In-silico study.

The haemolytic biovar of Gallibacterium anatis (G. anatis) is responsible for urogenital, gastrointestinal, and respiratory diseases in chickens. There are numerous reports on the resistance of G. anatis to antibiotics and recurrence of the disease, which raise concerns about antimicrobial treatment efficiency. Vaccination has been considered as the most feasible procedure of prevention in high risk farms. Subunit vaccines containing immunogenic components can have practical protective value in preventive measures regarding the infection. The present study aimed to introduce a polytopic vaccine candidate based on epitope detection. All registered sequences of four immunogenic proteins, includig Flfa, GTxA, Gab_1309, and Gab_2348 were retrieved and directed for variational analysis. A vaccine isolate was selected for each protein and tested for B-cell epitope mapping using different tools. Furthermore, consensus selected immunogenic regions with special patterns fused together by flexible linkers were integrated into two constructs and checked for the best status of proteasomal cleavage sites, as well as hydropathy plot. Moreover, back translations, along with codon optimization were performed, and then some tags were added to the constructs. The selected consensus B-cell immunogenic epitopes were for 12656: AA114-181, 7990: AA114-181, Avicor: AA42-77, 134-197, and IPDH: 61-155 for Flfa protein, AA185-235, AA372-457, and AA807-941 for GtxA-N, AA260-305, AA340-400, and AA110-146 for Gab-1309, and AA125-AA175 for Gab-2348. Two suitable patterns of attachment were selected from the different fusion patterns of epitopes in B-cell polytopic vaccinal constructs. Finally, the examination of these constructs showed their effect and efficacy for immune system stimulation. Based on bioinformatics results, these immunogens could be utilized as potential candidates to develop polytopic protective vaccines and design diagnostic kits.

[RPE morphometry in geographic atrophy due to AMD]. / RPE-Morphometrie bei AMD-bedingter geografischer Atrophie.

BACKGROUND: The geographic atrophy (GA) junctional zone includes changes in retinal pigment epithelium (RPE) and Bruch's membrane complex that are still difficult to interpret even with clinical high-resolution imaging technologies. We measured and evaluated degenerative RPE cell changes in histological sections of GA eyes. METHODS: In this study seven GA eyes were evaluated by light microscopy. In three eye sections, zones of typical RPE alterations were graded (0 = normal; 1 = irregular cells but intact layer; 2 = rounded, enlarged, and/or heaped cells; 3 = migrating cells in retina; 4 = RPE absent). In each graded zone, we measured and analyzed A) the total height of the RPE cell layer, B) the height of individual RPE cells and C) the height of basal deposits. RESULTS: From the outer macula towards the central RPE atrophic area the RPE passed almost steadily upward through grades of increased pathology. A) Zone 2 exhibited highly variable total RPE height (16.9 ± 5.6 µm) with hypertrophic and heaped cells next to atrophic ones. In comparison, zone 0 and zone 1 showed less variability and a regular total RPE height (10.9 ± 2.6 µm). B) In zone 2 the size of altered RPE cells varied widely (12.4 ± 5.2 µm, min. 5.1, max. 27.5). All detected migrating RPE cells were hypertrophic. C) In zone 0 basal deposits were found sporadically. With progressing RPE alterations, basal deposits became progressively continuous and thicker and reached a considerable height at the atrophic zone (9.5 ± 4.3 µm). CONCLUSION: Our measurements confirmed that degenerative RPE phenomena, particularly of degeneration grade 2/3 close to the actual RPE atrophy zone, are often large enough to be visualized in detail with already available modern imaging technologies (e. g. SD-OCT).

Sequencing and In Silico Multi-aspect Analysis of S1 Glycoprotein in 793/B Serotype of Infectious Bronchitis Virus Isolated From Iran in 2003 and 2011.

Infectious bronchitis (IB) is an acute, highly contagious, and economically important viral disease of chickens. The S1 subunit from Spike (S) protein plays the major role in protective immunity and is involved in the host-virus interactions, as well as infectious bronchitis virus (IBV) serotyping. Aim of the present study was multi-aspect analysis of the molecular and immunological features of 5&#39; part belonging to the S1 glycoprotein sequence of Iranian 793/B IBV strain isolates. This might ideally help in characterization, prevention, and vaccine development. The tissue samples were prepared, followed by virus isolation, reverse transcription polymerase chain reaction and restriction fragment length polymorphism analysis. In addition, sequencing and registration of the sequences in the National Center for Biotechnology Information were performed. Moreover, 12 sequences were retrieved from Fars province, Iran. The next steps included evaluation of conservation/variability along the sequences, phylogenetic analysis, estimation of the average evolutionary divergence over all the sequence pairs, predicting the phosphorylation/N-glycosylation/palmitoylation sites, and the final analysis of antigenicity. The findings of alignment, entropy plot, and pairwise similarity analysis revealed 17 hypervariable regions. The isolates belonging to Tehran were clustered in phylogenetic tree, and the most similar isolates to them were ADW11182 and ADW11183. Location of some of the N-glycosylation/phosphorylation/palmitoylation points indicated that these sites were conserved among the isolates. Furthermore, the frequency of epitopes and their scores reflect the high immunogenicity of S1 protein in 793/B serotype. Analysis of the primary and secondary structures demonstrated that their parameters had variable values and were different regarding the number and location of &alpha;-helix, &beta;-strand, and coils. According to our findings, the Iranian isolates of 793/B serotype change their molecular characteristics during time and in different geographical regions. These alterations might account for failure in prevention programs and differences in virulence and pathogenicity.

Production of a Human Recombinant Polyclonal Fab Antivenom against Iranian Viper Echis carinatus.

Venomous snakebite is a life-threatening injury in many tropical and subtropical areas including Iran. The gold standard treatment option for human envenomation is the use of antivenoms. Despite the unique effects of horse-derived antivenoms on the treatment of snakebite, they are not fully perfect and need improvements. In this study, human recombinant Fab fragment antivenom was produced in Rosetta-g bacterium using a gene library constructed in the previous study. The prepared Fab was purified in several steps, desalted, and lipopolysaccharide-depleted using ammonium sulfate solution and dialysis against phosphate buffer and Triton X-114 solution, respectively. Subsequently, the product was initially confirmed by the sodium dodecyl sulfate polyacrylamide gel electrophoresis and enzyme-linked immunosorbent assay (ELISA), respectively. Finally, the neutralization potency of the product was investigated in laboratory Syrian Mice. The obtained results showed corresponding reduced bands to Fab fragment with the molecular weight of about 28 kDa at a concentration of 3.1 mg/ml. There was a significant difference between the groups in terms of ELISA test (P&lt;0.05). The neutralization potency of the product against the venom of Echis carinatus (E. carinatus) was about 7 LD50/ml (54.6 &micro;g/ml) when tested on mice. Based on the results, the Fab fragment antivenom had the ability to neutralize the in vivo biological activity of the venom of Iranian E. carinatus. However, further studies are recommended to reach a suitable concentration of antivenom fragment.