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COVID-19 first appeared in December of 2019 in Wuhan, China. Since then it has become a global pandemic. A robust and scalable diagnostics strategy is crucial for containing and monitoring the pandemic. RT-PCR is a known, reliable method for COVID-19 diagnostics which can differentiate between SARS-CoV-2 and other viruses. However, PCR is location dependent, time consuming and relatively expensive. Thus, there is a need for a more flexible method which may be produced in an off-the-shelf format and distributed more widely. Paper-based immunoassays can fulfill this function. Here we present the first steps towards a paper-based test which can differentiate between different between the Spike protein of various coronaviruses, SARS-CoV-1, SARS-CoV-2 and CoV-HKU1 with negligible cross reactivity for HCoV-OC43 and HCoV-229E in a single assay which takes less than 30 minutes. Furthermore, our test can distinguish between fractions of the same Spike protein. This is done by an altered assay design with four test line locations where each antigen builds a unique, identifiable binding pattern. The effect of several factors, such as running media, immunoprobe concentration and antigen interference is considered. We find that running media has a significant effect on the final binding pattern where human saliva provides results while human serum leads to the lowest signal quality.

Lateral flow type detection is becoming interesting not only in regions with a poor medical infrastructure but also for practitioners in day-to-day clinical work or for veterinary control in case of possible epidemics. In this work, we describe the first steps of development of a multi-channel strip with potential internal calibration of multiparametric and colorimetric lateral flow assays for the simultaneous detection of the lipopolysaccharides (LPS) of Salmonella typhimurium (S. typhimurium) and Salmonella enteritidis (S. enteritidis). We structured four channels in the nitrocellulose membrane with a Yb:KGW solid-state femtosecond laser ("cold" ablation process) to form distinct tracks of porous material and used gold nanoparticles for the labeling of the antibodies. In addition, calibration curves of the spot intensities of both serovars are presented, and it was shown that no cross reactivity between the different capture antibodies and LPS occurred. Finally, we detected LPS of both Salmonella serovars simultaneously. The color changes (spot intensities of the reaction zones) were evaluated using the open-source image-processing program ImageJ. Graphical abstract Multiparametric testing, strip A was tested with LPS S. enteritidis ( c=0.01 g/L) and LPS S.typhimurium ( c=0.0001 g/L), strip B with LPS S. enteritidis ( c=0.001 g/L) and LPS S. typhimurium ( c=0.001g/L) and strip C with LPS S. enteritidis (c=0.0001 g/L) and LPS S. typhimurium ( c=0.01 g/L), and read-out.

Highly multiplexed immunoassays could allow convenient screening of hundreds or thousands of protein biomarkers simultaneously in a clinical sample such as serum or plasma, potentially allowing improved diagnostic accuracy and clinical management of many conditions such as autoimmune disorders, infections, and several cancers. Currently, antibody microarray-based tests are limited in part due to cross reactivity from detection antibody reagents. Here we present a strategy that reduces the cross-reactivity between nanoparticle-bound reporter antibodies through the application of ultrasound energy. By this concept, it was possible to achieve a sensitivity 10(3)-fold (5 pg mL(-1)) lower than when no ultrasound was applied (50 ng mL(-1)) for the simultaneous detection of three different antigens. The detection limits and variability achieved with this technique rival those obtained with other types of multiplex sandwich assays.

Inoculation with vaccine is the major intervention currently used to prevent influenza infections. However, it will be a challenge to produce and implement a new vaccine when a novel highly pathogenic influenza virus emerges in humans as significant infections. H7 subtype influenza viruses have similar epitopes on hemagglutinin, which can induce cross-reactive antibodies. In this study, a meta-analysis of the cross-reactivity of antibodies induced by one H7 subtype influenza vaccine against other H7 subtypes was performed. Database search was conducted in PubMed, Cochrane Library, EMBASE, MEDLINE, Chinese Biological Medicine Database (CBM), and Wanfang. A total of 9 articles comprising 811 human subjects were included in this meta-analysis. All assessed H7 influenza vaccines induced vaccine strain-specific protective antibodies [seroconversion rate (SCR) = 0.74, 95% CI (0.65, 0.82); seroprotection rate (SPR) = 0.81, 95% CI (0.78, 0.83)]. All H7 influenza virus monovalent vaccines exhibited cross-reactivity tested by hemagglutinin inhibition test (HI), microneutralization test (MN) and immunosorbent assay (ELISA) to other H7 subtype viruses. H7N1, H7N3, H7N7, and H7N9 vaccines elicited cross-reactive antibodies against other H7 subtype influenza viruses [SCR = 0.66, 95% CI (0.50, 0.82); SPR = 0.79, 95% CI (0.67, 0.91)]. The pooled SCR (95%CI) of cross-reactivity of H7N1 and H7N3 vaccines were 0.88 (0.85, 0.91) and 0.40 (0.26, 0.54), respectively. The consolidated SPR (95%CI) of H7N1 and H7N7 vaccines were 0.89 (0.86, 0.92) and 0.93 (0.81, 1.06). All H7 vaccines induced cross-reactive antibodies against H7N9 viruses [SCR = 0.69, 95% CI (0.52, 0.86); SPR = 0.85, 95% CI (0.76, 0.94)]. H7 vaccines can be used to limit influenza infection when a new highly pathogenic H7 virus appears.

Background Chronic hepatitis B virus (HBV) infections are a global health problem. There is a need for therapeutic strategies blocking continuous infection of liver cells. The grass pollen allergy vaccine BM32 containing the preS domain of the large HBV surface protein (LHBs) as immunogenic carrier induced IgG antibodies in human subjects inhibiting HBV infection in vitro. Aim of this study was the quantification, epitope mapping and investigation of HBV genotype cross-reactivity of preS-specific antibodies in subjects treated with different dosage regimens of BM32 Methods Hundred twenty eight grass pollen allergic patients received in a double-blind, placebo-controlled trial five monthly injections of placebo (aluminum hydroxide, n= 34) or different courses of BM32 (2 placebo + 3 BM32, n= 33; 1 placebo + 4 BM32, n= 30; 5 BM32, n= 31). Recombinant Escherichia coli-expressed preS was purified. Overlapping peptides spanning preS and the receptor-binding sites from consensus sequences of genotypes A–H were synthesized and purified. Isotype (IgM, IgG, IgA, IgE) and IgG subclass (IgG1-IgG4) responses to preS and peptides were determined by ELISA at baseline, one and four months after the last injection. IgG1 and IgG4 subclass concentrations specific for preS and the receptor-binding site were measured by quantitative ELISA. Findings Five monthly injections induced the highest levels of preS-specific IgG consisting mainly of IgG1 and IgG4, with a sum of median preS-specific IgG1 and IgG4 concentrations of >135 μg/ml reaching up to 1.8 mg/ml. More than 20% of preS-specific IgG was directed against the receptor-binding site. BM32-induced IgG cross-reacted with the receptor-binding domains from all eight HBV genotypes A-H. Interpretation BM32 induces high levels of IgG1 and IgG4 antibodies against the receptor binding sites of all eight HBV genotypes and hence might be suitable for therapeutic HBV vaccination. Funding This study was supported by the PhD program IAI (KPW01212FW), by Viravaxx AG and by the Danube-ARC funded by the Government of Lower Austria. Rudolf Valenta is a recipient of a Megagrant of the Government of the Russian Federation, grant No 14.W03.31.0024.

Bioinformatic amino acid sequence searches are used, in part, to assess the potential allergenic risk of newly expressed proteins in genetically engineered crops. Previous work has demonstrated that the searches required by government regulatory agencies falsely implicate many proteins from rarely allergenic crops as an allergenic risk. However, many proteins are found in crops at concentrations that may be insufficient to cause allergy. Here we used a recently developed set of high-abundance non-allergenic proteins to determine the false-positive rates for several algorithms required by regulatory bodies, and also for an alternative 1:1 FASTA approach previously found to be equally sensitive to the official sliding-window method, but far more selective. The current investigation confirms these earlier findings while addressing dietary exposure.

Biological drugs like therapeutic antibodies are widely used for the treatment of various diseases like inflammatory disorders and cancer. A drawback of these novel treatments is the substantial proportion of patients experiencing adverse reactions such as loss-of-drug effect or hypersensitivity reactions. These reactions are associated with pre-existing and/or developing anti-drug antibodies. Especially IgE development is a risk factor for life-threatening systemic anaphylaxis. Methods: In order to characterize the individual drug-specific serum IgE, an IgE cross-reactivity immune profiling (ICRIP) assay was developed. Individual IgG epitopes of anti-drug antibodies against adalimumab were identified by epitope mapping via peptide microarray. Results: ICRIP analyses of sera from patients treated with the therapeutic antibodies adalimumab (ADL) and infliximab (IFX) reveal individual, distinct IgE binding patterns. IgG epitopes were identified mostly located in the variable region of ADL. Conclusions: Using ICRIP and peptide microarrays for pharmacovigilance of the TNF-α blockers IFX and ADL, risk factors and biomarkers before and during therapy shall be identified. These diagnostic systems provide the basis for a safe and efficacious therapy decision for each patient in cases of adverse drug reactions mediated by different types of anti-drug antibodies.

Infection with one of the four dengue virus serotypes (DENV1-4) presumably leads to lifelong immunity against the infecting serotype but not against heterotypic reinfection, resulting in a greater risk of developing Dengue Hemorrhagic Fever/Dengue Shock Syndrome (DHF/DSS) during secondary infection. Both antibodies and T cell responses have been implicated in DHF/DSS pathogenesis. According to the T cell-based hypothesis termed “original antigenic sin,” secondary DENV infection is dominated by non-protective, cross-reactive T cells that elicit an aberrant immune response. The goal of our study was to compare the roles of serotype-specific and cross-reactive T cells in protection vs. pathogenesis during DENV infection in vivo. Specifically, we utilized IFN-α/βR−/− HLA*B0702 transgenic mice in the context of peptide vaccination with relevant human CD8 T cell epitopes. IFN-α/βR−/− HLA*B0702 transgenic mice were immunized with DENV serotype 2 (DENV2)-specific epitopes or variants found in any of the other three serotypes (DENV1, DENV3 or DENV4), followed by challenge with DENV. Although cross-reactive T cell responses were lower than responses elicited by serotype-specific T cells, immunization with either serotype-specific or variant peptide epitopes enhanced viral clearance, demonstrating that both serotype-specific and cross-reactive T cells can contribute to protection in vivo against DENV infection., Highlights • Serotype-cross-reactive CD8 T cells elicit a polyfunctional immune response similar to the response elicited by serotype-specific CD8 T cells. • Serotype cross-reactive CD8 T cells play a role in protection against DENV infection in a HLA-B*0702 Transgenic IFN-α/βR−/− mouse model. There are four major subtypes (serotypes) of the mosquito-borne Dengue virus. Infection with a first serotype is generally asymptomatic, but secondary infection with a different serotype is capable of causing severe disease. T cells previously exposed to a first serotype and which produce an immune response to a second serotype are said to be cross-reactive. Using a mouse model engineered with human T cell features, we characterized the cross-reactive T cell response to live dengue virus serotypes and viral protein fragments. Our results suggested cross-reactive T cells contribute to control of and protection against infection by a second dengue serotype, rather than leading to more severe disease.

Biomarkers may hold the key towards development and improvement of personalized cancer treatment. For instance, tumour expression of immune system-related proteins may reveal the tumour immune status and, accordingly, determine choice for type of immunotherapy. Therefore, objective evaluation of tumour biomarker expression is needed but often challenging. For instance, human leukocyte antigen (HLA) class I tumour epithelium expression is cumbersome to quantify by eye due to its presence on both tumour epithelial cells and tumour stromal cells, as well as tumour-infiltrating immune cells. In this study, we solved this problem by setting up an immunohistochemical (IHC) double staining using a tissue microarray (TMA) of rectal tumours wherein HLA class I expression was coloured with a blue chromogen, whereas non-epithelial tissue was visualized with a brown chromogen. We subsequently developed a semi-automated image analysis method that identified tumour epithelium as well as the percentage of HLA class I-positive tumour epithelium. Using this technique, we compared HCA2/HC10 and EMR8-5 antibodies for the assessment of HLA class I tumour expression and concluded that EMR8-5 is the superior antibody for this purpose. This IHC double staining can in principle be used for scoring of any biomarker expressed by tumour epithelium.

Autoimmunity is on the rise around the globe. Diet has been proposed as a risk factor for autoimmunity and shown to modulate the severity of several autoimmune disorders. Yet, the interaction between diet and autoimmunity in humans remains largely unstudied. Here, we systematically interrogated commonly consumed animals and plants for peptide epitopes previously implicated in human autoimmune disease. A total of fourteen species investigated could be divided into three broad categories regarding their content in human autoimmune epitopes, which we represented using a new metric, the Gershteyn-Ferreira index (GF index). Strikingly, pig contains a disproportionately high number of unique autoimmune epitopes compared to all other species analyzed. This work uncovers a potential new link between pork consumption and autoimmunity in humans and lays the foundation for future studies on the impact of diet on the pathogenesis and progression of autoimmune disorders., Highlights • We developed a comprehensive database of dietary-human autoimmune epitope overlap. • The Gershteyn-Ferreira index divides dietary antigens into three main groups with respect to overlap with human autoimmune epitopes. • Pig contains a disproportionately high number of unique human autoimmune epitopes.

To characterize the antigenicity of nucleocapsid proteins (NP) derived from canine coronavirus (CCoV) and canine respiratory coronavirus (CRCoV) in China, the N genes of CCoV (CCoV-BJ70) and CRCoV (CRCoV-BJ202) were cloned from swabs obtained from diseased pet dogs in Beijing and then sequenced. The recombinant NPs (rNPs) were expressed in Escherichia coli and purified by nickel-affinity column and size exclusion chromatography. Sequencing data indicated that the N genes of CCoV-BJ70 and CRCoV-BJ202 belonging to two distinctly different groups were relatively conserved within each subgroup. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results showed that rNPs of CCoV and CRCoV were expressed efficiently and isolated with a final purity of over 95%. Western blot analysis revealed the rNP from CRCoV could cross-react with mice antisera against human coronavirus (HCoV-229E, NL63, OC43, HKU1), while rNP of CCoV had cross-reactivity with only anti-sera against viruses belonging to the same group (HCoV-229E and NL63). In summary, CCoV and CRCoV rNPs were successfully expressed in E. coli and showed antigenic cross-reactivity with antisera raised against human coronaviruses. These findings indicate that further serologic studies on coronavirus infections at the animal-human interface are needed.

Anflu® is a seasonal trivalent inactivated split-virion influenza vaccine manufactured by Sinovac Biotech Co., Ltd. The objectives of this study were to evaluate the safety of Anflu® (2013–14 formulation: H1N1, H3N2 and BYAM) in infants and adults and its immunogenicity and cross-reactivity against mismatched influenza B lineage and avian influenza A(H7N9) viruses (hereafter BVIC and H7N9, respectively) in adults. In this phase IV open label trial, infants 6–35 months old (n=61) each received two injections with 28 days apart; adults 18–60 yrs old (n=60) and elderly >60 yrs old (n=61) each received one injection. Information of adverse events was collected through safety observation and follow-up visits. Pre- and post-immune blood samples (day 0 and 21) were collected from subjects ≥18 yrs old to detect hemagglutination inhibition antibody titers and calculate seroprotection rates (SPRs) and seroconversion rates (SCRs). The overall adverse reaction incidence was 1.6% (3/182), and no serious adverse event was reported during the study period. For subjects ≥18 yrs old, the SCRs, SPRs, and the geometric mean titers (GMTs) met the European criteria for all three strains. In addition, the point estimations of SCR, SPR and GMT for BVIC also met the European criteria. Six subjects were seroconverted against H7N9; however the serological results did not meet the European criteria. In conclusion, the results showed a satisfactory safety and immunogenicity profile of Anflu® and cross-reactivity against BVIC, but did not demonstrate cross-reactivity against H7N9 (Clinicaltrials.gov ID: NCT02269852).

Introduction Influenza A is known as a public health concern worldwide. In this study, a novel one-step real-time reverse transcription polymerase chain reaction (rtRT-PCR) assay was designed and optimized for the detection of influenza A viruses. Material and methods The primers and probe were designed based on the analysis of 90 matrix nucleotide sequence data of influenza type A subtypes from the GenBank database of the National Center for Biotechnology Information (NCBI). The influenza virus A/Tehran/5652/2010 (H1N1 pdm09) was used as a reference. The rtRT-PCR assay was optimized, compared with that of the World Health Organization (WHO), and its analytical sensitivity, specificity and reproducibility were evaluated. In total, 64 nasopharyngeal swabs from patients with influenza-like illness (ILI) and 41 samples without ILI symptoms were tested for the virus, using conventional cell culture, direct immunofluorescence antibody (DFA) methods, and one-step rtRT-PCR with the designed primer set and probe and the WHO’s. Results The optimized assay results were similar to the WHO’s. The optimized assay results were similar to WHO’s, with non-significant differences for 10–103 copies of viral RNA/reaction (p > 0.05). It detected 10 copies of viral RNA/reaction with high reproducibility and no cross reactivity with other respiratory viruses. A specific cytopathic effect was observed in 6/64 (9.37%) of the ILI group using conventional culture and DFA staining methods; however, it was not seen in non-ILI. Also, the results of our assay and the WHO’s were similar to those of viral isolation and DFA staining. Conclusions Given the high specificity, sensitivity and reproducibility of this novel assay, it can serve as a reliable diagnostic tool for the detection of influenza A viruses in clinical specimens and lab experiments.

Infections caused by Fasciola hepatica are of great importance in the veterinary field, as they cause important economic losses to livestock producers. Serodiagnostic methods, typically ELISA (with either native or recombinant antigens), are often used for early diagnosis. The use of native antigens, as in the MM3-SERO ELISA (commercialized as BIO K 211, BIO-X Diagnostics), continues to be beneficial in terms of sensitivity and specificity; however, there is interest in developing ELISA tests based on recombinant antigens to avoid the need to culture parasites. Of the antigens secreted by adult flukes, recombinant procathepsin L1 (rFhpCL1) is the most commonly tested in ELISA to date. However, although adult flukes produce three different clades of CLs (FhCL1, FhCL2, and FhCL5), to our knowledge, the diagnostic value of recombinant FhCL2 and FhCL5 has not yet been investigated. In the present study, we developed and tested three indirect ELISAs using rFhpCL1, rFhpCL2, and rFhpCL5 and evaluated their recognition by sera from sheep and cattle naturally infected with F. hepatica. Although the overall antibody response to these three rFhpCLs was similar, some animals displayed preferential recognition for particular rFhpCLs. Moreover, for cattle sera, the highest sensitivity was obtained using rFhpCL2 (97%), being equal for both rFhpCL1 and rFhpCL5 (87.9%), after adjusting cut-offs for maximum specificity. By contrast, for sheep sera, the sensitivity was 100% for the three rFhpCLs. Finally, the presence of truncated and/or partially unfolded molecules in antigen preparations is postulated as a possible source of cross-reactivity.

According to several recent studies, an unexpectedly high number of landmark papers seem to be not reproducible by independent laboratories. Nontherapeutic antibodies used for research, diagnostic, food analytical, environmental, and other purposes play a significant role in this matter. Although some papers have been published offering suggestions to improve the situation, they do not seem to be comprehensive enough to cover the full complexity of this issue. In addition, no obvious improvements could be noticed in the field as yet. This article tries to consolidate the remarkable variety of conclusions and suggested activities into a more coherent conception. It is concluded that funding agencies and journal publishers need to take first and immediate measures to resolve these problems and lead the way to a more sustainable way of bioanalytical research, on which all can rely with confidence.

The phenomenon of cross-reactivity between allergenic proteins plays an important role to understand how the immune system recognizes different antigen proteins. Allergen proteins are known to cross-react if their sequence comparison shows a high sequence identity which also implies that the proteins have a similar 3D fold. In such cases, linear sequence alignment methods are frequently used to predict cross-reactivity between allergenic proteins. However, the prediction of cross-reactivity between distantly related allergens continues to be a challenging task. To overcome this problem, we developed a new structure-based computational method, Cross-React, to predict cross-reactivity between allergenic proteins available in the Structural Database of Allergens (SDAP). Our method is based on the hypothesis that we can find surface patches on 3D structures of potential allergens with amino acid compositions similar to an epitope in a known allergen. We applied the Cross-React method to a diverse set of seven allergens, and successfully identified several cross-reactive allergens with high to moderate sequence identity which have also been experimentally shown to cross-react. Based on these findings, we suggest that Cross-React can be used as a predictive tool to assess protein allergenicity and cross-reactivity. Availability and Implementation: Cross-React is available at: http://curie.utmb.edu/Cross-React.html

Introduction : Differentiating between cross-reactivity and double sensitization is still a challenging issue in allergology. Aim : To differentiate cross-reactions accompanying latex allergy with the use of the ISAC test. Material and methods : Thirty-nine patients reporting immediate allergic reactions to latex were enrolled into the study (group A). The control group was comprised of 41 patients with allergic diseases not associated with latex (group B) and 20 healthy individuals (group C). Their history was recorded and skin prick tests were performed with latex, airborne and food allergens. Specific IgE against food allergens, latex (k82) and recombined latex allergens were determined. Immun o CAP ISAC test was performed with 103 molecules. Results : Sensitization to latex was found by means of skin tests in 16 cases and sIgE against latex was revealed in 12 cases (including 10 positive in both SPT and sIgE). In the ISAC test antibodies against recombined latex allergens were found in 8 patients with rHev β 6 as the most common. All the patients positive for rHev β 1, 5, 6, 8 had allergy or asymptomatic sensitization to food allergens cross-reacting with latex. Some reactions could not have been differentiated due to the lack of allergens in the ISAC test. Others, not related to latex-fruits syndrome were explained by cross-reactivity with other profilins or PR-10 proteins. Conclusions : Immun o CAP ISAC test could be useful in differentiating between cross-reactions and double sensitizations. However, in the case of latex its advantages are limited due to a small panel of allergens.

BACKGROUND—Pigeon breeder's disease is an extrinsic allergic alveolitis in the lungs of sensitised people, caused by hypersensitivity reactions to inhaled pigeon antigens. Antigens from different sources of the animal are used for diagnostic purposes, with serum being the most widely used. Bloom is rarely used; very little is known of its antigenicity and diagnostic performance, particularly when used with the enzyme linked immunosorbent assay (ELISA) method, which is the most popular test as it permits measurement of the antibody response. METHODS—To (a) standardise an ELISA for the measurement of specific IgG against pigeon serum and pigeon bloom extract; (b) to establish reference values for specific IgG in 73 non-exposed controls, (c) to show the presence of specific IgG against pigeon serum and bloom in serum samples of 17 patients with bird fancier's lung and 11 asymptomatic fanciers, and (d) to study the similarity of the two antigen sources by cross reactivity experiments. RESULTS—Reference values of specific IgG were defined with the 97.5 percentile (367.9 U/ml for pigeon serum and 953.7 U/ml for pigeon bloom extract). Of symptomatic patients 100% had values higher than the cut off for both antigens. In asymptomatic fanciers values were higher than the cut off for pigeon serum in 45% and bloom extract in 54%. Cross reactivity experiments showed that the two antigens differed in antigenic content although some components may be common to both. CONCLUSION—The ELISA methods used proved to be useful tools for evaluating specific IgG antibody responses against both antigens. The diagnostic performance of both ELISA methods performed with these antigen sources was similar, showing very high sensitivity but moderate specificity. Although some antigenic similarity was found between pigeon serum and bloom extract, cross reactivity studies showed that various antigens seemed to be specific to the bloom extract. However, the antigens responsible for pigeon breeder's disease seem to be present in both antigenic sources. Keywords: antibodies; pigeons; ELISA

Glycyrrhizic acid (GL) is a major active compound of licorice. The specific monoclonal antibody (MAb) (designated as 8F8A8H42H7) against GL was produced with the immunogen GL–BSA conjugate. The dissociation constant (K d) value of the MAb was approximately 9.96×10−10 M. The cross reactivity of the MAb with glycyrrhetic acid was approximately 2.6%. The conventional indirect competitive enzyme-linked immunosorbent assay (icELISA) and simplified icELISA adapted with a modified procedure were established using the MAb. The IC50 value and the detect range by the conventional icELISA were 1.1 ng mL−1 and 0.2–5.1 ng mL−1, respectively. The IC50 value and the detect range by the simplified icELISA were 5.3 ng mL−1 and 1.2–23.8 ng mL−1, respectively. The two icELISA formats were used to analyze GL contents in the roots of wild licorice and different parts of cultivated licorice (Glycyrrhiza uralensis Fisch). The results obtained with the two icELISAs agreed well with those of the HPLC analysis. The correlation coefficient was more than 0.98 between HPLC and the two icELISAs. The two icELISAs were shown to be appropriate, simple, and effective for the quality control of raw licorice root materials.

A complex and varied antibody response to allergens, in particular Dermatophagoides pteronyssinus, is observed in the serum of allergic patients. To examine the role of cross-reacting antigenic determinants in allergy, a strategy has been devised for isolating antibody populations that bind to individual polypeptides of an allergen source separated by sodium dodecyl sulfate-polyacrylamide electrophoresis and electroblotted to nitrocellulose. Polypeptides are identified on the blots, desorbed and coupled to CNBr-activated paper discs. The coupled discs are then used as immunoadsorbent surfaces for the binding of antibodies from allergic sera. Antibodies eluted from the polypeptide immunoadsorbents are then used to reprobe protein blots of the allergen. Results are presented which demonstrate strong cross-reaction between two major mite allergens, Der p I and Der p III, and some weaker intra-species cross-reactivities.

RAST-inhibition studies were performed by use of whole venom sac extracts to inhibit binding to purified venom allergens from various vespid wasps. Immunodiffusion studies were also performed with rabbit antisera raised against purified venom proteins. Immunologic cross-reactivity was demonstrated among the hyaluronidases, phospholipases, and antigen 5 s from both subgenera of yellow jackets, white-faced hornets, and paper wasps. The paper wasp hyaluronidase and antigen 5 were less related to those of the other three species than those of yellow jackets and hornets. It appears that immunologic cross-reactivity is the major mechanism of multiple allergic sensitivity to vespid venoms. Therapy with only the primary venom should be sufficient to protect against reactions from cross-reacting venoms.

We have used the RAST-inhibition technique in homologous and heterologous inhibitions of the Aspergillus fumigatus, Alternaria tenius and Cladosporium herbarum RAST assays followed by the immunoblot technique to assess the degree of shared allergenic determinants in these extracts. In the Aspergillus RAST, there was little or no inhibition with Cladosporium and Alternaria , but considerable cross-reactivity was found between Alternaria and Cladosporium . Inhibition by Alternaria of the Cladosporium RAST was found in a dose-dependent fashion in on serum, whereas all four sera in the Alternaria RAST were inhibited by Cladosporium in this fashion. Logit transformation of the Alternaria RAST-inhibition curves produced common slopes with Alternaria and Cladosporium in three of the four sera, indicating their immunologic identity. The immunoblot technique confirmed the degree of cross-reactivity found by RAST inhibition among the molds. This evidence of common allergenic determinants in these fungi could help to explain the observation that mold-allergic patients often have skin test reactions to several fungi.

Summary Bovine herpesvirus type 1 (BHV-1) glycoprotein gIV was purified by affinity chromatography. Purified preparations showed two distinct components of 71 K and 140 K following electrophoresis in sodium dodecyl sulphate polyacrylamide gels. The polypeptides were separated, excised from the gel and used to immunize rabbits; the resulting antisera showed a high degree of cross reactivity indicating that these polypeptides represent monomeric and dimeric forms of the same glycoprotein. Purified gIV was also used to develop a gIV-specific panel of monoclonal antibodies. Neutralizing monoclonal antibodies directed against gIV were conjugated to horseradish peroxidase and subjected to competition binding assays by ELISA. Three distinct neutralizing antigenic domains on gIV were identified. Domain 1 comprised two overlapping epitopes, whereas domain 2 was represented by a single monoclonal antibody. The third antigenic domain was made up of a complex of four identical or overlapping epitopes designated 3a, b, c, and d. Evidence is presented suggesting that domain 1 of gIV may be involved in penetration of the virus into the cell.

Summary A possible antigenic relationship between the porcine enteropathogenic coronavirus-like agent (CVLA) and 6 known coronaviruses was examined by immunoelectron microscopy (IEM) and by immunofluorescence (IF). CVLA did not show cross reactivity with infectious bronchitis virus, transmissible gastroenteritis virus (TGEV), canine coronavirus (CCV) hemagglutinating encephalomyelitis virus (HEV), neonatal calf diarrhea coronavirus (NCDCV) or feline infectious peritonitis virus (FIPV). Antigenic relationship was detected by IEM between TGEV and CCV, NCDCV and HEV and by IF between TGEV and CCV, TGEV and FIPV, HEV and NCDCV.

Graphical abstract, Highlights • A future COVID-19 vaccine is being developed at unprecedented speed to augment immune system defenses. • This paper presents myriad infection/vaccine-related mechanisms that could adversely impact vaccine effectiveness and safety. • It is unclear how the present accelerated vaccine development program can address these longer-term impact vaccine safety issues., A degraded/dysfunctional immune system appears to be the main determinant of serious/fatal reaction to viral infection (for COVID-19, SARS, and influenza alike). There are four major approaches being employed or considered presently to augment or strengthen the immune system, in order to reduce adverse effects of viral exposure. The three approaches that are focused mainly on augmenting the immune system are based on the concept that pandemics/outbreaks can be controlled/prevented while maintaining the immune-degrading lifestyles followed by much of the global population. The fourth approach is based on identifying and introducing measures aimed at strengthening the immune system intrinsically in order to minimize future pandemics/outbreaks. Specifically, the four measures are: 1) restricting exposure to virus; 2) providing reactive/tactical treatments to reduce viral load; 3) developing vaccines to prevent, or at least attenuate, the infection; 4) strengthening the immune system intrinsically, by a) identifying those factors that contribute to degrading the immune system, then eliminating/reducing them as comprehensively, thoroughly, and rapidly as possible, and b) replacing the eliminated factors with immune-strengthening factors. This paper focuses on vaccine safety. A future COVID-19 vaccine appears to be the treatment of choice at the national/international level. Vaccine development has been accelerated to achieve this goal in the relatively near-term, and questions have arisen whether vaccine safety has been/is being/will be compromised in pursuit of a shortened vaccine development time. There are myriad mechanisms related to vaccine-induced, and natural infection-induced, infections that could adversely impact vaccine effectiveness and safety. This paper summarizes many of those mechanisms.

We exploit the cross-reactivity of dengue (DENV) and Zika (ZIKV) virus polyclonal antibodies for nonstructural protein 1 (NS1) to construct a selective sensor that can detect yellow fever virus (YFV) NS1 in a manner similar to chemical olfaction. DENV and ZIKV antibodies were screened for their ability to bind to DENV, ZIKV, and YFV NS1 by enzyme linked immunosorbent assay (ELISA) and in pairs in paper immunoassays. A strategic arrangement of antibodies immobilized on paper and conjugated to different colored gold NPs was used to distinguish the three biomarkers. Machine learning of test area RGB values showed that with two spots, readout accuracies of 100% and 87% were obtained for both pure NS1 and DENV/YFV mixtures, respectively. Additional image preprocessing allowed differentiation between all four DENV serotypes with 92% accuracy. The technique was extended to hack a commercial DENV test to detect YFV and ZIKV by augmentation with DENV and ZIKV polyclonal antibodies.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Cultivated for the crispy petioles and round, fleshy, and flavored hypocotyl celery and celeriac have over two centuries of breeding history in Europe. In this review paper we summarized the most recent advances touching when necessary the historical context of celery and celeriac breeding. In the post genomic era of research, the genome sequence of celery is only partially available. We comprised however in this paper the most important aspects of celery genetics that are available today and have applicability in celery modern cultivars development. We discussed the problems and traits that drive the main celery and celeriac breeding goals, like hybrid seed production, disease resistance, and interesting enlarged hypocotyl and petiole characteristics. Besides the classical breeding traits we covered the potential of integration of existing cultivars as sources for consumer oriented traits like nutraceuticals and health promoting substances. Sustainability is a subject that is continuously growing in popularity and we looked at the genetic base of celery and celeriac that makes them sources for abiotic stress resistance and candidates for phytoremediation. We explored the fundamental concepts gained in various fields of celery and related species research, as resources for future improvement of celery and celeriac germplasm. We forecast what the next years will bring to Apium breeding.

The current paper suggests that egg allergy may arise due to microbial proteins that are homologous to egg allergens. These microbial proteins elicit an allergic response and lead to the development of specific microbial IgE molecules. These molecules cross-react with egg allergens and result in egg allergy. Some examples of microbial proteins that share similar sequences as egg allergens are presented in this paper. Food allergy has a negative impact on the quality of life and nutrition. In addition, it can lead to life-threatening reactions. The pathogenesis of food allergy is still not fully understood. Many young children develop food allergy with no known prior ingestion of the food allergen in question. The strong connection between food allergy and atopic dermatitis has been well-documented. While the prevalence of food allergy in the general pediatric population is 4–5%, the prevalence of food allergy in atopic dermatitis is at least 20% (1). This connection between food allergy and atopic dermatitis has led to the suggestion that the skin may be the site of food IgE sensitization, leading to food allergy. The dual-allergen-exposure hypothesis suggests that food allergens are sensitized via eczema, whereas early gastro-intestinal exposure leads to tolerance (2). This hypothesis is supported by multiple basic studies that provide evidence for IgE sensitization via the skin [reviewed in (3)]. In addition, it has been shown that environmental level of peanut allergens is increased in children who developed peanut allergy (4). The early introduction of peanut has led to the prevention of peanut allergy (5). Likewise, early introduction of egg has also met with some success, although the results were not as consistent (3). It has been suggested that improvement in eczema, in addition to early introduction of egg, is needed for successful prevention of egg allergy (6). This suggestion further highlights the importance of skin in the pathogenesis of food allergy. A hypothesis is proposed here that the interaction between the neonatal skin and microbial proteins is important for the development of IgE sensitization and egg allergy. The current hypothesis predicts the presence of microbial proteins that are homologous to egg allergens. Table 1 shows the microbial proteins that share homology with the IgE-binding domains of Gal d 1 (ovomucoid) (7). Four out of the 6 microbial proteins share > 60 % identity with a clinically-relevant IgE-binding region of Gal d 1 (FNPVCGTDGVTYDN) (8). Significant homology was also found between microbial proteins and Gal d 2 (ovalbumin), but no homology was found in the IgE-binding domains of Gal d 2 (9) (data not shown). To further confirm the correlation between microbial proteins and Gal d 1, prospective studies may be carried out to look for these microbial pathogens in atopy-prone neonates and correlate with neonates who eventually develop allergy to Gal d 1. These microbial pathogens can also be inoculated in animal models to show the development of specific IgE that cross-react with Gal d 1. Atopy-prone neonates who are born to parent with atopic dermatitis, asthma or allergic rhinitis have inherent skin barrier defects that predispose them to develop atopic dermatitis. Microbial pathogens are capable of evading these barrier defects to interact with the cutaneous immune system in these children. The processing of microbial proteins by antigen-presenting cells and subsequent presentation of antigenic peptides to T helper type 2 cells leads to the production of IL-4 and IL-13, which induce B cells to express specific IgE molecules. Bacterial allergy has been described more than half a century ago (10). It is also known that specific IgE to staphylococcal toxins can develop in young children with atopic dermatitis (11). Bacteria such as bacteroides can be acquired during birth or they can be part of neonates' microbiome (12). The skin also contains a wide array of microbial pathogens that can participate in the development of allergy (13). More recent data suggests that many microbial organisms (e.g., proteobacteria and archaea) previously thought to exist only in the environment such as soil, fresh or marine waters are now found to be part of the human skin microbiome (14, 15). Proteobacteria and bacteroides are also known to be present in atopic dermatitis lesions (16). The current proposal is conceptual that microbial proteins can be a sensitizing source in the development of food allergy in predisposed children. Preliminary data also suggests the presence of homologous proteins between microbes and other food allergens including peanut and cow's milk (unpublished data). Whether these microbial pathogens have a direct interaction with neonates, leading to the development of specific IgE, remains to be proven. It is possible that the prevention of egg allergy requires a different approach by targeting microbial pathogens. Table 1 Microbial proteins that share similar sequences as the IgE-binding domains of Gal d 1.

A large number of chemical structures that interact with G-protein coupled receptors (GPCRs) have been disclosed in patents or published papers. Most of these compounds are selective for a given protein target; however, it is well recognized that some GPCR-drugs interact with multiple targets. Using a literature database, we have identified compounds that act on different GPCRs. These protein targets are usually divided in three main classes, A, B and C, based on sequence similarity, but they can also be grouped pharmacologically based on endogenous ligand characteristics. In this paper, we specifically focus on compounds able to recognize two different classes or different pharmacological clusters within the same class. Despite the large number of GPCR ligands described in the literature, we identified a limited number of molecules acting on both classes A and B, only few acting on classes A and C and none acting on class B and C receptors. A search for bi- or multi-potent compounds exhibiting activities on different pharmacological clusters of class A receptors revealed cases of cross reactivity, the most frequent concerning amine and peptide receptor clusters.

The Zika Virus (ZIKV) was propelled to international attention during its outbreak from 2015-2016. Interestingly, the most recent outbreak was not ZIKV’s first, although it proved to be the most widespread and impactful, with millions affected in South America, Asia and Africa. Presently no longer considered a global emergency, ZIKV has managed to invoke fear and realization of the susceptibility of the global population to rapidly evolving viruses. In addition, the difficulty of diagnosing the virus demonstrates a deficiency in a rapid, virus specific, and accurate diagnostic tool for the family of flaviviruses that ZIKV belongs to. This paper reviews the approved identification methods along with an analysis of the advantage and disadvantages of each, as well as emerging alternative approaches in ZIKV diagnosis. Common problems with currently utilized methods include slow turnover time, limited throughput, need for further testing to confirm diagnosis, narrow sample compatibility, and cross reactivity to DENV and other similar viruses, Although newer methods discussed in the paper, namely Electrogenerated Chemiluminescence (ECL) and Reporter Virus Neutralization Test (RVNT), show improvement in throughput quantity, speed, and efficiency, it is not certain whether these tests are virus specific and will not react in the presence DENV. The rapidity of diagnosis is important in ensuring timely access to treatment as well as tracking and containing future possible epidemics, Concurrently, virus specificity is equally crucial in ensuring correct diagnosis. Thus, the challenge lies in finding the balance between the two.

The specificity of an immunohistochemical reaction is guaranteed by two sets of controls. Positive controls verify the specificity of the primary antibody and demonstrate that it binds only to the protein which was used as an immunogen. Negative controls ensure that the labelling technique is specific and that the primary antibody is responsible for generation of the immunostaining. In fact, the production of a labelling may also be related to cross reactivity or to non-specific physical or chemical interactions. This paper reviews the characteristics of various epitopes and antibodies, describes different strategies which prove the specificity of the immunohistochemical reaction in research or diagnostic pathology and point towards the essential information which should be reported in a paper.

Some infants allergic to cow milk-based formula are also sensitive to soybean-based formula. This paper aimed to explore the association of IgE and IgG cross-reactivity between αS1-casein in cow milk (CM) and soybean proteins. The IgE and IgG cross-reactive allergens and epitopes were identified using sera from infants allergic to CM or mice monoclonal antibodies. The AA sequence alignment was performed using bioinformatics software. Finally, the digestion and heating stability of the cross-reactive allergen were explored by sodium dodecyl sulfate (SDS)-PAGE and Western blotting. The results showed that the IgE and IgG cross-reactive allergen was α subunit of β-conglycinin named Gly m Bd 60K. The IgE and IgG epitopes were the sequences at AA 319-341 and AA 164-182. No intact Gly m Bd 60K allergen could be observed after 2 min in simulated gastric fluid by SDS-PAGE. Heating did not change IgE and IgG cross-reactivity by Western blotting. Therefore, the existence of cross-reactivity between CM αS1-casein and soybean proteins possibly contributes to the frequently observed cosensitization for these allergens in cow milk-allergic patients. The same IgE- and IgG-binding epitopes of cross-reactive allergens may provide important information for elucidation of the association between IgG and IgE antibody generation.

This paper describes data on allergies caused by food (vegetable) and their negative impact on the nutritional balance of the human body. Allergic responses to vegetables such as tomatoes, capsicum and spinach are next to fish, eggs and nuts. Epitopes such as NP24 (allergens) are one of the salt-induced allergenic proteins found in the thaumatin-like protein (TLP) family. The mechanism of allergenicity of TLP found in Solanum lycopersicum (Tomato) fruit is poorly studied. Here we demonstrated allergenicity conferred by the NP24 protein found in Tomato. The data on the cross-reactivity of NP24 protein was generated using Allergen Online and Allermatch tools. Tomato allergenic protein epitope shows a significant identity of with allergens reported in Capsicum, Olive, Kiwi, Tobacco and Banana allergens. Hence, the datasets of sequences, comparative analysis and homology epitope mapping over three dimensional (3D) structures revealed that NP24 has higher cross-reactivity to Capsicum and Tobacco proteins. Thus, this data probably act as limelight for planning wet lab experiments. Keywords: NP24 protein, Epitope mapping, Fruit and vegetable allergies, Oral allergy syndrome, IgE antibody

An enzyme immunoassay (EIA) for the measurement of serum testosterone has been developed using nitrocellulose paper discs as the solid support. The paper discs (6 mm diameter) coated with testosterone-specific antibody were incubated with testosterone and testosterone-peroxidase conjugate in glass tubes. The amount of testosterone present in samples could be estimated from the bound peroxidase activity. The assay was validated by comparison with a microtitre plate-based ELISA and a commercial radioimmunoassay (RIA) kit. The correlation coefficient between RIA and EIA was 0.84. No significant cross reactivity was observed with other steroids, except for dihydrotestosterone. The inter- and intra-assay coefficients of variation were 4.4 and 9.6% respectively. The preservation and transport of the coated paper discs are convenient and the overall cost of the method is less than that of other methods.

BackgroundProgress in characterising the humoral immune response to Severe Acute Respiratory Syndrome 2 (SARS-CoV-2) has been rapid but areas of uncertainty persist. Assessment of the full range of evidence generated to date to understand the characteristics of the antibody response, its dynamics over time, its determinants and the immunity it confers will have a range of clinical and policy implications for this novel pathogen. This review comprehensively evaluated evidence describing the antibody response to SARS-CoV-2 published from 01/01/2020-26/06/2020.MethodsSystematic review. Keyword-structured searches were carried out in MEDLINE, Embase and COVID-19 Primer. Articles were independently screened on title, abstract and full text by two researchers, with arbitration of disagreements. Data were double-extracted into a pre-designed template, and studies critically appraised using a modified version of the Public Health Ontario Meta-tool for Quality Appraisal of Public Health Evidence (MetaQAT) tool, with resolution of disagreements by consensus. Findings were narratively synthesised.Results150 papers were included. Most studies (113 or 75%) were observational in design, were based wholly or primarily on data from hospitalised patients (108, 72%) and had important methodological limitations. Few considered mild or asymptomatic infection. Antibody dynamics were well described in the acute phase, up to around three months from disease onset, but the picture regarding correlates of the antibody response was inconsistent. IgM was consistently detected before IgG in included studies, peaking at weeks two to five and declining over a further three to five weeks post-symptom onset depending on the patient group; IgG peaked around weeks three to seven post-symptom onset then plateaued, generally persisting for at least eight weeks. Neutralising antibodies were detectable within seven to 15 days following disease onset, with levels increasing until days 14–22 before levelling and then decreasing, but titres were lower in those with asymptomatic or clinically mild disease. Specific and potent neutralising antibodies have been isolated from convalescent plasma. Cross-reactivity but limited cross-neutralisation with other human coronaviridae was reported. Evidence for protective immunity in vivo was limited to small, short-term animal studies, showing promising initial results in the immediate recovery phase.ConclusionsLiterature on antibody responses to SARS-CoV-2 is of variable quality with considerable heterogeneity of methods, study participants, outcomes measured and assays used. Although acute phase antibody dynamics are well described, longer-term patterns are much less well evidenced. Comprehensive assessment of the role of demographic characteristics and disease severity on antibody responses is needed. Initial findings of low neutralising antibody titres and possible waning of titres over time may have implications for sero-surveillance and disease control policy, although further evidence is needed. The detection of potent neutralising antibodies in convalescent plasma is important in the context of development of therapeutics and vaccines. Due to limitations with the existing evidence base, large, cross-national cohort studies using appropriate statistical analysis and standardised serological assays and clinical classifications should be prioritised.

Various studies indicate that vaccination, especially with pneumococcal vaccines, protects against symptomatic cases of SARS-CoV-2 infection and death. This paper explores the possibility that pneumococcal vaccines in particular, but perhaps other vaccines as well, contain antigens that might be cross-reactive with SARS-CoV-2 antigens. Comparison of the glycosylation structures of SARS-CoV-2 with the polysaccharide structures of pneumococcal vaccines yielded no obvious similarities. However, while pneumococcal vaccines are primarily composed of capsular polysaccharides, some are conjugated to cross-reacting material CRM197, a modified diphtheria toxin, and all contain about three percent protein contaminants, including the pneumococcal surface proteins PsaA, PspA and probably PspC. All of these proteins have very high degrees of similarity, using very stringent criteria, with several SARS-CoV-2 proteins including the spike protein, membrane protein and replicase 1a. CRM197 is also present in Haemophilus influenzae type b (Hib) and meningitis vaccines. Equivalent similarities were found at lower rates, or were completely absent, among the proteins in diphtheria, tetanus, pertussis, measles, mumps, rubella, and poliovirus vaccines. Notably, PspA and PspC are highly antigenic and new pneumococcal vaccines based on them are currently in human clinical trials so that their effectiveness against SARS-CoV-2 disease is easily testable.

While COVID-19 continues to spread across the globe, diligent efforts are made to understand its attributes and dynamics to help develop treatment and prevention measures. The paradox pertaining to children being the least affected by severe illness poses exciting opportunities to investigate potential protective factors. In this paper, we propose that childhood vaccination against pertussis (whooping cough) might play a non-specific protective role against COVID-19 through heterologous adaptive responses in this young population. Pertussis is a vaccine-preventable infectious disease of the respiratory tract and it shares many similarities with COVID-19 including transmission and clinical features. Although pertussis is caused by a bacterium (Bordetella pertussis) while COVID-19 is a viral infection (SARS-CoV-2), previous data showed that cross-reactivity and heterologous adaptive responses can be seen with unrelated agents of highly divergent groups, such as between bacteria and viruses. While we build the arguments of this hypothesis on theoretical and previous empirical evidence, we also outline suggested lines of research from different fields to test its credibility. Besides, we highlight some concerns that may arise when attempting to consider such an approach as a potential public health preventive intervention against COVID-19.

It is known since Thomas Francis, Jr. published his first paper on original antigenic sin in 1960 that the first infection(s) with influenza virus leaves a special immunological imprint which shapes immune responses to future infections with antigenically related influenza virus strains. Imprinting has been implicated in both protective effects as well as blunting of the immune response to vaccines. Despite the fact that this phenomenon was already described almost 60 years ago, we have very little detailed knowledge of the characteristics and breadth of the immune response to the first exposure(s) to influenza virus in life and how this compares to later exposure as adults. Here, we investigate these immune responses in detail using an influenza virus protein microarray. While our findings are mostly descriptive in nature and based on a small sample size, they provide a strong basis for future large-scale studies to better understand imprinting effects., In contrast to influenza virus vaccination, natural infection induces long-lived and relatively broad immune responses. However, many aspects of the antibody response to natural infection are not well understood. Here, we assessed the immune response after H1N1 influenza virus infection in children and adults in a Nicaraguan household transmission study using an influenza virus protein microarray (IVPM). This technology allows us to simultaneously measure IgG and IgA antibody responses to hemagglutinins of many different virus strains and subtypes quantitatively with a high throughput. We found that children under 6 years of age responded to natural infection with a relatively narrow response that targeted mostly the hemagglutinin of the strain that caused the infection. Adults, however, have a much broader response, including a boost in antibodies to many group 1 subtype hemagglutinins. Also, a strong recall response against historic H1 hemagglutinins that share the K133 epitope with the pandemic H1N1 virus was observed. Of note, some children, while responding narrowly within H1 and group 1 hemagglutinins, induced a boost to H3 and other group 2 hemagglutinins when infected with H1N1 when they had experienced an H3N2 infection earlier in life. This is an interesting phenomenon providing evidence for immune imprinting and a significant new insight which might be leveraged in future universal influenza virus vaccine strategies. Finally, preexisting immunity to pandemic H1 hemagglutinins was significantly associated with protection from infection in both children and adults. In adults, preexisting immunity to non-H1 group 1 hemagglutinins was also significantly associated with protection from infection.

The development and introduction of new dietary protein sources has the potential to improve food supply sustainability. Understanding the potential allergenicity of these new or modified proteins is crucial to ensure protection of public health. Exposure to new proteins may result in de novo sensitization, with or without clinical allergy, or clinical reactions through cross-reactivity. In this paper we review the potential of current methodologies (in silico, in vitro degradation, in vitro IgE binding, animal models and clinical studies) to address these outcomes for risk assessment purposes for new proteins, and especially to identify and characterise the risk of sensitization for IgE mediated allergy from oral exposure. Existing tools and tests are capable of assessing potential crossreactivity. However, there are few possibilities to assess the hazard due to de novo sensitization. The only methods available are in vivo models, but many limitations exist to use them for assessing risk. We conclude that there is a need to understand which criteria adequately define allergenicity for risk assessment purposes, and from these criteria develop a more suitable battery of tests to distinguish between proteins of high and low allergenicity, which can then be applied to assess new proteins with unknown risks. © 2017 The Authors Chemicals/CAS: immunoglobulin E, 37341-29-0

Summary The antigenic reactivity of constituents of Schistosoma mansoni and peanut (Arachis hypogaea) was investigated to determine whether identical antigenic epitopes possessed by both organisms provided a possible explanation for the negative correlation between chronic schistosome infection and atopy to allergens. Aqueous extracts of peanuts were probed in Western immunoblots with rabbit IgG antibodies raised against the egg, cercarial and adult worm stages of S. mansoni. Several molecules in the peanut extract were antigenically reactive with antibodies from the various rabbit anti-schistosome sera. A pair of cross-reactive peanut molecules at ~30 000–33 000 molecular weight was purified and both proteins were identified by mass spectrometric analysis as the peanut allergen Ara h 1. Anti-S. mansoni soluble egg antigen antibodies that were eluted off the peanut molecules reacted with two S. mansoni egg antigens identified by mass spectrometry as IPSE/α-1 and κ-5. Alignments of the amino acid sequences of Ara h 1 and either IPSE/α-1 or κ-5 revealed a low level of peptide sequence identity. Incubation of nitrocellulose paper carrying electrophoresed peanut molecules, six constituents of other allergic plants and S. mansoni egg antigens in a mild solution of sodium metaperiodate before probing with antibodies, inhibited most of the cross-reactivities. The results are consistent with the antigenic cross-reactive epitopes of S. mansoni egg antigens, peanut and other allergic plants being cross-reactive carbohydrate determinants (CCDs). These findings are novel and an explanation based on ‘blocking antibodies’ could provide an insight for the inverse relationship observed between schistosome infection and allergies.

We read with interest the paper by Schmid et al. [(1)][1], who described elevated cardiac troponin T (cTnT) concentrations in patients with skeletal myopathies in which they suggested cross-reaction of the high-sensitivity (hs-)cTnT immunoassay with skeletal muscle troponin T (skTnT) isoforms as

BACKGROUND: Hymenoptera stings can cause severe anaphylaxis in untreated venom-allergic patients. A correct diagnosis regarding the relevant species for immunotherapy is often hampered by clinically irrelevant cross-reactivity. In vespid venom allergy, cross-reactivity between venoms of different species can be a diagnostic challenge. To address immunological IgE cross-reactivity on molecular level, seven recombinant antigens 5 of the most important Vespoidea groups were assessed by different diagnostic setups.METHODS: The antigens 5 of yellow jackets, hornets, European and American paper wasps, fire ants, white-faced hornets, and Polybia wasps were recombinantly produced in insect cells, immunologically and structurally characterized, and their sIgE reactivity assessed by ImmunoCAP, ELISA, cross-inhibition, and basophil activation test (BAT) in patients with yellow jacket or Polistes venom allergy of two European geographical areas.RESULTS: All recombinant allergens were correctly folded and structural models and patient reactivity profiles suggested the presence of conserved and unique B-cell epitopes. All antigens 5 showed extensive cross-reactivity in sIgE analyses, inhibition assays, and BAT. This cross-reactivity was more pronounced in ImmunoCAP measurements with venom extracts than in sIgE analyses with recombinant antigens 5. Dose-response curves with the allergens in BAT allowed a differentiated individual dissection of relevant sensitization.CONCLUSIONS: Due to extensive cross-reactivity in various diagnostic settings, antigens 5 are inappropriate markers for differential sIgE diagnostics in vespid venom allergy. However, the newly available antigens 5 from further vespid species and the combination of recombinant allergen-based sIgE measurements with BAT represents a practicable way to diagnose clinically relevant sensitization in vespid venom allergy.

Immunoaffinity columns (IACs) are a well-established tool in the determination of regulated mycotoxins in food and feed commodities. However, they also have the potential to become attractive pre-concentration and clean-up materials for the determination of masked (also called modified) mycotoxins, which have been recognised as important contributors to the toxicological hazard deriving from fungal spoilage of goods. However, the information available in the literature concerning the cross-reactivity of DON-IACs against the major conjugates (DON-3-G, 15-AcDON and 3-AcDON) is incomplete and often contradictory. We have carried out a detailed characterisation of the cross-reactivity of the four main IACs brands against DON and its conjugates as well as an assessment of the competition among the analytes. Only one IAC enabled the simultaneous analysis of all relevant DON forms while two missed 15-AcDON and the fourth one missed DON-3-G and 3-AcDON. In the case of the multivalent IAC, the analytes modified at the C-3 position compete for the antibody binding with preference for 3-AcDON (less spatially hindered) while DON-3-G has the more-hindered access to the active sites. Taking into consideration the levels of DON conjugates existing in real samples, the cross-reactivity of one DON-IAC allows a quantitative analysis of all of these analytes. Important but rather neglected aspects such as the continuous supply of IACs with identical characteristics, and of columns which are strictly blank, are also addressed in this paper., Graphical Abstract

Introduction: Diagnosing and managing drug hypersensitivity is challenging because there are no clear limits between different types of drug reactions. Distinguishing between type A (predictable) and type B (hypersensitivity) reactions when a drug is introduced on the market is not easy. When many people use a drug, adverse reactions can occur, conditioned by diverse genetic profiles, viral infections or concomitant therapy. Occasionally the only tool clinicians have on which to base the diagnosis is the clinical history. Skins tests or in vitro tests sometimes have low sensitivity or are unavailable, and drug provocation tests may be dangerous or strictly forbidden in case of severe cutaneous reactions. Areas covered: This paper reviews the diagnosis and management of the two main types of immunological reactions: IgE-mediated immediate drug hypersensitivity reactions (IDHRs) and non-immediate drug hypersensitivity reactions (NIDHRs). Expert commentary: Although Europe and the United States use different diagnostic methods, patients with history of drug hypersensitivity must avoid the suspicious drug, and clinicians must assess tolerance to safe alternatives under medical surveillance. Sometimes desensitization may be required. There is a consensus about the need to perform genetic testing for specific drugs and give patients proper documentation to prevent future exposure to culprit drugs.

Dengue is a mosquito borne infection, which in recent years has become a major international public health concern. Annually, 100 million dengue virus infections are reported worldwide. The nonstructural protein 1 (NS1) of dengue virus is a useful target for diagnostics of dengue infection since the protein is abundantly circulating in the blood during acute phase of the disease, in both primary and secondary infections. This research paper highlights the development of a panel of Mab and bsMab for dengue NS1 detection. The P148 series of Mabs showed high specificity for recombinant dengue NS1 antigen. These antibodies showed no cross reactivity with recombinant dengue envelope protein and other viral proteins. The hybrid-hybridoma approach to generate the P156.1 and P156.2 bsMabs from the P148 monoclonal antibody method was used during this study. Furthermore, the affinity purification provided good yields of quadromas associated with HRPO in two steps. Direct detection method involved coating of plates with different concentrations of recombinant antigen and detecting with bsMab. Sensitive sandwich assay with Mabs and bsMabs was also done. Detection of nonstructural dengue antigens may be of benefit for early and rapid diagnosis of dengue infection due to their long half-life in the blood.

In the paper, an enzyme-linked immunosorbent immunoassay (ELISA) for detection of enrofloxacin was described using one new derivative of enrofloxacin as coating hapten, resulting in surprisingly high sensitivity and specificity. Incorporation of aminobutyric acid (AA) in the new derivative of enrofloxacin had decreased the IC50 of the ELISA for enrofloxacin from 1.3 μg L−1 to as low as 0.07 μg L−1. The assay showed neglect cross-reactivity for other fluoroquinolones but ofloxacin (8.23%), marbofloxacin (8.97%) and pefloxacin (7.29%). Analysis of enrofloxacin fortified chicken muscle showed average recoveries from 81 to 115%. The high sensitivity and specificity of the assay makes it a suitable screening method for the determination of low levels of enrofloxacin in chicken muscle without clean-up step.

In our recent paper we presented data indicating a skeletal muscle origin of measured cardiac troponin T (cTnT) in skeletal myopathy patients [(1)][1]. We thank Dr. Vroemen and colleagues for their letter in response to this article. In contrast to our experiments where we used crude skeletal

The authors wish to make the following corrections to this paper [...]