Use of a baculovirus-expressed structural protein for the detection of antibodies to foot-and-mouth disease virus type A by a blocking enzyme-linked immunosorbent assay.
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ABSTRACT:
SUBMITTER: Ko YJ
PROVIDER: S-EPMC2812095 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Foot-and-mouth disease (FMD) is a highly contagious disease and one of the most economically important diseases of livestock. Vaccination is an important measure to control FMD and selection of appropriate vaccine strains is crucial. The objective of this study was to select a vaccine candidate and to evaluate the potential of a blocking ELISA for detecting neutralizing antibodies (NA-ELISA) in vaccine strain selection. Binary ethylenimine inactivated vaccines, prepared from four representative circulating strains (FMDV O/Mya/98, SCGH/CHA/2016, O/Tibet/99, and O/XJ/CHA/2017) belonging to four lineages within three different topotypes of FMD virus (FMDV) serotype O in China, were used to vaccinate cattle (12-13 animals for each strain), sheep (12-13 animals for each strain), and pigs (10 animals for each strain). The results of immunogenicity comparison showed that O/XJ/CHA/2017 exhibited the highest immunogenicity among the four strains in pigs, cattle, and sheep both by NA-ELISA and virus neutralizing test (VNT). Cross-neutralization analysis indicated that O/XJ/CHA/2017 displayed broad antigen spectrum and was antigenically matched with other three representative strains both by NA-ELISA and VNT. In addition, A significant correlation (p < 0.0001) was observed between the NA-ELISA titers and the VNT titers for four representative strains. The results showed that O/XJ/CHA/2017 was a promising vaccine strain candidate and NA-ELISA was comparable to VNT in neutralizing antibodies detection and could be used as the reference test system for vaccine strain selection.
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Project description:Feline morbillivirus (FeMV) has been associated with feline health, although its exact role in pathogenesis is still debated. In this study, an indirect enzyme-linked immunosorbent assay (i-ELISA) targeting a recombinant matrix protein of FeMV (rFeMV-M) was developed and assessed in comparison to a Western blotting (WB) assay. The i-ELISA was evaluated using blood samples from 136 cats that were additionally tested with real-time reverse-transcription PCR (RT-qPCR). The i-ELISA exhibited a sensitivity of 90.1%, specificity of 75.6%, positive predictive value of 88.2%, and negative predictive value of 79.1%. The agreement between i-ELISA and WB analyses was substantial (a κ coefficient of 0.664 with a 95% confidence interval of 0.529 to 0.799). Within the study group, 68.4% (93/136) of the cats were serologically positive in the i-ELISA and 66.9% (91/136) in the WB assay, with 11.8% (11/93) of false positivity with the i-ELISA. However, only 8.1% (11/136) of the cats tested positive for FeMV using RT-qPCR (p < 0.001). The developed i-ELISA proved effective in identifying FeMV-infected cats and indicated the prevalence of FeMV exposure. Combining FeMV antibody detection through i-ELISA with FeMV RT-qPCR could offer a comprehensive method to determine and monitor FeMV infection status. Nevertheless, this assay still requires refinement due to a significant number of false positive results, which can lead to the misdiagnosis of cats without antibodies as having antibodies. This study also provided the first evidence of seroprevalence against FeMV among cat populations in Thailand, contributing valuable insights into the geographic distribution and prevalence of this virus.
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Project description:The level of cotinine in biological specimens, such as serum, urine, and saliva, measured by gas or liquid chromatography is the most validated and reliable indicator of exposure to tobacco smoke. However, chromatographic methods are not always suitable for all types of situations.We validated a commercially available enzyme-linked immunosorbent assay (ELISA) that uses a polyclonal antibody to cotinine as a practical alternative to chromatographic methods.The cotinine antibody cross-reacts to 3-hydroxycotinine (3HC) and its glucuronide, thus generating a value for immunoreactive (IR) cotinine, which is a complex comprising cotinine, 3HC, and 3HC-glucuronide. The levels of IR cotinine in the urine of kindergarten children closely correlated with those of cotinine measured by gas chromatography-mass spectrometry (GC-MS) and reflected the smoking behavior of their parents more precisely than cotinine levels determined by GC-MS.Our findings showed that the cotinine-based ELISA can be a practical biomarker of exposure to tobacco smoke.
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Project description:The existing methods of callose quantification include epifluorescence microscopy and fluorescence spectrophotometry of aniline blue-stained callose particles, immuno-fluorescence microscopy and indirect assessment of both callose synthase and β-(1,3)-glucanase enzyme activities. Some of these methods are laborious, time consuming, not callose-specific, biased and require high technical skills. Here, we describe a method of callose quantification based on Sandwich Enzyme-Linked Immunosorbent Assay (S-ELISA). Tissue culture-derived banana plantlets were inoculated with Xanthomonas campestris pv. musacearum (Xcm) bacteria as a biotic stress factor inducing callose production. Banana leaf, pseudostem and corm tissue samples were collected at 14 days post-inoculation (dpi) for callose quantification. Callose levels were significantly different in banana tissues of Xcm-inoculated and control groups except in the pseudostems of both banana genotypes. The method described here could be applied for the quantification of callose in different plant species with satisfactory level of specificity to callose, and reproducibility. Additionally, the use of 96-well plate makes this method suitable for high throughput callose quantification studies with minimal sampling and analysis biases. We provide step-by-step detailed descriptions of the method.
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