Background
Tall fescue (Festuca arundinacea Schreb., Lolium arundinaceum Schreb. Darbysh) and perennial ryegrass (Lolium perenne) are important cool‐season forage and amenity grasses that have a ...mutualistic association with an endophytic fungus. Endophytes confer insect and drought resistance to plants but can produce mammalian toxins. Novel endophytes that do not produce mammalian toxins have been introduced to elite cultivars for commercial production. Seed companies need to maintain adequate levels of novel endophytes within the elite forage cultivars. Endophyte detection is performed using immunochemical and molecular techniques because of their speed and reliability. Early detection in seedlings is essential to evaluate the viability of the endophyte within seed lots.
Methods
This research aimed to identify the earliest growth stage in which immunochemical and molecular methods can detect viable endophyte in seedlings of tall fescue cultivars BarOptima (e34), Texoma MaxQII (584), and Jesup MaxQ (542), as well as the perennial ryegrass cultivar Remington (NEA2).
Results
Immunochemical testing detected endophytes in seedlings 14 days after germination (DAG), but the detection rate increased until 42 DAG in some cultivars tested. The molecular marker Tef1exon detected endophytes at a lower rate than the immunochemical method at 28–42 DAG. However, there was insufficient DNA to detect endophytes in 14 DAG seedlings using markers.
Conclusions
We conclude that the most accurate detection of viable endophytes in seedlings was 42 DAG, at which sufficient and consistent endophyte colonization occurred.
Many forage grasses have a beneficial fungus living inside them. The presence of this fungus is a necessity to many producers. Knowing when and how to test for it has been a problem in the past. Here, we compare different methods of fungal testing and different time points that will help producers make testing decisions.
Scope
LTP‐syndrome is characterized by sensitization (IgE) to multiple non‐specific lipid transfer proteins (nsLTPs) with a variable clinical outcome. The treatment is primarily based on offending ...food avoidance. However, the determination of Pru p 3‐specific IgE is currently the main diagnostic tool to assess sensitization to nsLTPs. Herein, the study evaluates improvement of LTP‐syndrome diagnosis and clinical management using a new IgE multiplex‐immunoblot assay with a high diversity of food nsLTPs.
Methods and results
An EUROLINE‐LTP strip with 28 recombinant nsLTPs from 18 allergenic sources is designed. In total the study investigates 38 patients with LTP‐syndrome and compares results from the nsLTPs (LTP‐strip) with the respective food extracts of Prick‐by‐prick (PbP) testing. The agreement exceeds 70% for most nsLTPs, e.g., Pru p 3 (100%), Mal d 3 (97%), Pru av 3 (89%), Pha v 3 isoforms (87%/84%), Ara h 9 (82%), Cor a 8 (82%), and Jug r 3 (82%). The functionality and allergenic relevance of nine recombinant nsLTPs are proven by Basophil activation testing (BAT).
Conclusions
The new IgE multiplex‐immunoblot nsLTP assay shows a good diagnostic performance allowing culprit food assessment. Negative results from LTP‐strip may indicate potentially tolerable foods, improving diet intervention and patients’ quality of life.
The multiparameter‐immunoblot assay containing a high diversity of plant‐food‐nsLTPs shows a good diagnostic performance compared to Prick‐by‐Prick, representing an opportunity to improve the LTP‐syndrome allergological work‐up and clinical management. True positive tests allow culprit food assessment while true negatives may provide potentially tolerable foods, improving diet intervention and patient quality of life.
Rapid and simple serological assays for characterizing antibody responses are important in the current COVID-19 pandemic caused by SARS-CoV-2. Multiplex immunoblot (IB) assays termed COVID-19 IB ...assays were developed for detecting IgG and IgM antibodies to SARS-CoV-2 virus proteins in COVID-19 patients.
Recombinant nucleocapsid protein and the S1, S2 and receptor binding domain (RBD) of the spike protein of SARS-CoV-2 were used as target antigens in the COVID-19 IBs. Specificity of the IB assay was established with 231 sera from persons with allergy, unrelated viral infections, autoimmune conditions and suspected tick-borne diseases, and 32 goat antisera to human influenza proteins. IgG and IgM COVID-19 IBs assays were performed on 84 sera obtained at different times after a positive RT-qPCR test from 37 COVID-19 patients with mild symptoms.
Criteria for determining overall IgG and IgM antibody positivity using the four SARS-CoV-2 proteins were developed by optimizing specificity and sensitivity in the COVID-19 IgG and IgM IB assays. The estimated sensitivities and specificities of the COVID-19 IgG and IgM IBs for IgG and IgM antibodies individually or for either IgG or IgM antibodies meet the US recommendations for laboratory serological diagnostic tests. The proportion of IgM-positive sera from the COVID-19 patients following an RT-qPCR positive test was maximal at 83% before 10 days and decreased to 0% after 100 days, while the proportions of IgG-positive sera tended to plateau between days 11 and 65 at 78-100% and fall to 44% after 100 days. Detection of either IgG or IgM antibodies was better than IgG or IgM alone for assessing seroconversion in COVID-19. Both IgG and IgM antibodies detected RBD less frequently than S1, S2 and N proteins.
The multiplex COVID-19 IB assays offer many advantages for simultaneously evaluating antibody responses to different SARS-CoV-2 proteins in COVID-19 patients.
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•A series of β-carboline-cinnamide conjugates was synthesized as cytotoxic agents.•Compound 7h displayed potent IC50 of 0.70 µM in HCT-15, Entinostat IC50 of 3.87 µM.•Immunoblot ...analysis showed that the compound 7h is class I selective HDAC inhibitor.•Molecular docking revealed that 7h bind nicely within active pocket of the HDAC 2.•7h can be promising lead for development of potentially inhibiting HDAC inhibitors.
The effect of β-carboline motif as cap for HDAC inhibitors containing cinnamic acid as linker and benzamides as zinc binding group was examined in this study. A series of β-carboline-cinnamide conjugates have been synthesized and evaluated for their HDAC inhibitory activity and in vitro cytotoxicity against different human cancer cell lines. Almost all the compounds exhibited superior HDAC inhibitory activity than the standard drug Entinostat for in vitro enzymatic assay. Among the tested compounds, 7h displayed a noteworthy potency with an IC50 value of 0.70 ± 0.15 µM against HCT-15 cell line when compared to the standard drug Entinostat (IC50 of 3.87 ± 0.62 µM). The traditional apoptosis assays such as nuclear morphological alterations, AO/EB, DAPI, and Annexin-V/PI staining revealed the antiproliferative activity of 7h while depolarization of mitochondrial membrane potential by JC-1 was observed in dose-dependent manner. Cell cycle analysis also unveiled the typical accumulation of cells in G2M phase and sub-G1/S phase arrest. In addition, immunoblot analysis for compound 7h on HCT-15 indicated selective inhibition of the protein expression of class I HDAC 2 and 3 isoforms. Molecular docking analysis of compound 7h revealed that it can prominent binding with the active pocket of the HDAC 2. These finding suggest that the compound 7h can be a promising lead candidate for further investigation in the development of novel anti-cancer drug potentially inhibiting HDACs.
Following discontinuation of the recombinant immunoblot assay (RIBA), the only available supplementary test for the detection of hepatitis C virus (HCV) is the nucleic acid amplification test (NAAT). ...However, the NAAT does not adequately detect past HCV. Consequently, it is hard to distinguish between past HCV infection and biological false positivity with an anti-HCV result alone. We assessed the diagnostic performance of two immunoassays: the ARCHITECT anti-HCV chemiluminescent microparticle immunoassay (CMIA; Abbott Diagnostics, Wiesbaden, Germany) and the Access HCV Ab PLUS chemiluminescent immunoassay (CIA; Bio-Rad, Marnes-la-Coquette, France). We also explored an optimized algorithm to determine the anti-HCV results.
We tested 126,919 patients and 44,556 individuals who underwent a medical checkup. RIBA and NAAT were conducted for samples that tested anti-HCV-positive using CMIA and CIA. We assessed the optimal signal-to-cutoff (S/CO) ratio in HCV-positive samples.
In total, 1,035 blood samples tested anti-HCV-positive. Of these, RIBA was positive in 512, indeterminate in 160, and negative in 363 samples. One hundred sixty-five samples were NAAT-positive. Diagnostic sensitivity and positive predictive value (PPV) were 96.7% and 52.1%, respectively, for CMIA, and 94.7% and 72.3%, respectively, for CIA. The optimal S/CO ratio was 5.2 for CMIA and 2.6 for CIA at 95% PPV. In total, 286 samples tested positive in CMIA and 444 in CIA, while 443 samples tested positive in both assays.
It is hard to determine anti-HCV positivity based on the S/CO ratio alone. However, this study elucidated the role of the S/CO ratio by using the NAAT and RIBA.
To evaluate the clinical application value of electrochemiluminescence immunoassay analyzer (ECLIA) and chemiluminescent magnetic microparticle immunoassay (CMIA) in the detection of Treponema ...pallidum (TP).A total of 1225 patients in Peking University Third Hospital was enrolled from June 2014 to October 2014. ECLIA and CMIA were applied to detect the serum anti-TP. The positive rate was analyzed. RIBA was taken as a golden standard to evaluate the sensitivity, the specificity, the positive predictive value, the negative predictive value, and the accuracy of ECLIA and CMIA. A correlation analysis between 2 assays was conducted, and that between assay and RIBA. We also evaluate the clinical value of TPPA in the detection of T pallidum.The positive rate of CMIA and ECLIA is 10.63% and 9.89%, respectively, showing no statistically significant difference (P > .05). For CMIA, ECLIA, and TPPA, the sensitivity is 99.16%, 99.16%, and 99.16%, the specificity is 98.99%, 99.82%, and 100%, the positive predictive value is 91.47%, 98.33%, and 100%, the negative predictive value is 99.91%, 99.91%, and 99.91%, the coincidence rate is 99.01% (Kappa = 0.895), 99.75% (Kappa = 0.997), and 99.92% (Kappa = 0.998), respectively.The result shows high correlation between ECLIA and CMIA. Both have high sensitivity and specificity and can be used as screening tests for the diagnosis of T pallidum in common condition.
Background
Hepatitis C virus antibody (anti‐HCV) test had been approved as a preliminary screening test for HCV infection. Light‐initiated chemiluminescent assay (LiCA) was a homogenous method. We ...aimed to assess the clinical diagnostic performance of LiCA and compare it with that of chemiluminescence immunoassay (CLIA) which was widely used in clinical laboratories.
Methods
A total of 10 772 patients from the Peking University Third Hospital were enrolled. The serum samples were detected on the ChIVD LiCA500 and Abbott Architect i2000SR platforms. Recombinant immunoblot assay (RIBA) and HCV RNA assay were used for confirmation.
Results
The negative agreement rate between ChIVD LiCA anti‐HCV assay and Abbott Architect anti‐HCV assay was 99.91%, the positive agreement rate was 37.31%, the total agreement rate was 98.74%, and the kappa coefficient (κ) was 0.519. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of ChIVD LiCA anti‐HCV assay were 96.39%, 99.95%, 89.58%, and 99.97%, respectively, which were superior to those of Abbott Architect anti‐HCV assay (93.98%, 99.25%, 51.90%, and 99.95%, respectively).
Conclusion
ChIVD LiCA anti‐HCV assay was a highly sensitive, specific homogenous method with good diagnostic performance, and was applicable for the routine screening of HCV infection in clinical laboratories.
The measurement of antibodies against hepatitis C virus (HCV) is important to screen HCV infection. The aim of this study is to investigate the reliability of 2 commercially available anti-HCV ...antibody kits used in routine laboratory testing in Egypt. One thousand nine hundred and thirty-one serum samples were analyzed using 2 anti-HCV test systems: Cobas e 411® Elecsys Anti-HCVII and Vidas® Anti-HCV Biomerieux. Discrepant samples were tested using the recombinant immunoblot assay Innogenetics® INNO-LIA HCV Score. Overall agreement of the 2 tests was 94%. Following discrepant sample testing by LIA, sensitivity and specificity using Vidas were 94% and 99%, respectively, while those for Cobas were 97% and 96%, respectively. This study demonstrates superior specificity by Vidas and higher sensitivity by Cobas. Both methods are suitable for laboratory and/or blood screening programs. The concomitant use of a supplementary or confirmatory assay is necessary to compare the accuracy of HCV serological assays.
Background/Aim
To investigate the frequency and clinical relevance of an extended autoantibody profile in patients with systemic sclerosis (SSc).
Materials and Methods
In this cross‐sectional study, ...serum from 100 consecutive patients was subjected to indirect immunofluorescence (IIF) (HEp‐20‐10/primate liver mosaic) and Systemic Sclerosis Profile by EUROIMMUN to evaluate anti‐nuclear antibodies (ANA) and autoantibodies against 13 different autoantibodies in patients with SSc less than 3 years.
Results
Ninety‐three of 100 patients were positive for ANA by IIF. Fifty‐three patients showed single positivity, 26 anti‐topoisomerase antibodies (anti‐Scl70 ab), 16 anticentromere antibodies (ACAs), six anti‐RNA polymerase III antibodies (anti‐RNAPIII ab), one anti‐Ku antibody, one anti‐PM/Scl100 antibody, two anti‐PM/Scl75 antibodies, one anti‐Ro52 antibody, whereas 32 patients had multiple autoantibody positivities. Among classic SSc‐specific autoantibodies, anti‐Scl70 and anti‐RNAPIII abs showed the highest cooccurrence (n = 4). One patient was simultaneously positive for anti‐RNAPIII ab and ACA, and one was positive for ACA and anti‐Scl70 ab. The clinical features were not statistically different between single and multiple autoantibody‐positivity for classic SSc‐specific autoantibodies (ACA, anti‐Scl70 ab, and anti‐RNAPIII ab), except for digital ulcer in the multiantibody positive ACA group (p = .019).
Conclusion
Based on our results, coexpression of autoantibodies is not uncommon in SSc patients. Although autoantibodies specific to SSc in early disease show generally known clinical features, it remains to be investigated how the coexpression of autoantibodies will affect clinical presentation.
The coexpression of autoantibodies is not uncommon in systemic sclerosis (SSc) patients. Although SSc‐specific autoantibodies generally show known clinical features, the clinical presentation of the coexpression in specific and nonspecific autoantibody positivity continues to be necessary.
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