Journal of immunoassay & immunochemistry最新文献

Immunoassay tests are used in clinical and forensic toxicology laboratories to determine illicit drug use in biological samples. Therefore, this study aims to optimize the cutoff concentrations of DOA I Plus in the blood and compare the LC-MS/MS results. 680 authentic forensic whole blood specimens with Randox Evidence DOA I Plus array were screened for drug of abuse and confirmed by LC-MS/MS. Regarding the manufacturer recommended threshold values, 139 out of 680 authentic blood samples were positive for one or more analytes, while 541 were negative. Nearly all of the 139 positive blood samples confirmed by LC-MS/MS were true positive for one or more analytes, while 522 of the 541 negative blood samples were true negative. The overall sensitivity and the specificity were 87.8% and 99.6%, respectively. THC was considered in detail, and a receiver operator characteristic curve analysis was performed to determine the optimum cutoff for THC, as it accounts for 78% of all positive results according to the manufacturer's recommended thresholds. The optimal threshold value for THC was determined at a concentration of 23 ng/mL, while these values for other parameters were defined as recommended by the manufacturer.

Several diagnostic measures have been employed to precisely detect the SARS-CoV-2 viral infection using viral antigens, nucleic acids, and other serological approaches. The sensitivity and specificity of the serological tests remain a challenging need. Here, we describe the detection of human anti-SARS-CoV-2 IgG and IgM antibodies qualitatively through two optimized in-house ELISA and lateral flow immunoassay. Both approaches are based on the prokaryotic expression of 50 kDa SARS-CoV-2 recombinant nucleocapsid protein. This SARS-CoV-2rN-6×His was used either to coat ELISA plates or to be conjugated to gold nanoparticles followed by colorimetric detection of bound human IgG or IgM. In the LFA, we show the optimization of nanoparticle size, protein-binding capacity, membrane treatment, and finally testing the potential capacity of using either the optimized ELISA or LFA in detecting antibodies raised against viral infection. Assessment of both methods was carried out using human sera-positive and negative SARS-CoV-2 antibodies. The ELISA and LFA tests showed 86%, 96.5% sensitivity, 92%, 93.75% specificity, 97%, 98.2% PPV, and 64%, 88.2% NPV, respectively. In conclusion, both approaches were able to successfully detect human antibodies against SARS-CoV-2 nucleocapsid protein. The importance of both protocols cannot be overstated in the detection and diagnosis of viral infections, especially in developing countries.

Tetanus is an acute and often fatal infectious disease caused by Clostridium tetani. Tetanus toxin (TT) is responsible for spastic paralysis observed in tetanus. Anti-tetanus antibodies obtained from horses and humans are the most antitoxins used for tetanus treatment, although some clinical side effects and disadvantages have been reported in their application. The aim of this study is the production of anti-TT IgY and evaluation of its protective effects in a mouse model. Anti-TT IgY was purified from the egg yolk using PEG6000 precipitation and water dilution methods, and its purity was verified by SDS-PAGE. Finally, the potency of purified anti-TT IgY in neutralizing the lethal effects of TT was studied in vivo using a mouse model. PEG6000 precipitation method had better results. Animal studies showed that the purified IgY neutralized the toxic effects of 100 MLD of TT and multiple intravenous-dose injections of anti-TT IgY also had a continuous effect of TT neutralization. The purified anti-TT IgY was effective in neutralizing the lethal activity of TT in a mouse model. Our results suggested that IgY could be an alternative therapeutic source for the management of tetanus in the future.Abbreviations Anti-TT, Anti-tetanus toxin; ELISA, Enzyme-linked immunosorbent assay; IgY, Immunoglobulin Y; MLD, Minimum lethal dose; PBS, Phosphate buffer solution; PEG, Polyethylene glycol; SDS-PAGE, Sodium dodecyl sulfate polyacrylamide gel electrophoresis; TIG, Tetanus immune globulin; TT, Tetanus toxin; WD, Water dilution; RT, Room temperature.

Probiotics positively influence age-related macular degeneration (ARMD) given their propensity to attenuate oxidative and inflammatory stress. We addressed the impact of probiotics on metabolic profiles, clinical indices, inflammatory and oxidative stress parameters in ARMD patients. We performed a randomized, double-blind, placebo-controlled trial analyzing 57 subjects with ARMD aged between 50 and 85 years. Subjects were randomized into two groups, and received daily for 8 weeks either probiotic capsule or placebo. Fasting blood samples were obtained at baseline and after the 8-week intervention for the determination of metabolic profiles and oxidative stress biomarkers. After the 8-week intervention, compared with the placebo, probiotic supplementation significantly increased means HDL-cholesterol (Probiotic group: +3.86±4.42 vs. Placebo group: -0.55±4.93 mg/dL, P = .001), plasma total antioxidant capacity (TAC) (Probiotic group: +77.43±168.30 vs. Placebo group: -23.12±169.22 mmol/L, P = .02) and significantly decreased malondialdehyde (MDA) levels (Probiotic group: -0.18±0.46 vs. Placebo group: +0.18±0.25 µmol/L, P = .001). There was no significant effect of probiotic administration on other metabolic profiles and clinical symptoms. Overall, an eight-week probiotic administration among ARMD patients had beneficial effects on TAC, MDA and HDL-cholesterol levels; however, it did not affect clinical signs and other metabolic profiles.

Changes in the immune system participate in the pathogenesis and development of infectious diseases. Previous studies have indicated immune dysregulation in patients suffering from COVID-19 and mucormycosis. Therefore, this study investigated whether interleukin-27 (IL-27) and interleukin-32 (IL-32) levels may participate in the development and outcome of COVID-19 associated mucormycosis (CAM). The blood samples were obtained from 79 patients suffering from COVID-19 and mucormycosis and 25 healthy subjects. The serum samples were isolated from the whole blood and frequencies of some immune cells were measured by a cell counter. The levels of IL-27 and IL-32 were assessed by enzyme-linked immunosorbent assay. IL-27 and IL-32 levels were significantly lower in patients with COVID-19 and mucormycosis than healthy subjects (P < .05), although there was no significant difference in IL-27 between patients with COVID-19 and CAM. IL-27 level was significantly higher in severe COVID-19 survivors than dead cases (P < .01). Patients with CAM had significant increases in NLR compared to COVID-19 patients and healthy individuals (P < .0001-0.01). NLR was significantly associated with COVID-19 outcome (P < .05). Severe COVID-19 survivors had a significant reduction in NLR compared to non-survivors (P < .05). Changes in IL-27 and IL-32 levels may contribute to the pathogenesis of CAM. IL-27 may relate to the pathogenesis and outcomes of mucormycosis in COVID-19 patients.

The status of Infectious bursal disease (IBD) in indigenous chickens and backyard poultry in Rwanda has not been previously elucidated. This cross-sectional study was to determine the seroprevalence of infectious bursal disease in indigenous chickens and to identify the associated factors. The study was been done in three districts in the Eastern province of Rwanda where blood from 364 indigenous chickens were collected. ID Screen® IBD indirect enzyme-linked immunosorbent assay (ELISA) test was used to detect IBD antibodies in these birds. 145 questionnaires were also administered to poultry farmers to obtain information on biosecurity measures and associated factors to IBD outbreaks. The study revealed 48.4% (176/364) prevalence of the chicken with IBDV antibodies with statistical significance (P < .05) among/between location and age groups. The questionnaire revealed that there were other important associated factors which included chicken scavenging for seed as a source of food (59.3% of farmers reported), absence of routine vaccination (53.8%), live chickens are purchased from the open market with no information about IBD outbreaks and vaccination (30.0%), open disposal of dead chickens suspected of IBD (58.9%). IBD virus antibodies are present in indigenous chicken in Eastern Rwanda hence further investigation to better understand the epidemiology of IBD virus in indigenous chickens is desired and more research is needed to identify the role of indigenous chickens in the spread of IBD virus in Rwanda.

This study was designed to evaluate the immunohistochemical expression of programmed death ligand-1 (PD-L1) in tumor cells (TCs) and tumor-infiltrating immune cells (TICs) in hepatocellular carcinoma (HCC) and to correlate its expression with clinicopathological parameters. Seventy-two formalin-fixed paraffin-embedded blocks of HCC were collected. The data were collected from the patients' records. The blocks were stained with hematoxylin and eosin. Additionally, they were immunostained with PD-L1. Membranous staining was considered positive expression including the entire membrane or part of it ± cytoplasmic staining, and the percentage of total cancer cells ≥ 5% was evaluated as positive staining for TCs. The TICs were considered positive if they expressed membranous ± cytoplasmic staining of PD-L1 ≥ 1%. Of the total cases, 34.7% expressed PD-L1 positively in TCs and 15.3% expressed PD-L1 positively in TICs. Significant associations were observed between PD-L1 expression in TCs and tumor grade, capsular and/or vascular invasion, tumor stage, nodal metastasis, and the expression of PD-L1 in paracancerous tissue. The cases that positively expressed PD-L1 exhibited reduced overall survival (OS). PD-L1 was expressed in HCC TCs and TICs. Its expression in TCs was associated with higher HCC grades, advanced stages, capsular and/or vascular invasion, and nodal metastasis, and cases that expressed PD-L1 displayed reduced OS. Therefore, PD-L1 might serve as a poor prognostic indicator and a tumor immunotherapy target.

Psoriasis is characterized by cutaneous hyperproliferation, secondary to immune system dysregulation. Vitamin A regulates the immune response and sustains epithelial tissue hemostasis. The CYP1A1 gene, has many biological actions, including vitamin A metabolism. To evaluate CYP1A1 gene polymorphism and serum vitamin A level in patients with psoriasis vulgaris, a case-control study involving two groups was conducted: group 1 (45 patients with psoriasis vulgaris) served as the cased group and group 2 (45 healthy participants who were sex and age matched) acted as the control group. CYP1A1 (rs1048943) gene polymorphism and vitamin A serum level were assessed by TaqMan allelic discrimination (PCR) and ELISA, respectively. AG genotype was present only in cases (22.2%), while AA genotype was present in all controls (P=.001). Vitamin A levels were lower in cases than in controls (32.0 ± 7.41 vs. 46.2 ± 15.7 μg/ml, respectively) (P<.001). AG genotype was associated with a lower vitamin A level (P=.001). The detected genotype difference between psoriasis patients and controls, which was associated with a lower serum vitamin A level and was also lower in more severe cases, suggests a role of the CYP1A1 gene and vitamin A in disease pathogenesis and prognosis.

Alopecia areata (AA) is a disorder with several etiologies. The evidence suggests that the absolute copy number of mitochondrial deoxyribonucleic acid (mtDNA), as well as proportion of mutated mtDNA copies, determines disease onset. This study aims to quantify the relative index of the mtDNA copy number in patients with AA and healthy controls and correlate the results with the existing clinical ‎information. This case-control study included 50 patients with AA and 50 age- and sex-coordinated healthy persons as controls. The severity of AA was weighed using the Severity of Alopecia Tool and Kavak's classification. The relative index of the mtDNA copy number was measured by real-time qPCR. Significant statistical difference was observed between cases and controls regarding mean mtDNA copy number, p < .001. There was significant positive correlation with SALT score (p =  0.001). A cutoff value of >1.619 N/µL could significantly diagnose AA cases (p < .001), and a cutoff value of > 1.36 N/µL could discriminate mild AA cases from those with moderate AA (p =  0.007). The relative index of mtDNA copy number is significantly elevated in AA cases and could be helpful in diagnosing and evaluating AA severity.