Journal of immunoassay & immunochemistry最新文献

The pattern recognition receptors (PRRs) trigger signaling cascades, such as nuclear factor kappa B (NF-κB) and interferon regulatory factors (IRFs). Rotavirus (RV) countermeasures against innate responses and understanding of these processes will improve our knowledge regarding immunopathogenesis of RV infection. In this study, we investigated the effect of RV RF strain on the important ISG candidate genes engaging in virus infections for which little information is known in RV RF strain. To this end, MA104 cells were mock/infected with RF followed by incubation in the presence or absence of IFN-α and the expression of MX1, OAS1, STAT1, ISG15, and ISG56 mRNA was analyzed by real-time PCR. All of ISGs' mRNAs showed higher expression levels in IFN I treated cells compared to virus-infected cells except for ISG56. Infecting the cells with RV and treatment with IFN type I led to overexpression of ISG56 compared to cells were either infected with the virus or only treated with IFN I. In conclusion, we showed that the RV RF strain efficiently blocks type I IFN-induced gene expression particularly ISG15, MX1, STAT, and OSA1 as antiviral proteins. Furthermore, viruses may use some ISGs such as ISG 56 to regulate IFN I signaling pathway, negatively.

Membrane proteins are difficult to be extracted and to be coated on the substrate of the immunoassay reaction chamber because of their hydrophobicity. Traditional method to prepare membrane protein sample requires many steps of protein extraction and purification that may lead to protein structure deformation and protein dysfunction. This work proposes a simple technique to prepare and immobilize the membrane protein suspended in an unprocessed crude cell lysate sample. Membrane fractions in crude cell lysate were incorporated with the large unilamellar vesicle (LUV) that was mainly composed of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) before coating in the polystyrene plate by passive adsorption technique. Immunofluorescence staining and the Enzyme-Linked Immunosorbent Assay (ELISA) examination of a strictly conformation-dependent integral membrane protein, Myelin Oligodendrocyte Glycoprotein (MOG), demonstrate that LUV incorporated cell lysate sample obviously promotes MOG protein immobilization in the microplate well. With LUV incorporation, the dose-response curve of the MOG transfected cell lysate coating plate can be 2-9 times differentiated from that of the untransfected cell lysate coating plate. The LUV incorporated MOG transfected cell lysate can be efficiently coated in the microplate without carbonate/bicarbonate coating buffer assistance.

Quantum dots have been widely used for biomedical applications like imaging, targeted drug delivery, and in-vitro diagnostics for better sensitivity. In-vitro diagnostic, lateral flow-based assay systems are gaining attention in the field of biomarker analysis mainly due to ease of test and quick availability of results. In the study, the potential of water-soluble carboxylic (-COOH) functionalized photoluminescent Cadmium Telluride Quantum Dots (CdTe) nanoparticles for lateral flow-based detection of N-terminal Natriuretic Peptide (NT-proBNP) biomarker (for heart failure) detection has been evaluated. Monoclonal antibodies were conjugated with COOH functionalized CdTe with EDC-NHS coupling chemistry, and conjugation was confirmed using FTIR. The CdTe nanoparticle exhibited an emission maximum at 715 nm when it is excited with 375 nm. The COOH functionalized CdTe showed an antigen concentration-dependent linearity in the lateral flow applications when the dye was prepared freshly and used. However, a relative reduction in CdTe quantum dot fluorescence intensity with time was observed. Factors such as low stability could be due to the quenching of the fluorescence of CdTe. This limits its commercial viability as an in-vitro diagnostic tool; thus, modifications of the quantum dots are required to have a stable preparation for its commercial potential for quantifications.

B-cell-activating factor (BAFF) is a crucial cytokine supporting survival and differentiation of B cells. Dysregulation of BAFF is involved in the pathogenesis of B-cell related autoimmune diseases including immune thrombocytopenia (ITP). The aim of this study was to evaluate the significance of BAFF expression in pediatric ITP patients. Eighty pediatric patients with ITP are subdivided in three groups. Group I included (32 patients) diagnosed with acute ITP less than 3 months, group II (48 patients) diagnosed with persistent ITP (from 3 to 12 months) and chronic ITP (more than 12 months) and group III 20 healthy controls. Complete blood picture, autoimmune profile, antiplatelet antibodies, coagulation profile, bone marrow examination, and RT-PCR were performed to detect the expression for BAF for all participants in this study. BAFF expression levels significantly increased in cases rather than in controls. BAFF Expression Value significantly increased in groups I & II (3.10 ± 1.99&3.29 ± 2.58) compared to controls (0.83 ± 0.45) as p < .001 for both. On the other hand, groups I & II were comparable in BAFF Expression Value (p = .470). BAFF expression increased in ITP patients, implying a function in the disease's pathogenesis.

There are limited data on inflammatory cytokines and chemokines; the humoral immune response; and main clinical laboratory parameters as indicators for disease severity and mortality in patients with critical and mild COVID-19 without comorbidities or immune-mediated diseases in Saudi Arabia. We determined the expression levels of major proinflammatory cytokines and chemokines; C-reactive protein (CRP); procalcitonin; SARS-CoV-2 IgM antibody and twenty-two clinical laboratory parameters and assessed their usefulness as indicators of disease severity and in-hospital death. Our results showed a significant increase in the expression levels of SARS-CoV-2 IgM antibody; IL1-β; IL-6; IL-8; TNF-α and CRP in critical COVID-19 patients; neutrophil count; urea; creatinine and troponin were also increased. The elevation of these biomarkers was significantly associated and positively correlated with in-hospital death in critical COVID-19 patients. Our results suggest that the levels of IL1-β; IL-6; IL-8; TNF-α; and CRP; neutrophil count; urea; creatinine; and troponin could be used to predict disease severity in COVID-19 patients without comorbidities or immune-mediated diseases. These inflammatory mediators could be used as predictive early biomarkers of COVID-19 disease deterioration; shock and death among COVID-19 patients without comorbidities or immune-mediated diseases.

Acute coronary syndrome (ACS) is defined as a range of conditions which the blood flow to the heart was reduced or stopped. This disorder is correlated to a systemic inflammatory response and some biochemical factors. Therefore, the aim of this study was investigations of serum C-reactive protein (CRP) and uric acid levels in ST-segment elevation myocardial infarction (STEMI) and non-ST-elevation ACS (NSTE ACS), as common subtypes of ACS. Patients with ACS (n = 140) were assessed with coronary arteriography and divided into STEMI and NSTE ACS groups. The serum levels of hs-CRP and uric acid were investigated using a routine clinical chemistry analyzer. Patients with STEMI showed a significant increase in uric acid level compared to those with NSTE ACS (P < .0001). Other data indicated that hs-CRP level in patients with STEMI was significantly higher than individuals with NSTE ACS (P < .0001). Modeling analysis revealed that the increased levels of acid uric and hs-CRP in patients with STEMI were independent of the effects of age, gender, background diseases, and familial history (P < .001). The current study provides further evidence to indicate that hs-CRP and uric acid may be considered as biofactors for comparing STEMI from NSTE ACS and determining disease outcome.

Lupus nephritis (LN) is the main manifestation of systemic Lupus Erythematosus (SLE). MicroRNAs (miRNAs) and autoantibodies could be suitable candidate biomarkers of LN. This study evaluates the expression of circulating miR-148a and miR-126 along with anti-dsDNA, anti-C1q, and anti-C3b autoantibodies in SLE patients with LN (SLE + LN). 30 women with SLE, 30 women with SLE + LN, and 25 women as healthy controls (HCs) were enrolled in this study. The plasma expression of selected miRNAs was evaluated by real-time PCR. The serum level of anti-dsDNA, C1q, and C3b antibodies was measured by the ELISA. The expression of miR-148a was significantly increased in SLE and SLE+LN groups compared with the control group. No significant difference was found in the expression of miR-126 among the groups. The frequency of autoantibodies was significantly higher in the SLE + LN group than SLE. The Higher levels of circulating miR-148a in the SLE samples compared with the HCs suggest that this miRNA could be a reliable biomarker for SLE patients (with or without LN). Also, autoantibodies against dsDNA, C1q, and, C3 could be used for the prediction of SLE nephritis, independently. However, further studies are needed to confirm these findings.

Identification of biomarkers is crucial in guiding the treatment decision and improving the future outcomes of DLBCL. The aim of the current study is to detect the biochemical and clinical impacts of miR-150 and miR-21 expression levels in DLBCL. Quantification of serum miR-150 and miR-21 expression levels by real-time PCR after micro-RNA extraction and RT-PCR. At a cutoff point of 2.3 for miR-21, the sensitivity, specificity, positive predictive, and negative predictive values for diagnosis of DLBCL were 98%, 90%, 90.7%, and 97.8%, respectively. At cut-off point (≤19.12) the sensitivity, specificity, the positive predictive and negative predictive values of miR-21 to discriminate stage IV vs stage II DLBCL patients were 68.42%, 80%, 86.7%%,and 57.1%, respectively. Serum miR-150 and serum miR-21 can be used as diagnostic markers for DLBCL patients, but miR-21 is more sensitive than miR-150. Serum miR-21 can be used as prognostic marker for DLBCL patients. It was more sensitive and more specific than miR-150.

Leptin, produced by adipocytes, regulates metabolism, hunger, and immune response. The inflammatory role of leptin has been linked to autoimmune diseases. To assess leptin gene polymorphism and serum level in alopecia areata and their relation to metabolic syndrome (MS). This case-control study was conducted on 100 alopecia areata patients (50 with MS and 50 without MS) and 50 age- and gender-matched controls. Leptin gene polymorphism and serum level were assessed through the use of PCR and ELISA, respectively. GG genotype was the highest in AA with MS (54%), lower in AA without MS (42%), and the lowest in controls (20%). G allele was more expressed in cases, than in controls (P < .001). The serum leptin level was the highest in AA with MS, lower in AA without MS, and the lowest in controls (P value = 0.001). Leptin level was significantly higher in GG polymorphism than AG and AA. Leptin gene polymorphism (GG genotype) and serum level appear to play a significant role in AA. Absent difference regarding leptin gene polymorphism and MS might indicate a separate inflammatory role of leptin or the future risk of MS development in AA patients.

Urinary bladder cancer incidence varies all over the world. Egypt displays high incidence rates. Molecular subtyping helps risk stratification and personalized treatment. Cancer-associated fibroblasts (CAFs) in the tumor microenvironment may provoke tumor-promotion or tumor suppression. Fibroblast activation protein (FAP) is a marker of CAFs, suggested to accelerate tumor progression in various cancers. In urothelial carcinoma, investigations regarding impact of FAP expression on prognosis are needed. This work aims to study impact of FAP expression in urothelial carcinoma and find its relation to CK 5/6 (basal) expressed and CK 20 (luminal) expressed immunohistochemical markers. This retrospective study included 70 urothelial carcinoma specimens. Immunohistochemistry was performed and results were analyzed. FAP was expressed in 67.1% of cases and showed significant association with advanced tumor stage, muscle invasion, mitoses in tumor cells and stratified groups; as 73.9% of FAP positive cases were of Ck5/6+/Ck20- (basal subtype). All studied parameters did not show significant association with patient's overall survival. In conclusion, FAP could have a role in modulating tumor microenvironment and promoting tumor invasion. FAP is correlated with basal subtype of urothelial carcinoma, which may be an indicator of tumor aggressiveness. FAP antagonists may be helpful in preventing tumor progression.