Enzyme-linked Immunosorbent Assay Kit Research Articles

Development and Validation of an Indirect and Blocking ELISA for the Serological Diagnosis of African Swine Fever

African swine fever (ASF) is an economically devastating viral disease of pigs caused by the ASF virus (ASFV). The rapid global spread of ASF has increased the demand for ASF diagnostics to be readily available and accessible. No commercial ASF enzyme-linked immunosorbent assay (ELISA) kits are manufactured and licensed in North America. Here, we report the development of two serological diagnostic assays, a blocking ELISA (bELISA) based on ASFV glycoprotein p54 and an indirect ELISA (iELISA) based on ASFV glycoproteins p54 and p72. The assays showed high sensitivity and specificity and detected anti-ASFV antibodies in serum samples from experimentally infected animals as early as 8 days post-infection. The two assays were produced commercially (AsurDx™ bELISA and iELISA) and subjected to extensive validation. Based on data from a set of characterized reference sera, the prototype commercial assays, while maintaining 100.00% specificity, showed 97.67% (AsurDx™ bELISA) and 83.72% (AsurDx™ iELISA) sensitivity. Both prototype assays detected anti-ASFV antibodies in serum samples collected from pigs experimentally infected with multiple ASFV strains and field samples collected from sick, recovering, and vaccinated animals. The two commercially available assays can be used in routine ASF diagnostics, serological surveys, and for evaluating serological responses to ASF vaccine candidates.

Determination of the Level of von Willebrand Factor, ADAMTS13, and Ratio of ADAMTS13:von Willebrand Factor in Sickle Cell Disease Patients

Abstract Background: Sickle cell anemia (SCA) is a hypercoagulable state characterized by a significant alteration in hemostatic parameters which may predispose an increased risk of vaso-occlusive crisis (VOC). Sickle cell disease (SCD) is the most common genetic disorder in sub-Saharan Africa. Nigeria bears a high disease burden with an estimated prevalence of 1%–3% of its population being affected by the disease. The study seeks to determine the role of von Willebrand factor (VWF), ADAMTS13, and the ratio of ADAMTS13:VWF antigen in the pathogenesis of VOC. Objective: The objective of this study is to evaluate the level of VWF, ADAMTS13, and their ratio in SCD subjects in Calabar and to determine their role in the pathogenesis of VOC. Methodology: This is a comparative study carried out at the University of Calabar Teaching Hospital (UCTH), Calabar. Sixty SCA patients were evaluated in VOC and steady states as well as five parented healthy controls. VWF: Ag and ADAMTS13:Ag were evaluated using Assaypro enzyme-linked immunosorbent assay kits with Lot nos. 01751728 and 04222167R, respectively. Data were analyzed by IBM SPSS Chicago software version 21. The study was approved by the UCTH Institution Ethical Review Board. Results: The mean ages of the SCA subjects and controls were 23.5 ± 7.2 years and 26.5 ± 5.6 years, respectively (P = 0.706). There were 23 (38.3%) males in the SCA group and 21 (42.0%) females in the controls. There was no significant difference in their sex distribution (P = 0.063). The mean (standard deviation [SD]) of VWF in VOC, steady state, and controls were 2.52 ± 0.34, 1.34 ± 0.23, and 1.41 ± 0.23 IU/mL, respectively. The differences in mean were significantly higher in VOC state (P = 0.003). The mean ± SD of ADAMTS13 in VOC, steady state, and controls were 0.61 ± 0.10, 0.44 ± 0.06, and 0.62 ± 0.10 μg/L, respectively. ADAMTS13 levels did not differ significantly across the groups (P = 0.270). Similarly, there was no significant difference between ADAMTS13:VWF ratios across the groups (P = 0.318). Conclusion: VWF level is elevated in VOC state and thus may be implicated in the pathogenesis of VOC. ADAMTS13 and the ratio of ADAMTS13:VWF are not significantly affected in VOC.

Diagnosing Autoimmune Bullous Diseases—An Indian Perspective

Abstract Introduction: Autoimmune bullous diseases (AIBDs) are a group of illnesses characterized by autoantibodies targeting adhesion molecules in the skin and mucosa. Accurate diagnosis of the specific subtype of AIBD is crucial for effective management and predicting prognosis, especially in cases with an increased risk of malignancy. However, differentiating between subtypes can be challenging due to overlapping symptoms. Overview of diagnostic tests: Direct immunofluorescence microscopy (DIF) detects in vivo bound antibodies in perilesional tissue biopsies and provides details about the probable site of autoantibody deposition within the skin/mucosae, immunoglobulin type, and pattern of antibody deposition. Indirect immunofluorescence (IIF) microscopy with organ substrate is a minimally invasive serological test that detects circulating autoantibodies. Enzyme-linked immunosorbent assay (ELISA) quantifies serum autoantibodies against specific autoantigens. Quantitative ELISA is useful for diagnosis, monitoring therapy, and assessing disease activity. Commercially available ELISA kits, including the multi-variant ones, can detect antibodies associated with AIBDs. BIOCHIP is a technique based on IIF that offers a sensitive and specific diagnostic alternative to ELISA. It uses microarrays with multiple antigenic substrates to simultaneously screen common AIBDs. The BIOCHIP slides contain different substrates, allowing the identification of multiple types of autoantibodies in a single test. Indian context: While these diagnostic tests offer valuable insights into target antigens, antibody patterns, and disease subtypes, it is important to note that the availability of these tests is limited in most centers across India. This limitation can be attributed to factors such as the relatively higher cost of these investigations, challenges related to the stability of immuno-reactants, and a shortage of trained personnel capable of performing such tests. Conclusion: This review discusses the diagnosis of AIBDs based on resources available in India, as of today. It also provides with practically applicable diagnostic algorithms for pragmatic diagnosis of AIBDs in Indian scenario.

Review of domestic test systems for detecting current or past SARS‑CoV‑2 infection using enzyme‑linked immunosorbent assay

Introduction. Rapid and reliable assessment of circulating antibody levels to SARS-CoV-2 remains the current challenge for monitoring and preventing the spread of COVID-19. As the risk of emergence and dissemination of new viral strains continues to grow, epidemiologic research is also becomes increasingly important. The enzyme-linked immunosorbent assay (ELISA) is useful to assess the vaccination effectiveness and the need for revaccination, the proportion of post-disease and vaccinated individuals, and to monitor population immunity. This article provides an overview of the local ELISA kits currently available for the detection of antibodies against SARS-CoV-2 viral proteins, with a comparison to the certified kit by Mediagnost Ltd. (Germany).The aim. Laboratory testing of seven domestic reagent kits for the determination of antibodies to SARS-CoV-2 viral proteins by ELISA in comparison with the certified test system “Anti-SARS-CoV-2 ELISA E 111-IVD” (Mediagnost Ltd., Reutlingen, Germany).Materials and methods. The study included 80 serum samples from conditionally healthy donors: experimental group (n=70, serum samples collected after 2019) and control group (n=10, serum samples collected before 2018). The experimental group is comprised of non-infected and post-disease individuals; a proportion of those were vaccinated against COVID-19.Results and conclusions. The data obtained indicate high reproducibility, sensitivity (100 %), and specificity for all domestic ELISA kits in detecting both post-disease (90–95 %) and vaccinated volunteers (100 %). The main quality criteria of the obtained results were comparable to those for the certified test system, “Anti-SARS-CoV-2 ELISA E 111-IVD” (Mediagnost Ltd., Germany). All kits detecting IgG to the receptor-binding domain (RBD) and to the S-glycoprotein were effective for the reliable assessment of antibody production after the vaccination. Kits for the detection of antibodies to the RBD and/or viral nucleocapsid are useful for identifying unvaccinated post-disease individuals.

An NLR family member X1 mutation (p.Arg707Cys) suppresses hepatitis B virus infection in hepatocytes and favors the interaction of retinoic acid-inducible gene 1 with mitochondrial antiviral signaling protein.

NLR family member X1 (NLRX1) is an important member of the NOD-like receptor (NLR) family and plays unique roles in immune system regulation. Patients with hepatitis B virus (HBV) infection are more likely to have the NLRX1 mutation p.Arg707Cys than healthy individuals. It has been reported that NLRX1 increases the infection rate of HBV in HepG2 cells expressing sodium taurocholate cotransporting polypeptide (NTCP). However, the role of NLRX1 mutation (p.Arg707Cys) in hepatitis remains unclear. We constructed Huh7 cells that stably overexpressed NTCP, using LV003 lentivirus. First, wild-type (WT) and mutant (MT) NLRX1 overexpression plasmids were constructed. The MT plasmid contained a point mutation at position 707 of the WT overexpression plasmid. Then, Huh7-NTCP cells were transfected with the WT or MT NLRX1 overexpression plasmid, and subsequent NLRX1 expression was analyzed using real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. HBV RNA levels were determined using RT-qPCR. HBsAg and HBcAg levels were confirmed immunohistochemically. Interferon alpha (IFN-α), interleukin 6 (IL-6), and type I interferon beta (IFN-β) levels were determined using enzyme-linked immunosorbent assay kits. p-p65, p-interferon regulatory factor (IRF) 3, and p-IRF7 expression levels were examined using western blot. The interaction of NLRX1 and retinoic acid-inducible gene (RIG)-1/mitochondrial antiviral signaling (MAVS) protein was confirmed by coimmunoprecipitation. The interaction of NLRX1 with IFN-α, IL-6, or IFN-β was analyzed by dual luciferase reporter gene assay. The levels of HBV RNA, HBsAg, and HBcAg in infected cells transfected with the WT NLRX1 or MT NLRX1 expression plasmid were higher than those in the untransfected control group; and these levels were lower in the cells transfected with MT NLRX1 than in those transfected with WT NLRX1. The levels of IFN-α, IFN-β, IL-6, p-p65, p-IRF3, and p-IRF7 were lower in cells transfected with WT NLRX1 or MT NLRX1 than in control cells. The levels of IFN-β, p-p65, p-IRF3, and p-IRF7 were higher in cells transfected with MT NLRX1 than in those transfected with WT NLRX1. Moreover, NLRX1 competitively inhibited RIG1 binding to MAVS, but the mutation in MT NLRX1 reduced this inhibitory effect. In addition, NLRX1 decreased the promoter activity of IFN-α, IFN-β, and IL-6. Our findings revealed that NLRX1 is a regulatory factor that inhibits the anti-HBV ability of hepatocytes and that the mutation p.Arg707Cys in NLRX1 suppresses HBV infection and activates the IFN/nuclear factor κB pathway.

α-Linolenic acid promotes testosterone synthesis by improving mitochondrial function in primary rooster Leydig cells

The present study aimed to investigate the direct effects of α-Linolenic acid (ALA) on the in vitro production of testosterone and the expression of key enzymes and proteins related to steroidogenesis in Leydig cells of roosters. MethodsPurified primary Leydig cells isolated from 65-week-old roosters were purified and treated with different concentrations of ALA treatments: (0 μm/L [control], solvent control group (DMSO), 20 μM/L, 40 μM/L, and 80 μM/L) and cell counting-8 (CCK-8) for cell viability assay, Enzyme-linked immunosorbent assay (ELISA) kit for the determination of testosterone in cell supernatants, quantitative (real-time) PCR, and analysis of activities of antioxidants catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA), evaluation of mitochondrial membrane potential, pro- and anti-apoptotic proteins/genes Bcl-2, Bcl-2-associated X protein (Bax), apoptosis-inducing factor (AIF) were done respectively. ResultsOur results showed that ALA significantly increased testosterone secretion in primary rooster Leydig cells (P < 0.05), and 40 μM/L is the optimal dose. Leydig cells supplemented with ALA (20, 40, 80 μM) increased the expression of key enzymes and proteins 3β-hydroxysteroid dehydrogenase (3β-HSD), steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc) concerning steroidogenesis, enhanced antioxidant capability, improved mitochondrial biogenesis, and markedly improved the mitochondrial membrane potential (P < 0.05). Furthermore, the expression of the apoptosis-suppressive gene Bcl-2 was significantly increased, but Bax and AIF expression was decreased in the ALA group compared to that in the control group (P < 0.05). ConclusionALA promoted testosterone production, enhanced steroidogenic enzyme expression, improved mitochondrial function, and antioxidant capacity, and reduced apoptosis in primary rooster Leydig cells, with 40 μM/L identified as the optimal concentration.

Prognostic value of serum amyloid A protein as a biomarker in the diagnosis of 2019 novel coronavirus disease (COVID-19)

BackgroundThe current study aimed to evaluate the prognostic value of serum amyloid A protein )SAA( protein as a biomarker in diagnosing 2019 novel coronavirus disease )COVID-19( infection. MethodsThe study was conducted on 123 patients with definitive COVID-19 infection referred to Shahid Beheshti and Sina hospitals in Hamedan province, Iran. Five-milliliter blood samples were taken from all included patients and serum was isolated using a centrifuge at 10,000 rpm for 10 minutes. Laboratory tests were conducted, including c-reactive protein (CRP), Erythrocyte Sedimentation Rate (ESR), potassium level, sodium blood test, platelets (PLT), complete blood count (CBC), lymphocyte count, and neutrophil count. The SAA enzyme-linked immunosorbent assay (ELISA) Kit was applied to measure the SAA level in serum samples. Results123 patients included 73 males and 50 females, age ±50. Sixty-six (53.7%) patients had negative CRP while 80 (65%) patients had normal ESR. Potassium levels were not normal among 111 (94.9%) patients. Seventy-seven (63.1%) patients had normal CBC, while 108 (87.8%) patients had neutrophils above the normal range. 94 (97.9%) patients over the age of 50 were positive for SAA. In terms of gender, men were the most frequent patients with SAA. There was a statistically significant relationship between the serum level of SAA and outcomes of patients with COVID-19 (p = 0.0001). 94% of patients with SAA ≤50 were recovered from COVID-19 infection. The sensitivity rate of SAA compared to polymerase chain reaction (PCR) and computed tomography scan (CT scan) tests was 93% and 99%, respectively. Moreover, the accuracy of SAA compared to PCR and CT scan tests was 52% and 96%, respectively. ConclusionResults indicate the SAA is a sensitive, but not specific biomarker in the early detection of COVID-19. The quantitative levels of SAA can be useful in predicting treatment outcomes among patients with COVID-19.

An antibody nanopore-enabled microsensor for detection of osteoprotegerin

This paper introduces an antibody-based nanopore thin film sensor for detecting osteoprotegerin (OPG) in buffer solutions and serum samples, offering significant improvements over current commercial enzyme-linked immunosorbent assay kits in terms of cost, specificity, and selectivity. Capable of detecting OPG concentrations as low as 0.25 pg ml−1—far below the limits of traditional lab equipment—this sensor requires only about 1 μl of serum for effective measurement. The utilization of reference sensors helps reduce non-specific binding, enhancing the sensor’s accuracy. Its affordability and operational simplicity make it ideal for point-of-care monitoring of OPG levels in real-time patient care.

Astaxanthin improves assisted reproductive technology outcomes in poor ovarian responders through alleviating oxidative stress, inflammation, and apoptosis: a randomized clinical trial

BackgroundPoor ovarian response (POR) to controlled ovarian stimulation (COS) remains challenging, especially in advanced-age women with diminished ovarian reserve, resulting in low live birth rates. Many patients prefer to conceive with their eggs, underscoring the need for improved treatments. This study explores astaxanthin potential as a COS adjuvant to improve ovarian response and assisted reproductive technology (ART) outcomes, considering its impact on oxidative stress (OS), inflammation, and apoptosis, which are key factors in POR.MethodsIn this randomized, triple-blind, placebo-controlled trial, 60 infertile POR patients from POSEIDON Group 4 (the poorest prognosis category, age > 35 and poor ovarian reserve (anti-müllerian hormone < 1.2 ng/ml or antral follicle count < 5) undergoing intracytoplasmic sperm injection were enrolled. Patients were assigned to receive either 12 mg/day AST or placebo for eight weeks. All patients underwent a gonadotropin-releasing hormone antagonist regimen for COS. ART outcomes were compared between groups. Blood serum and follicular fluid (FF) were analyzed for OS markers (superoxide dismutase [SOD], total antioxidant capacity [TAC], and malondialdehyde [MDA]), and pro-inflammatory cytokines (interleukin-6 [IL-6], interleukin-8 [IL-8], and vascular endothelial growth factor [VEGF]) via enzyme-linked immunosorbent assay kits, and cell-free DNA [cfDNA] (apoptotic marker) via ALU quantitative polymerase chain reaction.ResultsAfter the intervention, the AST group exhibited a significant elevation in serum (P = 0.013) and TAC (P = 0.030), accompanied by a significant reduction in serum MDA (P = 0.005). No significant differences between AST and placebo groups were observed in OS markers in FF. AST group showed significant reductions in the serum IL-6 (P < 0.001), IL-8 (P = 0.001), and VEGF (P = 0.002) levels following AST therapy. In the AST group, FF levels of IL-6 (P = 0 < 001), IL-8 (P = 0.036), VEGF (P = 0.006), and cfDNA (P < 0.001) were significantly lower than in the placebo group. Between-group comparisons showed significant differences in the alterations of serum SOD (P = 0.027), IL-6 (P < 0.001), and IL-8 (P = 0.035) levels between AST and placebo groups. The AST group showed significant increases in the number of retrieved oocytes (P = 0.003), MII oocytes (P = 0.004), frozen embryos (P = 0.037), and high-quality embryos (P = 0.014) compared to the placebo group.ConclusionAST shows promise as a COS adjuvant therapy, potentially enhancing some ART outcomes in POR through alleviating OS, inflammation, and apoptosis.Trial registrationClinical trial registration number: IRCT20230223057510N1, URL: https://irct.behdasht.gov.ir/trial/68870, registration date: 2023 March 16.Graphical

Novel Insights Into The Role Of Serum Interleukin-39 In Patients With Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that has a genetic predisposition and a complex pathogenesis in which the involvement of various cytokines has been shown. In the current study, serum interleukin-39 (IL-39) levels from 99 patients with SLE and 33 healthy control subjects who attended the Baghdad Teaching Hospital Rheumatology Unit were examined. Patients were divided into 3 subgroups according to disease status: inactive (n = 33), active moderate (n = 33), and active severe (n = 33). This topic has not yet been explored, so the significance of IL-39 as a biomarker for SLE was evaluated. Cytokine levels were measured using enzyme-linked immunosorbent assay kits. Full medical histories, body mass index, gender, and clinical disease activity, the latter evaluated using the SLE disease activity index, were documented. Laboratory disease parameters, including anti-dsDNA antibodies, C3 and C4 levels, erythrocyte sedimentation rate, and C-reactive protein titres, were measured. The mean age of patients was 33.92 ± 0.91 years. The IL-39 level was higher in patients (13.70 ± 0.35 ng/l) than in controls (10.67 ± 0.19 ng/l, p &lt; 0.01). The mean of IL-39 levels was highest in patients with active severe SLE (17.42 ± 0.48 ng/l) and then became incrementally lower with reducing disease severity, i.e., active moderate, 13.34 ± 0.23 ng/l; inactive, 10.93 ± 0.24 ng/l. Serum IL-39 showed good validity for the diagnosis of SLE. With a cut-off value ≥ 10.25 ng/l and an area under the curve of 0.79, diagnostic sensitivity, accuracy, and specificity were 79.98%, 51.5%, and 71.97%, respectively. In conclusion, serum IL-39 levels were significantly higher in patients with SLE than in healthy controls and were correlated with disease activity. This interleukin may be useful in predicting disease severity.

Human β-Defensin 2 as a Link between EBV Infection and Multiple Sclerosis

Multiple sclerosis (MS) is the result of a complex interaction between environmental factors and genetic predisposition. Epstein-Barr virus (EBV) is a significant environmental risk factor associated with MS. Human- beta defensin 2 (HβD-2) is a crucial innate immune response proteint that as an inflammatory marker protects humans from pathogens invading the body like viruses. Therefore, a case-control study was conducted on 90 subjected (48 patients with MS and 42 healthy controls) to examine the role of HβD-2 in EBV viral load infection and MS patients. An enzyme-linked immunosorbentassay kit was used to measure HβD-2 levels in serum. The viral load of EBV was determined using a real-time PCR. The findings revealed that median HβD-2 levels in patients were significantly lower than in controls. (96.62 vs.124.8 ng/mL; p &lt; 0.01). According to the receiver operating characteristic curve analysis, HβD-2 is good predictor for MS disease (area under the curve=0.739; p &lt; 0.001). EBV frequency was lower in MS patients compared to the controls, although difference was not statistically significant (25 vs. 38.1%; p = 0.181).On other hand, the median (EBV) load was substantially greater in patients than in healthy controls ( 8.55 vs. 1.4 DNA copy/100 cells). This study concluded that HβD-2 levels showed no significant differences in each age group, sex, Expanded Disability Status Scale (EDSS) of multiple sclerosis patients, rather showed an inverse significant correlation (correlation coefficient = - 0.432) with EBV load, which protect against human viral infections.

Cardiac Myosin-binding Protein-C as a Biomarker in the Early Diagnosis of Acute Coronary Syndrome and Differentiation of Its Types

Abstract Background: Coronary heart disease (CHD) is primarily caused by atherosclerotic lesions within the intima of coronary arteries and acute coronary syndrome (ACS) is the main acute clinical manifestation of CHD. The ACS is manifested in one of three subtypes and it is the leading cause of mortality worldwide. The three subtypes of ACS include: acute myocardial infarction (MI) with the electrocardiogram (ECG) presenting ST-segment elevation (STEMI), MI with no ST-segment elevation on ECG (NSTEMI), and the third subtype is unstable angina (UA). The early diagnosis of is important in decreasing morbidity and mortality in ACS patients. Objective: To investigate the role of serum level of cardiac myosin-binding protein-C (cMyC), compared with high-sensitivity cardiac troponin-I (hsCTn-I) in the early diagnosis of ACS and differentiation of its subtypes. Materials and Methods: One hundred and twenty patients (72 males and 48 females), aged ≥30 years selected from those who were admitted to emergency department (ED) of Al-Yarmouk teaching hospital and diagnosed with ACS by cardiologists. The duration between the onset of chest pain and admission to ED should not exceed 3 h in any cohort. Apparenty healthy subjects as controls group for the study were recruited from those who had no current illness, particularly CHD, no other systemic disease and each had a normal ECG. For each study subject, cMybp-C and hsCT-I serum levels on admission were measured using the enzyme-linked immunosorbent assay kits. For each ACS patient, serum level of hsCTn-I level was measured 3 h after admission. Results: The comparison of cMybp-C levels among study groups revealed an overall significant difference and on paired comparison of study groups, the cMybp-C mean level was significantly higher in each ACS subgroup than in controls group (P &lt; 0.001), except in UA subgroup versus controls group. The cMybp-C levels showed a significant positive correlation with hsCT-I levels on admission in STEMI and NSTEMI subgroups but not in UA subgroup. The cMybp-C levels also showed a significant positive correlation with hscT-I levels 3 h after admission in STEMI and NSTEMI subgroups but not in UA subgroup. On receiver operating characteristic curve analysis, the cMybp-C level had a better diagnostic accuracy than hscTn-I on admission in differentiation of patients with “STEMI or NSTEMI” from those with UA or from controls. Conclusion: Serum cMybp-C mean level is significantly higher in ACS patients with STEMI and NSTEMI than in controls and the increase was more significant than hsCTn-I mean level on admission, so it could help the early diagnosis of ACS patients. The serum levels of cMybp-C also had a better diagnostic accuracy than hsCTn-I on admission in differentiation of ACS patients with STEMI or NSTEMI from those with UA or from controls.

Fast and label-free detection of procalcitonin in human serum for sepsis using a WTex-based extended-gate field-effect transistor biosensor

In this article, we present the first instance of depositing WTex-sensitive films with varying thicknesses (3, 4, and 5 nm) onto flexible polyimide substrates using radio-frequency sputtering. These films were used to create an extended-gate field-effect transistor (EGFET) for pH sensing and detecting procalcitonin (PCT) in the sera of patients with sepsis or bacterial infections. Among the films, the 4 nm WTex film exhibited high sensitivity (59.57 mV/pH), minimal hysteresis (∼0.8 mV), and a low drift rate (0.14 mV/h). Additionally, this WTex-based EGFET sensor retained a pH sensitivity of 59.2 mV/pH even after 180 days of operation and exhibited excellent mechanical flexibility, enduring 500 bending cycles without degradation. Moreover, PCT antibodies, activated using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinimide, were immobilized on the WTex film functionalized with 3-aminopropyl triethoxysilane. This effective immobilization enabled the specific binding of PCT antigens. The WTex-based EGFET biosensor demonstrated high sensitivity (18.12 mV/pCPCT) across a wide dynamic range (1 fg/mL to 1 μg/mL). Furthermore, the PCT concentrations in patient sera, whether from individuals with or without sepsis or bacterial infections, measured by our biosensor were comparable to results obtained using clinical enzyme-linked immunosorbent assay kits.

Assessment of the humoral immunity against diphtheria, tetanus, and hepatitis B among children with acute lymphocytic leukemia

Background: Approximately all systemic therapies for childhood affect the immune system. The behavior of the immune system in leukemia patients following chemotherapy is not yet clearly defined. The probability of vaccination failure and the need for revaccination remain challenging for these patients. Aim: To evaluate the humoral immunity against diphtheria, tetanus, and hepatitis B in children with acute lymphocytic leukemia (ALL) immediately and 6 months after chemotherapy. Materials and Methods: In the present prospective cohort study, 21 patients with ALL referred to Mofid Children’s Hospital were studied immediately and 6 months after chemotherapy. Serum samples were collected from patients, and the levels of immunoglobulins (IgG, IgM, IgE, and IgA) antibodies against diphtheria, tetanus, and hepatitis B were determined using specific enzymelinked immunosorbent assay kits. The obtained data were analyzed using Statistical Package for Social Sciences 21 software. Results: A total of 13 males and 8 females with an average age of 8.6 ± 2.5 years were included in the present study. Six months after chemotherapy, the mean level of IgG, IgM, IgE, and IgA displayed an increase of 563.1 units in IgG, 11 units in IgM, 11.3 units in IgE, and 5 units in IgA levels. Moreover, data revealed that 6 months after chemotherapy, the mean level of IgG antibodies displayed an increase of 7.09, 3.43, and 1.03 units against hepatitis B, diphtheria, and tetanus, respectively. A significant relationship was found between the antibody level against diphtheria and the age group of the patients (p = 0.003). Conclusion: Humoral immune status was boosted after 6 months of chemotherapy, though all patients had some extent of lasting immune dysfunction. We indicate that survivors of childhood cancer have ongoing humoral immunological defects and may remain at risk for infectious complications after completion of therapy. Relevance for Patients: The present study indicated that systemic therapies for pediatrics with leukemia affect the immune system. Pediatrics with leukemia may remain at risk for infectious complications after completion of therapy.

Co-frequency or contrary? The effects of Qiwei Baizhu Powder and its bioactive compounds on mucosa-associated microbiota of mice with antibiotic-associated diarrhea

Qiwei Baizhu Powder (QWBZP) has been proven effective in treating antibiotic-associated diarrhea (AAD), and the mechanism is associated with regulating the gut microbiota. However, the role of the bioactive compounds of QWBZP in regulating the gut microbiota is still unclear. In this study, 24 mice were divided into a normal control group (N), a model group (R), a QWBZP decoction group (TW), and a QWBZP-TG group (TG). AAD mouse models were established by mixed antibiotic administration. After modeling, mice in the TW group and TG group were treated with QWBZP decoction and QWBZP-TG, respectively. Mice in the N group and R group were gavaged with sterile water. 16S rRNA gene sequencing was used to investigate the changes of mucosa-associated microbiota (MAM) in the small intestine of mice. Moreover, the levels of diamine oxidase (DAO), D-Lactate, secretory immunoglobulin A (sIgA), interleukin 6 (IL-6), IL-10, and tumor necrosis factor-α (TNF-α) were detected using enzyme-linked immunosorbent assay (ELISA) kits. The results showed that QWBZP-TG significantly altered the diversity, structure, and abundance of MAM in the AAD mice. QWBZP-TG exerted a stronger suppression effect on Escherichia and Clostridium compared with QWBZP decoction. Meanwhile, QWBZP-TG downregulated the abundance of Lactobacillus, which elicited an opposite effect to QWBZP decoction. Prevotella was the signature bacteria that responded to the QWBZP-TG intervention. Furthermore, both QWBZP decoction and QWBZP-TG decreased the levels of DAO, D-Lactate, sIgA, IL-6, and TNF-α in the AAD mice. The role of glycosides is to help QWBZP ameliorate diarrhea symptoms by inhibiting the proliferation of diarrhea-associated bacteria, reducing inflammation and regulating immunity.

Serum Malondialdehyde-Modified Low-Density Lipoprotein Level May Be a Biomarker Associated with Aortic Stiffness Among Patients Undergoing Peritoneal Dialysis

Background: Serum malondialdehyde-oxidized low-density lipoprotein (MDA-oxLDL) is associated with atherosclerosis and increased risk of cardiovascular disease (CVD). Vascular calcification frequently occurs with arterial stiffness in patients on peritoneal dialysis (PD). This cross-sectional study aimed to elucidate the correlation between aortic stiffness and MDA-oxLDL levels in patients on PD. Methods: Overall, 92 patients on PD were included. The carotid–femoral pulse wave velocity (cfPWV) was evaluated using cuff-based volumetric displacement, and blood samples were obtained from all patients. Aortic stiffness was classified based on cfPWV values (&gt;10 m/s indicating aortic stiffness). Serum MDA-ox-LDL levels were quantified using commercial enzyme-linked immunosorbent assay kits. Results: In total, 33 (35.9%) patients were classified into the aortic stiffness group. Factors, including systolic blood pressure (SBP), serum triglyceride levels, C-reactive protein levels, age, weight, body mass index (BMI), waist circumference, MDA-oxLDL levels, and diabetes mellitus (DM) prevalence, were significantly higher in the aortic stiffness group. Multivariable logistic regression analysis revealed significant associations between aortic stiffness and MDA-oxLDL levels, BMI, and SBP. Furthermore, multivariable forward stepwise linear regression analysis revealed serum MDA-oxLDL levels as a significant independent predictor of cfPWV values. Conclusions: Serum MDA-oxLDL levels correlate positively with cfPWV values and may predict aortic stiffness among PD patients, highlighting its potential role in assessing CVD risk in this population.

Metformin Exhibits Anti-Inflammatory Effects by Regulating microRNA-451/CXCL16 and B Cell Leukemia/Lymphoma 2 in Patients With Osteoarthritis.

Osteoarthritis (OA) is the most common cause of chronic disability in joints among older individuals. The primary goal of OA treatment is pain relief to improve the quality of life. Inflammation and aging are involved in the pathogenesis of pain in OA. In this study, we evaluated the ability of metformin to regulate microRNAs, such as miR-451 and miR-15b, and their target proteins, CXCL16 and B cell leukemia/lymphoma 2 (BCL-2), involved in inflammation and apoptosis. In this double-blind placebo-controlled clinical trial, patients were randomly divided into two groups: one receiving metformin and the other receiving a placebo for four months (starting at 0.5 g/day for the first week, increasing to 1 g/day for the second week, and increasing to 1.5 g/day for the remaining period). In addition to evaluating the clinical response using the Knee Injury and Osteoarthritis Outcome Score questionnaire, miR-451 and miR-15b expression levels were detected using real-time polymerase chain reaction. The serum levels of CXCL16 and BCL-2 were evaluated using enzyme-linked immunosorbent assay kits before (time zero) and after treatment (month four). Metformin increased miR-451 expression levels simultaneously with pain reduction, whereas miR-15b expression did not change significantly after four months of treatment. Also, metformin decreased the serum levels of BCL-2 and CXCL16 in patients with OA. The effects of metformin in reducing pain can be attributed to many factors, including its anti-inflammatory and antiaging effects. Our findings suggest that metformin may reduce pain and inflammation in patients with OA through the regulation of miR-451/CXCL16 and BCL-2.

Poly (ADP-Ribose) Polymerase 1 Induces Cyclic GMP-AMP Synthase-stimulator of Interferon Genes Pathway Dysregulation to Promote Immune Escape of Colorectal Cancer Cells

Colorectal cancer (CRC) ranks third globally in cancer incidence and mortality, posing a significant human concern. Recent advancements in immunotherapy are noteworthy. This study explores immune modulation for CRC treatment. Initially targeting poly (ADP-ribose) polymerase 1 (PARP-1), a gene overexpressed in CRC tissues per The Cancer Genome Atlas, we examined its correlation with immune cell infiltration using the Tumor Immune Estimation Resource tool. Quantitative reverse transcription polymerase chain reaction assessed PARP-1 mRNA and inflammation-related gene expression in tumor tissues and cells. Assessing CD8+ T-cell proliferation and cytotoxicity towards HCT116 cells involved carboxyfluorescein diacetate succinimidyl ester and lactate dehydrogenase kits. Chemotaxis was gauged using a Transwell system in a CD8+ T-cell coculture setup, with immunofluorescence revealing cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING) levels in HCT116 cells. Enzyme-linked immunosorbent assay kits measured CD8+ T-cell cytokine secretion. The findings suggested that PARP-1 was overexpressed in CRC tissues and cells and this overexpression was positively correlated with Treg cell infiltration. Overexpression of PARP-1 could significantly reduce the proportion of cGAS and STING-positive cells in HCT116 cells, dampen the proliferation, tumor-killing capacity, and chemotaxis of CD8+ T cells, and inhibit the secretion of related cytokines. The introduction of STING agonists could reverse the effects caused by overexpressed PARP-1. In vivo experiments affirmed the independent anti-tumor effects of PARP-1 inhibitors and STING agonists, synergistically inhibiting tumor growth. Silencing PARP-1 in HCT116 cells potentially boosts CD8+ T-cell activity against these cells through the cGAS-STING pathway.

Hepatitis E Virus Infection in Patients with Systemic and Cutaneous Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by a multifactorial etiology in which genetic and environmental factors interplay. An exclusively cutaneous condition has been described and defined as cutaneous lupus erythematosus (CLE). In Italy, a nationwide blood donor survey found an overall HEV prevalence of 8.7%, with an interregional variation from 2.2% to 22.8%. In this study, we aimed to estimate HEV seroprevalence in a cohort of patients affected by SLE and CLE attending the Lupus Clinic, Sapienza University of Rome. Serum samples were tested for anti-HEV immunoglobulin Ig G and M antibodies using commercial enzyme-linked immunosorbent assay (ELISA) kits. Statistical analysis was performed. In total, 138 patients were enrolled, 92 (67%) affected by SLE and 46 by CLE. The prevalence of HEV infection was 23.9% in the CLE group and 7.6% in the SLE group. The anti-HEV+ prevalence was significantly more frequent in CLE. Some mechanisms may be linked to increased susceptibility to HEV such as a molecular mimicry associated with the CLE condition or with the skin compartment/skin self-antigens, as well as the involvement of the genetic background. Regarding the possible risk factors, no association was found, although, of note, the odds of HEV+ relative to contact with animals and to eating raw seafood were strongly higher than the unit in the CLE group.

Clinical study on the utility of allergy tests to detect IgE-mediated anaphylaxis after diclofenac etalhyaluronate administration.

This study was conducted to investigate factors involved in anaphylaxis related to diclofenac etalhyaluronate (DEH) [product name: Joyclu® (JCL)] (containing DEH and macrogol 400), which is used to treat patients with osteoarthritis. Patients with osteoarthritis were divided into two groups that had (experienced patients) or had not experienced anaphylactic symptoms after JCL administration (non-experienced patients). Five tests performed to assess factors related to anaphylaxis consisted of a skin prick test (SPT) as the primary endpoint and the other tests including basophil activation test, allergen-specific IgE (sIgE) tests using enzyme-linked immunosorbent assay or immunochromatographic kits, and genetic study were secondary endpoints. The SPT showed 4 (wheal)/7 (erythema) of 15 experienced patients and 0/3 of 19 non-experienced patients were positive for any of the test reagents containing DEH. The basophil activation test showed two experienced patients were positive for test reagents containing DEH. DEH- and diclofenac-sIgE were detected in 3 and 1 of 12 experienced patients, respectively. No clear results were shown in the other tests. DEH may be the main factor involved in the development of anaphylaxis. The SPT was more sensitive than the basophil activation and allergen-sIgE tests for identifying factors associated with anaphylaxis.