Serum Enzyme-linked Immunosorbent Assay Research Articles

Hamster neogenin, a host-cell protein contained in a respiratory syncytial virus candidate vaccine, induces antibody responses in rabbits but not in clinical trial participants

ABSTRACT A recombinant respiratory syncytial virus (RSV) fusion glycoprotein candidate vaccine (RSV-PreF) manufactured in Chinese hamster ovary cells was developed for immunization of pregnant women, to protect newborns against RSV disease through trans-placental antibody transfer. Traces of a host-cell protein, hamster neogenin (haNEO1), were identified in purified RSV-PreF antigen material. Given the high amino-acid sequence homology between haNEO1 and human neogenin (huNEO1), there was a risk that potential vaccine-induced anti-neogenin immunity could affect huNEO1 function in mother or fetus. Anti-huNEO1 IgGs were measured by enzyme-linked immunosorbent assay in sera from rabbits and trial participants (Phase 1 and 2 trials enrolling 128 men and 500 non-pregnant women, respectively; NCT01905215/NCT02360475) collected after immunization with RSV-PreF formulations containing different antigen doses with/without aluminum-hydroxide adjuvant. In rabbits, four injections administered at 14-day intervals induced huNEO1-specific IgG responses in an antigen-dose- and adjuvant-dependent manner, which plateaued in the highest-dose groups after three injections. In humans, no vaccination-induced anti-huNEO1 IgG responses were detected upon a single immunization, as the values in vaccine and control groups fluctuated around pre-vaccination levels up to 90/360 days post-vaccination. A minority of participants had anti-huNEO1 levels ≥ assay cutoff before vaccination, which did not increase post-vaccination. Thus, despite detecting vaccine-induced huNEO1-specific responses in rabbits, we found no evidence that the candidate vaccine had induced anti-huNEO1 immunity in human adults. The antigen purification process was nevertheless optimized, and haNEO1-reduced vaccines were used in a subsequent Phase 2 trial enrolling 400 non-pregnant women (NCT02956837), in which again no vaccine-induced anti-huNEO1 responses were detected.

IL36 indicating good prognosis in human Hepatocellular Carcinoma.

Background and Aim: Hepatocellular carcinoma (HCC) is the leading cause of cancer death in men before the age of 60 years in China. Interleukin (IL)36 played important roles in antitumor immune responses, but its role in HCC is still unknown. We aimed to explore the correlation between IL36 and prognosis of HCC patients.Methods: The expression of IL36 was measured by Immunohistochemistry (IHC), serum Enzyme-linked Immunosorbent Assay (ELISA) and flow cytometry (FCM). Chi-square test was performed to analyze the relationship between IL36 expression and clinical parameters of HCC patients. The correlation between IL36 expression and prognosis of HCC patients was evaluated by Kaplan-Meier method and Cox regression analysis.Results: The IL36 expression in HCC tumor samples was lower than that in paired peri-tumor samples; the analyses suggested that there was no correlation between IL36 expression and age, gender, and tumor size, but tight relationship between IL36 expression and liver cirrhosis, metastasis and some other clinical parameters. The results of Kaplan-Meier analysis indicated positive expression of IL36 could induce high survival rate of patients. The detection of IL36 with ELISA suggested that expression of IL36 in serum was the highest in patients of HCC, other than the chronic hepatitis patients and the healthy. The result of FCM suggested the expression of IL36 was higher in CD4+ T cells than other immune cells.Conclusions: There is a close relationship between the expression of IL36 and the prognosis of HCC, higher expression of IL36 suggested better prognosis and longer survival of HCC.

Results of enzyme-linked immunoassay of the blood serum in patients with infectious and inflammatory diseases of the anterior segment of the eye

To evaluate the clinical significance of the data obtained during enzyme-linked immunosorbent assay (ELISA) of the blood serum of patients with inflammatory diseases of the anterior eye segment compared to a group of healthy volunteers. A retrospective analysis of the results of serum ELISA of 200 patients with chronic keratoconjunctivitis and keratouveitis was performed using the solid-phase IFA method in order to detect the presence of antibodies to herpes simplex viruses of 1 and 2 types (HSV 1, 2), cytomegalovirus (CMV), Chlamydia trachomatis and Toxoplasma gondii. The control group consisted of 34 healthy volunteers with no signs of inflammation of the eye tissue. There were no significant differences in the frequency of detection of both «acute-phase» IgM and «chronic» IgG to HSV and CMV between patients of the main group and healthy volunteers (ρ≥0.05). The levels of IgG to Toxoplasma gondii in the study group were three times higher than in the control group (ρ≤0.05). A comparative analysis of the frequency of detection of antibodies to Chlamydia trachomatis showed statistically significant differences between the groups and more frequent detection of immunoglobulins classes A, G and M in patients with inflammatory eye diseases (ρ≤0.05). The absence of statistical differences with the control group in the frequency of detection of active HSV and CMV infections in patients with inflammatory diseases of the anterior eye segment when using ELISA allows to recommend the use of additional methods of examination such as polymerase chain reaction (PCR) and real-time PCR (RT-PCR) with identification of pathogen genetic material in the available biological secretions. The increase in IgG titers to toxoplasma in patients of the main group apparently indicates a cross-stimulation of antibody synthesis against the background of a chronic inflammatory process. Significant differences in the frequency of detection of all types of antibodies to chlamydiae suggests a significant etiological role of this pathogen in the development and maintenance of chronic inflammation in the anterior eye segment.

A case report of intraparenchymal neurocysticercosis in a postpartum female in Saudi Arabia

Neurocysticercosis is a neurologic disease caused by infection with larvae of Taenia solium. It is most commonly transmitted by consumption of pork, water contaminated with T. solium or poor hygiene habits. As only few cases of neurocysticercosis have been documented in Saudi Arabia, the authors report a case of neurocysticercosis in a young Indian female residing in Saudi Arabia who presented with generalized tonic–clonic seizures 6 days after a normal vaginal delivery. Her physical and laboratory investigations as well as chest X-ray and electroencephalogram were all normal. Computed tomography of the head revealed multiple calcified nodular lesions, and magnetic resonance imaging showed ring-enhancing lesion in the left frontoparietal area. Serum enzyme-linked immunosorbent assay (qualitative) was positive for immunoglobulin G antibodies for cysticercosis. A diagnosis of neurocysticercosis was made, and the patient was treated with dexamethasone and levetiracetam for 4 days before discharge. At the 3-month follow-up, the patient's condition had significantly improved, and her seizures had not recurred. This report recommends considering neurocysticercosis as a differential diagnosis in patients presenting with new-onset seizures, even if the symptoms do not initially indicate neurocysticercosis or if the patient resides in an area where the disease is rare.

Albumin-corrected serum calcium and serum parathyroid hormone-related polypeptide determined by enzyme-linked immunosorbent assay in cirrhotic patients with hepatocellular carcinoma \u2013 a pilot study

ContextParathyroid hormone-related peptide (PTHrP) is produced by many malignant tumors. It is responsible for most cases of hypercalcemia in patients with malignancy. Few published studies shed light on the relation between serum calcium levels and serum PTHrP levels in cirrhotic patients with hepatocellular carcinoma (HCC).AimThe aim of the current work was to evaluate serum PTHrP in cirrhotic patients with HCC and a possible correlation between serum PTHrP levels and albumin-corrected serum calcium levels in these patients.Patients and methodsThis is a cross-sectional study. The study included 35 cirrhotic patients with HCC (diagnosed depending upon α-fetoprotein and abdominal imaging studies). Data about their serum albumin and albumin-corrected serum calcium levels were collected. Sera of the studied patients were collected for determination of PTHrP levels by enzyme-linked immunosorbent assay (ELISA). Numerical data were summarized in the form of mean±SD. Strength of association between variables was tested using Pearson correlation coefficient.ResultsApproximately 8.6% of studied patients were hypercalcemic, and no statistically significant positive correlation was detected between serum PTHrP determined by ELISA and albumin-corrected serum calcium in these patients.ConclusionApproximately 8.6% of studied cirrhotic patients with HCC were hypercalcemic. NO statistically significant positive correlation was detected between serum PTHrP determined by ELISA and albumin-corrected serum calcium in these patients. Studies involving a larger number of patients could clarify the exact role of PTHrP in the development of hypercalcemia in cirrhotic patients with HCC.

Effect of herbal cake-partitioned moxibustion on Leptin/JAK2/STAT3 in lipid-lowering pathway of hyperlipidemia rabbits

Objective: To observe the lipid-lowering effect of different transdermal absorption enhancers applied to the herbal cake-partitioned moxibustion in hyperlipidemia model rabbits, and to explore the possible mechanism. Methods: Forty New-Zealand rabbits were randomly divided into 5 groups using the random number table method, with 8 rats in each group. Rabbits in the blank group were fed routinely with normal diet; rabbits in the other groups were fed with high-fat diet for 12 weeks to establish the hyperlipidemia model. Rabbits in the blank and the model groups were not treated. After the model was prepared, rabbits in the non-transdermal absorption enhancer group received herbal cake-partitioned moxibustion without transdermal absorption enhancer; rabbits in the laurocapram group and the borneol group received herbal cake-partitioned moxibustion with laurocapram or borneol respectively. After 4 weeks of treatment, serum was collected for enzyme-linked immunosorbent assay (ELISA), and the liver tissues were isolated for immunohistochemistry, quantitative polymerase chain reaction (qPCR) and Western-blotting (WB) detection. Results: Serum ELISA results showed that leptin was significantly decreased in the model group compared with the blank group (P 0.05). The qPCR results of rabbit liver tissues showed that the mRNA expressions of leptin, Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) in the model group were significantly lower than those in the blank group (all P<0.05); compared with the model group, the mRNA expressions of leptin, leptin receptor (LR), JAK2 and STAT3 in the non-transdermal absorption enhancer, the laurocapram and the borneol groups were significantly increased (all P<0.05); compared with the non-transdermal absorption enhancer group, the mRNA expressions of leptin, LR, JAK2 and STAT3 in the laurocapram and the borneol groups were significantly increased (all P<0.05); compared with the laurocapram group, the mRNA expressions of leptin, LR, JAK2 and STAT3 in the borneol group were significantly increased (P<0.05). The trend of immunohistochemistry and WB detection results was basically consistent with the qPCR assay results. The immunohistochemistry and WB detection results of phosphorylated JAK2 (phospho-JAK2) and phosphorylated STAT3 (phospho-STAT3) were basically consistent with those of JAK2 and STAT3. Conclusion: The molecular expression of Leptin/JAK2/STAT3 pathway in the hyperlipidemia model rabbits was decreased. The molecular expression of Leptin/JAK2/STAT3 pathway was significantly increased after the herbal cake-partitioned moxibustion. The application of laurocapram and borneol, as transdermal absorption enhancers, in the herbal cake-partitioned moxibustion could more obviously up-regulate the factors of the Leptin/JAK2/STAT3 lipid-regulating pathway than the herbal cake-partitioned moxibustion alone.

Association of Helicobacter pylori infection and vitamin B12 deficiency

H. pylori infection is strongly related with chronic gastritis of the stomach, which causes impairment in gastric acid and secretion of pepsin, and linked to male absorption of food-vitamin B12. The most important virulence genes accompanying stomach and intestine disease are (cag A and vac A). Aim of the study: To evaluate the rate of gastric ulcer infection with H. pylori and its detect CagA gene in H.pylori isolated from gastritis. Materials and Methods: A total of 200 biopsy patients aged from (10- ≥ 60) years were collected from Baqubah teaching hospitals. They were suffering from gastric upset and attended to endoscopic unit of department of medicine. Questioners including, sex, age, smoking, presence of cancer, and biopsies of gastric tissue were collected from the corpus or the ant rum or corpus and ant rum of the patient's stomach. Three biopsies were taken from each patient. Histopathologic study, gram staining, and rapid urease test working for each patients. Serology Test Serum Enzyme-Linked Immunosorbent Assay Testing (ELISA).

Galactose-deficient IgA1 in skin and serum from patients with skin-limited and systemic IgA vasculitis

IgA vasculitis (IgAV) encompasses a systemic form involving kidneys, gut, skin, or joints, and a skin-limited form. One characteristic feature of systemic IgAV is deposition of galactose-deficient IgA1 (GD-IgA1) in kidneys (as in IgA nephropathy). The relevance of GD-IgA1 for cutaneous vasculitis is unknown. We investigated whether GD-IgA1 is deposited perivascularly in systemic and also skin-limited IgAV and whether its serum levels differ between both forms. In a case-control study, deposition of GD-IgA1 was analyzed immunohistochemically by KM55 antibody in skin biopsy specimens from 12 patients with skin-limited IgAV and 4 with systemic IgAV. GD-IgA1 levels were compared by enzyme-linked immunosorbent assay in sera from 15 patients each with skin-limited and systemic IgAV and from 11 healthy individuals. All biopsy samples from systemic IgAV, and also from skin-limited IgAV, revealed perivascular GD-IgA1 deposition. The average GD-IgA1 concentration in serum was significantly higher in systemic IgAV than in skin-limited IgAV, despite overlap between the groups. Although high GD-IgA1 levels may be predictive of systemic IgAV, patient numbers were too low to determine cutoff values for systemic versus skin-limited IgAV. Perivascular GD-IgA1 deposition is a prerequisite for systemic and skin-limited IgAV; however, high GD-IgA1 levels in some patients with systemic IgAV suggest a dose-dependent effect of GD-IgA1 in IgAV.

Changes of myelin basic protein expression in the white matter of young rats with sepsis

Objective To investigate the relationship between endotoxin infection and demyelinating lesions of the central nervous system, and to provide evidence for clinical diagnosis and treatment. Methods Sixty-four healthy Wistar rats(18 days old), weighting 30~40 g, were randomly divided into two groups: control group and lipopolysaccharide(LPS) group.LPS group was intraperitoneally injected with 5 mg/kg LPS to prepare rat sepsis model.The control group received intraperitoneal injection of saline of equal volume.All surving animals were anesthetized at the set time 2 h, 4 h, 6 h and 12 h after administration(8 rats at each time point), and craniotomy after taking blood from the heart.Myelin injury was staining by luxol fast blue.Immunohistochemical staining and Western blot were used to detect the expression of myelin basic protein(MBP) in rat brain at different time points.The expression of the MBP protein in serum detected by enzyme-linked immunosorbent assay(ELISA). Results Observed under light microscope after luxol fast blue staining, the myelin sheath of the white matter in the brain was disordered, sparse and stained lightly from 6 to 12 hours in LPS group.Western blot analysis and immunohistochemical test results showed that the expression of MBP protein in the LPS group was decreased compared with the control group at 2 h, 4 h and 6 h after LPS administration(P 0.05). The serum ELISA results showed that the serum MBP levels in the LPS group increased at 2 h, 4 h, 6 h and 12 h after LPS administration(P<0.05). Conclusion In rat sepsis model, MBP protein expression in the white matter decreases, serum MBP protein content increases, and the decreasing of MBP protein expression is associated with matter myelin injury. Key words: Lipopolysaccharide; Sepsis; Demyelination; Myelin basic protein

In vitro modulatory effect of dehydroepiandrosterone sulfate on apoptosis and expression of apoptosis-related genes in patients with systemic lupus erythematosus.

Systemic lupus erythematosus‌ (SLE) is an autoimmune disease characterized by hyperactive B cells that produce various autoantibodies. Sex hormones have been documented to influence the development of SLE, in which women with SLE have low plasma level of dehydroepiandrosterone sulfate (DHEAS). A strong conclusion about the effect of DHEAS on apoptosis in SLE patients has not been provided. The aim of this study was to assess apoptotic effects of DHEAS on peripheral blood lymphocytes (PBLs) from SLE patients. Twenty SLE patients and 20 age- and sex-matched healthy controls were included into this study. Concentration of DHEAS was measured using enzyme-linked immunosorbent assay in serum from all participants. Freshly isolated PBLs from each individual were treated with 7.5-µmol of DHEAS for 24 hr in cell culture medium to assess the effect of DHEAS on apoptosis using fluorescein isothiocyante-conjugated annexin V and propidium iodide. The messenger RNA (mRNA) expression level of apoptosis-related genes (Fas, Fas-L, Bcl-2, and Bax) in PBLs was measured using real-time PCR before and after treating with DHEAS. Level of DHEAS was low in SLE patients compared with healthy controls (p < 0.05). After treating with DHEAS, the percentage of apoptotic cells in SLE patients was decreased in comparison with healthy controls. DHEAS treatment increased the mRNA expression level of Bcl-2 in PBLs from SLE patients. DHEAS reduced the apoptosis rate in PBLs from SLE patients and may decrease the load of autoantigens. Therefore, DHEAS might be considered as a therapeutic tool in SLE patients.

Importance of Serum Adiponectin Levels in The Prediction And Ethiopathogenesis of Intrauterine Growth Restriction

Objective: Our aim in this study is to determine the possibility that patients with IUGR (intrauterine growth retardation)will be able to look at the adenonetin levels in the early stages of pregnancy and to have IUGR. In this respect, earlydiagnosis, treatment and follow-up are aimed to reduce morbidity and mortality due to IUGRMaterial Method: The study was carried out between January and March 2010. Zekai Tahir Burak Women’s HealthEducation and Research Hospital 28-40. with 89 patients aged 18-40 years. Groups 44 IUGR patients and 45 normalpregnant patients were divided into two groups. To examine the serum adiponectin level, an average of 3 cc was takenin the gel biochemistry tubes and then centrifuged at 4000 10 minutes. The resulting serum samples were collectedfor the ELISA kit and kept at -80 C until the day the patients were collected. In the study, adiponectin levels weremeasured by serum ELISA (enzyme linked immunosorbent assay) method.Results: In adiponectin evaluation, mean value was 105.90 19.02 pg / ml for the IUGR group, 65.96 16.5 pg / mlfor the control group and 1.6 times higher for the IUGR group( p < 0.001)Conclusion: In this study, the increased adiponectin levels of IUGR patients were significantly higher than those ofIUGR patients, suggesting that pregnancies of IUGR patients can be measured high in the first weeks and if IUGR ishigh in those weeks, IUGR can be predicted.

Using a panel of multiple tumor-associated antigens to enhance the autoantibody detection in the immunodiagnosis of ovarian cancer.

Ovarian cancer (OC) is a major malignancy affecting a large population over the world, and a biomarker that holds diagnostic potential is of critical importance. Recently, autoantibodies have been indicated as biomarkers in multiple cancer research. The current study was designed to explore the practice of using autoantibodies in diagnostic settings by the enzyme-linked immunosorbent assay of sera with a panel of tumor-associated antigens (TAAs). A panel of 12 TAAs was selected to detect the corresponding autoantibodies in sera sampled from 132 OC patients as case group and 147 normal healthy individuals as the control group. The diagnostic potential of this panel was evaluated by conventional evaluation, receiver operating characteristic (ROC) curve analyses, and classification tree analysis. When the cutoff values were set as mean ± 2 SD for normal healthy individuals, the positive rates of antibodies to any single TAA were less than 20% both in OC and in normal healthy individuals. In a parallel screening approach, a panel ofnine TAAs (p53, C-myc, p90, p62, AHSG, 14-3-3zeta, RalA, Koc, and p16), obtained optimal diagnostic performance in OC with the sensitivity of 61.4% at the 85.0% specificity. In addition, when the nine TAAs were combined with CA125, the sensitivity and specificity were improved to 94.7% and 78.2%, respectively. The ROC curve analyses showed that only the area under the receiver operating characteristic curves (AUCs) of antibodies against C-myc, Koc, and RalA was beyond 0.6, which were 0.732, 0.668, and 0.665, respectively. The AUC of the combination was up to 0.914 (P < 0.05). Decision tree analysis showed that C-myc, HCC1.3, RalA, and CA125 held high potential in the detection of OC. The panel ofnine TAAs also identified 78.8% of OC patients who had normal CA125 levels in their serum samples, indicating that elevated CA125 and anti-TAA antibodies appeared to be independent but supplementary biomarkers for diagnosing OC. In summary, the current study further supports that a customized TAA panel can serve as a promising and powerful tool for immunodiagnosis of OC and may be particularly useful in patients with normal CA125 levels.

Trypanosoma cruzi infection in naturally infected dogs from an endemic region of Cundinamarca, Colombia.

The seropositivity and risk factors for Trypanosoma cruzi infection in dogs from a municipality of Cundinamarca, a central state of Colombia were studied. A total of 356 client-owned dogs from urban, peri-urban and rural areas of La Mesa municipality, (Cundinamarca, Colombia) were randomly selected. Blood samples were collected by venipuncture. Anti-T. cruzi antibodies were determined using the enzyme-linked immunosorbent assay (ELISA) method. Reactive ELISA sera were processed by indirect immunofluorescence to confirm the presence of anti-T. cruzi antibodies. Chi-square tests were conducted for statistical analysis. Serologic tests for T. cruzi infection showed a prevalence of 29.49% (105/356), the rural area show a highest T. cruzi infection pattern in comparison with the other zone locations. Two triatomine species were found through the study: Panstrongylus geniculatus (53.4%) and Rhodnius colombiensis (46.6%). The prevalence of positive vectors for parasite was of 52.1% (38/73). Additionally, a very close relation between triatomine bugs and dogs in the rural zone (1:3.1) was observed. These results are the first report of natural infection by T. cruzi in domestic dogs in La Mesa municipality. In conclusion, the presence of anti-T. cruzi antibodies in dogs in this area suggest vector transmission. There is a need for active surveillance programs throughout the La Mesa municipality and vector control strategies should also be implemented.

New insight into the diagnostic cut-off value of serum anti-ToxocaraIgG for ocular toxocariasis in uveitis patients

Uveitis is one of the commonest causes of vision loss worldwide and its exact etiology is still not clarified in most patients. The current study is a trial to assess the efficacy of serum anti-Toxocara immunoglobulin G (IgG) by enzyme-linked immunosorbent assay (ELISA) as a diagnostic tool for ocular toxocariasis (OT) and to detect OT prevalence and the associated ocular manifestations in sera of patients with uveitis. One hundred and twelve patients (62 females and 50 males) with uveitis were diagnosed by ophthalmologists, radiologists and rheumatologists according to ocular manifestations, laboratory and radiological investigations. Serum anti-Toxocara IgG titers were determined by ELISA in sera of all patients. Our results revealed that OT is highly associated with intermediate and posterior uveitis. Children and young adult females, especially those residing in rural areas, complained mainly of diminution of vision in the left eye, with strabismus and leukocoria. At a cut-off value of 0.258, the sensitivity and specificity of IgG ELISA were 93.3% and 100%, respectively. In conclusion, at a novel cut-off value of 0.258 the serum anti-Toxocara IgG ELISA is predicted to be a diagnostic tool for OT regarding sensitivity and specificity. Also, it has potential importance in the interpretation and differential diagnosis of OT. Thus, serum anti-Toxocara IgG ELISA should be a routine test for screening of suspected cases.

Detection of toxoplasma-specific immunoglobulin G in human sera: performance comparison of in house Dot-ELISA with ECLIA and ELISA.

In the current study, performance of electrochemiluminescence immunoassay (ECLIA) in detection of anti-toxoplasma IgG in human sera was compared with that of enzyme-linked immunosorbent assay (ELISA). Furthermore, performance of an in house Dot-ELISA in detection of anti-toxoplasma IgG was compared with that of ECLIA and ELISA. In total, 219 human sera were tested to detect anti-toxoplasma IgG using Dynex DS2® and Roche Cobas® e411 Automated Analyzers. Discordant results rechecked using immunofluorescence assay (IFA). Then, sera were used in an in house Dot-ELISA to assess toxoplasma-specific IgG. Of the 219 samples, two samples were found undetermined using ECLIA but reactive using ELISA. Using IFA, the two sera were reported unreactive. Furthermore, two samples were found reactive using ECLIA and unreactive using ELISA. These samples were reported reactive using IFA. The overall agreement for the two former methods was 98% (rZ0.98.1; P< 0.001). The intrinsic parameters calculated for in house Dot-ELISA included sensitivity of 79.5, specificity of 78.2, and accuracy of 78.9%, compared to ECLIA and ELISA. Positive and negative predictive values included 82.9 and 74.2%, respectively. A 100% sensitivity was found in in house Dot-ELISA for highly reactive sera in ECLIA and ELISA. ECLIA is appropriate for the first-line serological screening tests and can replace ELISA due to high speed, sensitivity, and specificity, particularly in large laboratories. Dot-ELISA is a rapid, sensitive, specific, cost-effective, user-friendly, and field-portable technique and hence can be used for screening toxoplasmosis, especially in rural fields or less equipped laboratories.

Combined use of CEMIP and CA 19-9 enhances diagnostic accuracy for pancreatic cancer

Carbohydrate antigen (CA) 19-9 is the only diagnostic marker used in pancreatic cancer despite its limitations. Here, we aimed to identify the diagnostic role of CEMIP (also called KIAA1199) combined with CA 19-9 in patients with pancreatic cancer. A retrospective analysis of prospectively collected patient samples was performed to determine the benefit of diagnostic markers in the diagnosis of pancreatic cancer. We investigated CEMIP and CA 19-9 levels in 324 patients with pancreatic cancer and 49 normal controls using serum enzyme-linked immunosorbent assay. Median CA 19-9 and CEMIP levels were 410.5 U/ml (40.8–3342.5) and 0.67 ng/ml (0.40–1.08), respectively, in patients with pancreatic cancer. The AUROC for CA 19-9 and CEMIP were 0.847 (95% confidence interval [CI]: 0.806–0.888) and 0.760 (95% CI: 0.689–0.831), respectively. Combination of CA 19-9 with CEMIP showed markedly improved AUROC over CA 19-9 alone in pancreatic cancer diagnosis (0.94 vs. 0.89; P < 0.0001). CEMIP showed a diagnostic yield of 86.1% (68/79) in CA 19-9 negative pancreatic cancer. Combined use with CEMIP showed significantly improved diagnostic value compared with CA 19-9 alone in pancreatic cancer. Especially, CEMIP may be a complementary marker in pancreatic cancer patients with normal CA 19-9 levels.

Proteomic identification and characterization of hepatic glyoxalase 1 dysregulation in non-alcoholic fatty liver disease

BackgroundNon-alcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide. However, its molecular pathogenesis is incompletely characterized and clinical biomarkers remain scarce. The aims of these experiments were to identify and characterize liver protein alterations in an animal model of early, diet-related, liver injury and to assess novel candidate biomarkers in NAFLD patients.MethodsLiver membrane and cytosolic protein fractions from high fat fed apolipoprotein E knockout (ApoE−/−) animals were analyzed by quantitative proteomics, utilizing isobaric tags for relative and absolute quantitation (iTRAQ) combined with nano-liquid chromatography and tandem mass spectrometry (nLC-MS/MS). Differential protein expression was confirmed independently by immunoblotting and immunohistochemistry in both murine tissue and biopsies from paediatric NAFLD patients. Candidate biomarkers were analyzed by enzyme-linked immunosorbent assay in serum from adult NAFLD patients.ResultsThrough proteomic profiling, we identified decreased expression of hepatic glyoxalase 1 (GLO1) in a murine model. GLO1 protein expression was also found altered in tissue biopsies from paediatric NAFLD patients. In vitro experiments demonstrated that, in response to lipid loading in hepatocytes, GLO1 is first hyperacetylated then ubiquitinated and degraded, leading to an increase in reactive methylglyoxal. In a cohort of 59 biopsy-confirmed adult NAFLD patients, increased serum levels of the primary methylglyoxal-derived advanced glycation endproduct, hydroimidazolone (MG-H1) were significantly correlated with body mass index (r = 0.520, p < 0.0001).ConclusionCollectively these results demonstrate the dysregulation of GLO1 in NAFLD and implicate the acetylation-ubquitination degradation pathway as the functional mechanism. Further investigation of the role of GLO1 in the molecular pathogenesis of NAFLD is warranted.

False positive acetylcholine receptor antibodies in a case of unilateral chronic progressive external ophthalmoplegia: case report and review of literature

ABSTRACTMethods: We present a rare case with atypical presenting features of unilateral CPEO with a false positive Acetylcholine Receptor Antibody (AchRA) test resulting in diagnostic delay. We illustrate the unilateral nature of this case and demonstrate the caveats of performing myogenic ptosis correction in such patients. We also discuss the differential diagnosis of false positive AchRA, a test commonly performed in the investigation of ptosis.Results: A 34-year old female presented with a more than 3-year history of slowly-progressive, unilateral, right-sided restriction in eye movements and ptosis. Clinical examination showed EOM were grossly restricted in the right eye with a ptosis and normal in the left eye. Serum AchRA was positive on serum enzyme-linked immunosorbent assay (ELISA) however, following two months of oral pyridostigmine therapy there were no signs of clinical improvement. The initial serum sample sent was retested for AchRA by radio-immunoassay (RIA) which came back negative. Subsequently a muscle biopsy was requested which showed the presence of ragged red fibres.Conclusion: Unilateral ptosis and ophthalmoplegia is an unusual presentation for CPEO which characteristically produces bilateral symmetrical motility defects. In addition to Myasthenia Gravis elevated AchRA levels have been reported in other autoimmune conditions such as Primary biliary cirrhosis, Eaton Lambert syndrome and Graves’s ophthalmopathy. We also highlight the superiority of RIA versus ELISA in the detection of AchRA and illustrate the diagnostic challenge of investigating and managing myogenic ptosis in this complex cohort of patients.

Comparison of the serum and salivary antibodies to detect gastric Helicobacter pylori infection in Kashan (Iran)

Background and aimHelicobacter pylori (H. pylori) is an important and common contagious human pathogen which may cause peptic ulcer and also gastric cancer. The definite diagnosis of it is made through invasive tests. Recently, non-invasive tests including serologic tests of serum and saliva have been conducted for diagnosis of H. pylori infection. In this research, the diagnostic values of serum and salivary serology were compared together to use salivary anti-H. pylori test as an alternative method in the future.MethodsDuring this prospective case-control study on patients who were candidates for endoscopy and gastric biopsy from March 2015 to April 2016 in Shahid Beheshti hospital, Kashan, Iran, serum and salivary samples were obtained for measurement of Immunoglobulin G (IgG) antibody levels against H. pylori by enzyme-linked immunosorbent assay (ELISA). Histopathology was the gold standard test. Statistical analysis was performed by SPSS software version 16. Statistical tests included Kolmogorov-Smirnov, independent-samples t-test, Chi-square, Mann-Whitney U, Kruskal-Wallis, McNemar and correlation.ResultsOf 123 patients, sixty-one patients (49.6%) were H. pylori-positive according to histology. The median levels of anti-H. pylori antibodies in serum (p<0.001) and saliva (p<0.001) of H. pylori-positive cases were significantly higher than H. pylori-negative cases. Sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and accuracy of serologic tests in serum were 75%, 79%, 3.5, 0.3, 77% and for saliva were 85%, 82%, 4.7, 0.18, 84% respectively.ConclusionDiagnostic values of salivary ELISA are comparable to serum ELISA and can be used as an alternative modality for diagnosis of H. pylori infection.

PREPARATION AND EVALUATION OF CHEMICALLY INACTIVATED SALMONELLA ENTERITIDIS VACCINE IN CHICKENS

Objective: Salmonella enteritidis ghosts (SEGs) is a non-living empty bacterial cell envelopes which were generated using a different concentration of sodium hydroxide (NaOH) 6.4 mg/mL and evaluated as a vaccine candidate in specific pathogen-free (SPF) chicken. SEGs have been produced by chemical-mediated lysis and evaluated the potential efficacy of chemically induced SEG vaccine and its ability to induce protective immune responses against virulent S. enteritidis challenge in SPF chickens.Methods: SPF chickens were divided into three groups: Group A (non-vaccinated control), Group B (vaccinated with prepared vaccine), and Group C (vaccinated with commercial vaccine).Results: Vaccination of SPF chicken with SEGs induced higher immune responses before and after virulent challenge. SPF chicken vaccinated with SEGs showed increasing in serum enzyme-linked immunosorbent assay (ELISA) antibodies. During the vaccination period, Groups B and C showed higher serum antibody titer compared to Group A. The minimal inhibitory concentration (MIC) of NaOH was capable of inducing non-living SEGs, and it has successfully generated non-living SEGs by MIC of NaOH.Conclusion: It is a one-step process which means easy manufacturing and low production cost compared to protein E-mediated lysis method. Chemically induced SEG vaccine is a highly effective method for inducing protective immunity. This study strongly suggests that SEGs will be a permissive vaccine, as the method of inhibition of S. enteritidis was safe and cheaper than other methods, and it gave a good protection.