Enzyme-linked Immunoassay Method Research Articles

CHARACTERISTICS OF CYTOKINE REGULATION AND CARTILAGINOUS TISSUE METABOLISM AT PRIMARY AND POST-TRAUMATIC OSTEOARTHRITIS

Cytokine system and cartilaginous tissue metabolism of 27 women affected by 0-1 stage of primary and 19 women affected by 0-1 stage of post-traumatic osteoarthritis (OA) have been studied as well as of 10 healthy persons of the control group. The enzyme-linked immunoassay method was used to determine the content of cartilage oligometric matrix protein (СOMP), cartilage glucoprotein (YKL-40), interleukins (ILs) 4 and 1β in blood serum, collagen fragments (CTX II) in urine. It was found that СOMP, CTX II, YKL-40 and IL-1β concentrations grew at more intensive rate in case of primary OA compared to post-traumatic OA. The diminution of correlation relationship strength in IL-1β serum concentrations is noted with the system level of cartilaginous tissue degradation products on the background of YKL-40 regulatory influence buildup which is more prominent with primary OA than it is with the post-traumatic one. Conclusions: early stages of primary OA are characterized by more prominent degenerative changes in the joint hyaline cartilage due to losses of type II collagen and hyperproduction of proinflammatory link cytokines with the background of YKL-40 regulatory influence buildup and reduction of the IL-1β influence. The changes of cartilaginous tissue metabolism with post-traumatic OA were characterized by preservation of dependence on IL-1β serum concentration with the background of reduction of YKL-40 influence.

The results of the diagnosis of giardiasis in children with the use of the method of microscopy, enzyme immunoassay, polymerase chain reaction.

Giardiasis in children remains an urgent problem, the importance of which is determined by the endemicity for many countries and regions, polymorphism of the clinic, which causes difficulties in clinical diagnosis of this pathology. The report presents the results of the diagnostic significance of methods of laboratory diagnostics giardiase: microscopy of native and stained with Lugol solution preparation fecal enzyme-linked immunoassay (ELISA) for the presence of antibodies of class IgM and IgG antibodies to the antigens of Giardia in the serum and antigen of G. Lamblia in feces, polymerase chain reaction (PCR) - detection of Giardia DNA in feces obtained during the examination of 160 patients with giardiasis children aged 3 to 14 years. The results of our study suggest that the most informative methods of diagnosis of giardiasis in children is coproscopy (specificity - 96.2%, sensitivity - 98.1%), PCR coprofiltrate. (specificity - 85%, sensitivity 82.5%), determination of antigen in feces by ELISA (specificity - 87,5%, sensitivity - 60%). With high specificity of the blood ELISA method (90%), the sensitivity of the method is only 27.5%. It should be recognized that the «gold standard» diagnosis of giardiasis remains microscopy of native and stained with a solution of Lugol preparation of feces with a three - time study at intervals of 2-3 days.

P6314Left- versus Bi-Ventricular Assist Device-mediated anti-HLA alloantibody induction in bridge-to-transplantation patients

Abstract Background Presence of anti-Human Leukocyte Antigen (anti-HLA) alloantibodies (allosensitization) in a heart transplantation candidate impedes the chance of finding a compatible transplant. Left Ventricular Assist Devices (LVADs) have been shown to induce allosensitization. Purpose The purpose of this study is to characterize anti-HLA allosensitization in adult patients receiving left (LVAD) versus biventricular- (BiVAD) assist devices as bridge to transplantation (BTT). Methods This study retrospectively assessed anti-HLA antibody induction in all adult patients who have received either LVAD or BiVAD as BTT at our institution. Anti-HLA alloantibody screening was performed before, and at multiple time-points after VAD implantation. Anti-HLA antibody detection was performed with a cytotoxic panel-reactive antibody (PRA) method until 2003, enzyme-linked immunoassay (ELISA) method from 2003 to 2005 and SAB assays on the Luminex platform from 2005 to 2017. Sensitization was defined either as PRA >10%, or as peak anti-HLA antibody mean fluorescence intensity (MFI) values of more than 1000. Baseline characteristics and sensitization in the two patient groups were evaluated using descriptive statistics. Results Between 2003 and 2018, 154 patients were placed on VAD support at our institution as BTT. Sensitization data were available for 130 patients. Fifty five (36.6%) patients were supported with an LVAD (median age, 48 years, 26% female) and 95 (63.4%) with a BiVAD (median age, 45 years, 28% female). Twenty-five (45.4%) of the LVAD and 49 (51.2%) of the BiVAD patients were eventually transplanted (p=0.47) with an average time to transplantation 588 and 562 days respectively. Fifteen (27.2%) LVAD and 35 (36.8%) BiVAD patients died before transplantation (p=0.23) with an average time on VAD support before death 269 and 302 days respectively. Evidence of sensitization pre-VAD was found in 17.1% of the LVAD and 12.5% of the BiVAD patients (p=0.53); these percentages rose to 40.0% (p=0.001) and 43.7% (p=0.0001) respectively at 2 to 12 months post-VAD implantation. However, the post-VAD allosensitization status was not statistically different between the LVAD and the BiVAD group. Among the 25 LVAD patients who were transplanted 7 (30.4%) were sensitized immediately before transplantation, compared to 22 (46.8%) out of 49 BiVAD patients (p=0.16). Conclusion Both LVADs and BiVADs are a factor of de novo anti-HLA antibody development in adult patients after implantation. Sensitization pattern does not seem to differ between LVAD and BiVAD patients. Additionally, patients supported by LVAD or BiVAD have similar chances of finding a suitable donor heart.

Zika fever immunodiagnostics: overview of test systems.

The threat of rapid spread of Zika virus beyond endemic regions has given rise to more research in field of epidemiology and clinic, as well as to the search for Zika fiver new diagnostic and preventive tools. Between 2013 and 2017 in Russia 18 cases of infection transmission by travellers were reported. Fever Zika reference monitoring center in Volgograd Research AntiPlague Institute (Volgograd, Russian Federation) provides counseling and methodological assistance on laboratory diagnosis and monitoring of Zika fever. In this regard, a literature review of commercial test systems for immunodiagnostics of this infection was performed. Currently, a number of test systems for solid-phase enzyme-linked immunoassay method (ELISA), immunochromatography and indirect immunofluorescent method (IIFT) have been developed for immunodiagnostics of Zika fever. Euroimmun Ltd. remains the only manufacturer that has access to detailed information on validation of the specificity of the produced diagnostic kits. Independent studies confirm that Euroimmun test systems have high specificity and high sensitivity, which is proved by the study of the material from various populations, including Europeans travelling to Zika virus endemic regions and people residing in these regions. A detailed overview of characteristics of Euroimmun test systems for immunodiagnostics of Zika fever allows us to conclude that there is a rationale for the use of these test systems for Russian Federation sanitary protection by identification of antibodies in patients, presumably infected with the Zika virus.

Асоціації рівня макрофаг-інгібуючого фактора зі ступенем тривожності та депресії у хворих, що перенесли інфаркт міокарда з елевацією сегмента ST

The aim – to investigate associations between macrophage inhibitory factor (MIF) and anxiety, depression and stress in patients with ST-segment elevation myocardial infarction (STEMI).Materials and methods. 73 patients with STEMI with TIMI-III aged 58.37±10.34 were enrolled in the study. Revascularization tactics distribution was the following: 43 (58.9 %) patients underwent primary percutaneous coronary intervention using bare-metal coronary stent, 30 (41.1 %) – pre-hospital thrombolysis with subsequent percutaneous coronary intervention within12 hours. Echocardiography was done during the period of hospitalization. Serum MIF was studied using the enzyme linked immunoassay method. Anxiety, depression and stress were determined with DASS-21 questionnaire.Results and discussion. When comparing the median values of the MIF level in STEMI patients, a statistically significant increase in the MIF concentration relative to control group (2582.80 [1308.40–4122.20] and 573.75 [397.80–1016.75] ng/ml, p<0.001). It evidences the activation of the formation of MIF after the index event. Positive correlation was determined between the level of MIF, the level of troponin I (r=0.33; р=0.045) and blood leukocytes (r=0.36; р=0.039) that confirms its involvement into inflammation and necrosis processes. The univariant and multivariate analysis revealed the effect of the degree of depression and anxiety growth on the DASS-21 questionnaire (p=0.0138 and p=0.0050, respectively) on the increasing of the MIF.Conclusions. The data obtained confirm the relationship between the MIF and anxiety and depression prior the onset of myocardial infarction, indicating their influence on the processes of inflammation.

Difference of Vitamin D and Interleukin-6 Levels in Children with Steroid- Resistant, Steroid-Sensitive and Idiopathic Nephrotic Syndrome

Idiopathic nephrotic syndrome (INS) is the most prevalent autoimmune glomerular disease in children and its pathogenesis is correlated with high level of interleukin 6 (IL-6) and low level of vitamin D. This study was aimed to investigate the difference of vitamin D and IL-6 level in steroid resistant nephrotic syndrome (SRNS), steroid sensitive (SSNS), and idiopathic nephrotic syndrome (INS). This research was designed as cross sectional involving 45 subjects which then divided into 3 groups as follows: SRNS, SSNS, and INS. A level of serum 25 (OH)D was measured by Enzyme-linked Immuno Assay Method then categorized as sufficiency, insufficiency, and deficiency. Level of IL-6 serum was measured by ELISA method. Results showed that IL-6 level was significantly different among three groups, in which SRNS had the highest value (ANOVA, p < 0.05). Further analysis demonstrated that IL-6 level correlated with steroid resistance (Spearman correlation test, p = 0.000, r = 0.692). Vitamin D status was significantly different among three groups (Chi square, p = 0.03) and associated with steroid resistance (Spearman correlation test, p = 0.000, r = 0.568). Moreover, IL-6 level associated with 25 (OH) D level in SRNS group (Pearson correlation test, p = 0.020, r = 0.591) but not in the SSNS and INS group. We conclude that IL-6 levels were significantly higher in SRNS group as compared to other groups. Otherwise, vitamin D status were significantly lower in SRNS compared with other groups. An IL-6 level was negatively correlated with vitamin D status in patients with NS, specifically in SRNS group.

The relationship of macrophage migration inhibitory factor with clinical and anamnestic parameters in patients with ST-segment elevation myocardial infarction

Objective — to investigate the association between levels of the macrophage migration inhibitory factor (MIF) and the clinical and anamnestic parameters in patients with ST segment elevation myocardial infarction (STEMI) and to determine the factors, affecting this relationship.Materials and methods. The study involved 73 patients with previous confirmed STEMI and successful recovery of blood flow (TIMI-III). The mean patients’ age was (58.37 ± 10.34) years. Clinical, anamnestic and biochemical indicators were determined. The level of MIF was studied using the enzyme linked immunoassay method (RayBio® Human MIF ELISA KIT, USA). The general group of patients was divided by the median MIF level: the first group consisted of patients with low or moderate MIF (≤ 2582.80 ng/ml), n = 36, the second group consisted pf patients with high levels of this cytokine (&gt; 2582.80 ng/ml), n = 37. Mono- and multivariate regression analysis was carried out to identify factors, that mostly significantly affected the MIF levels.Results and discussion. The comparison of median values of the MIF levels in STEMI patients revealed the significant increase in MIF levels vs the control group (2582.80 [1308.40—4122.20] and (573.75 [397.80—1016.75] ng/ml, p &lt; 0.001), that confirmed activation of the MIF formation because of the index event. Positive correlation was determined between the MIF levels, levels of troponin I (r = 0.33; p = 0.045) and blood leukocytes (r = 0.36; p = 0.039). Anterior myocardial infarction (p = 0.047) and injury of left anterior descending artery (p = 0.016) were detected more frequently in the group of patients with high MIF levels, that indicates its directly connection with the size of the myocardial infarction in combination with significantly higher troponin level. Multivariate and univariate logistic analysis showed the reliable independent association between the anterior localization of the MI and the elevated MIF level (OR = 0.2282, 95 % CI [0.1035—0.7680], p = 0.0133), as well as the presence of stable angina to the index event and the lower level of cytokine (OR = 0.2318, 95 % CI [0.0637—0.8438], p = 0.0266).Conclusions. The macrophage migration inhibitory factor at STEMI is involved in the processes of inflammation and necrosis, its increase indicates the direct relationship with the myocardial infarction extension. The presence of stable angina prior the myocardial infarction promotes the pre-conditioning effect and a lower degree of damage, and proportionally lower MIF levels.

Low levels of vitamin D and the relationship between vitamin D and Th2 axis-related cytokines in neuromyelitis optica spectrum disorders

ObjectiveThe impact of vitamin D have been studied in neuroinflammation disorders, and as the newly discovered Th2-related cytokines, IL-25, IL-31 and IL-33 may also play important roles in the lesions of neuromyelitis optica spectrum disorders (NMOSD). This study sought to investigate the clinical profiles of vitamin D and Th2 axis-related cytokines and their relationships in patients with NMOSD. MethodsEighty-four NMOSD patients and 84 healthy controls (HC) were evaluated for serum levels of the total vitamin D [25(OH)D], 25-hydroxyvitamin D2 [25(OH)D2] and 25-hydroxyvitamin D3 [25(OH)D3] by means of high performance liquid chromatographytandem mass spectrometry (HPLC-MS/MS). Meanwhile, serum AQP4-IgG (n = 84) were detected by an AQP4-transfected cell-based assay (CBA) and IL-25, IL-31, IL-33 (n = 32) were performed using enzyme-linked immunoassay (ELISA) method. ResultsThe serum levels of 25(OH)D, 25(OH)D2 and 25(OH)D3 were significantly lower in NMOSD group as compared to HC group. There were also significant differences in serum vitamin D levels between the acute phase group and remission group except the 25(OH)D2 levels (p = 0.070). No correlations were detected between vitamin D and disease activity or vitamin D and disease disability. Furthermore, serum 25(OH)D, 25(OH)D2, and 25(OH)D3 levels were not correlated with serum IL-25, IL-31, and IL-33 levels, the location of lesions and the number of lesion locations. ConclusionOur result showed hypovitaminosis D in NMOSD patients. The activity of 25(OH)D3 seemed to be closer to 25(OH)D than 25(OH)D2. Low levels of 25(OH)D/25(OH)D3 might represent a risk factor for the disease activity in patients with NMOSD.

Reduced cerebrospinal fluid levels of interleukin-10 in children with febrile seizures

PurposeThe exact etiology of febrile seizures (FS) is still unclear. However, it is thought that cytokine network activation may have a causative role. Therefore, this study aimed to evaluate the levels of interleukin-12 (IL-12) as a proinflammatory cytokine, interleukin-10 (IL-10) as an anti-inflammatory cytokine, and interferon-β (IFN-β), a marker of toll-like receptor-3 activation as a host response to viruses. These cytokine levels were analyzed in the cerebrospinal fluid (CSF) of children after a FS. MethodsWith the approval of the Human Research Ethics Committee, 76 patients with FS, who underwent lumbar puncture (LP) for the exclusion of central nervous system (CNS) infection, and who didn’t have CSF pleocytosis, were included in the study. The control group consisted of 10 patients with similar ages, with an acute febrile illness and who required LP to exclude CNS infection. The analyses were made by the enzyme-linked immunoassay method. ResultsAge, gender distribution and CSF IL-12 and IFN- β levels did not differ, but CSF IL-10 levels were significantly lower in the FS group as compared to the control group (0.78 ± 4.5 pg/ml, versus 27 ± 29 pg/ml, p < 0.0001). ConclusionThe low-level of CSF IL-10, considering its anti-inflammatory properties, may play a role in the etiopathogenesis of FS.

Seroprevalence of TORCH Infections in Children

Background : The acronym “TORCH” was introduced to highlight a group of pathogens that cause a congenital and perinatal infections: Toxoplasma gondi, rubella virus, Cytomegalovirus (CMV) and Herpes Simplex Virus (HSV). These pathogens are often associated with congenital anomalies. Congenital malformations have a direct impact on the family. This study was undertaken to detect the serological evidence of TORCH infections in children, by establishing the presence of specific IgM antibodies.&#x0D; Methods: During the period 1st June 2016 to 30th May 2017, 58 suspected TORCH infection cases were included from Paediatrics Department of CMOSH for TORCH antibody detection. The children were in the age of 0 day to 1 year with an average age of 3.3±2.59 months. The serum samples were tested forIgM and IgG antibodies against TORCH agents by using enzyme linked immunoassay method (ELISA).&#x0D; Results: Among the 58 children, seropositivity was found in 55 (94.82%) cases. Of the 55 seropositive cases serological evidence for combination of IgM and IgG with any one of the TORCH agents was detected in 25 (43.10%) and IgG alone was detected in 30 (51.72%) children. IgM/IgG antibody positivity to Toxoplasma, Rubella, CMV and HSV was 21(36.21%), 50(86.21%), 52(89.66%) and 8(13.79%) respectively. IgM anti-CMV were observed in highest number of cases 19(32.76%), followed by IgM anti-rubella 7(12.07%). IgM against HSV and Toxoplasma were found in equal number of cases, 3(5.17%) each.&#x0D; Conclusions: Awareness amongst clinicians and mothers about TORCH infections and their consequences should be developed.&#x0D; Chatt Maa Shi Hosp Med Coll J; Vol.17 (1); Jan 2018; Page 13-16

Comparison of Serum Interleukin-6 (Il-6) Levels Between Patients With HELLP Syndrome Versus Normotensive Pregnant

Background: Interleukin-6 (IL-6) has been known associated with oxidative stress and endothelial dysfunction, and also has important roles in pathogenesis of preeclampsia. Several studies have demonstrated that there was a significant increase of serum IL-6 levels in preeclamptic compared to normotensive pregnant women. However, study to evaluate serum IL-6 level in pregnant women with HELLP (hemolysis, elevated liver enzyme, and low platelet) syndrome is not well documented yet. This study aims to evaluate serum IL-6 levels between women with HELLP syndrome compared to normotensive pregnant.Methods: The research design was observational cross-sectional study. Samples were recruited consecutively using inclusion and exclusion criteria from emergency department and inpatient wards at Dr. Sardjito General Hospital, Yogyakarta, Indonesia. Serum IL-6 were collected from venous blood sample and measured by enzyme-linked immunoassay (ELISA) method. Independent-samples t-test or Mann-Whitney test was used to compare serum IL-6 values between women with HELLP syndrome and normotensive pregrnancy. A statistical measurement conducted using SPSS IBM Statistics 19® and considered significance when p&lt;0,05.Results: There were 46 subjects, consisted of 23 women with HELLP syndrome and 23 women with normotensive pregnancy. Serum IL-6 level was a mean of 0,11±0,08 pg/mL and 0,15±0,20 pg/mL in women with HELLP syndrome compared to normotensive pregnancy respectively. There was no significant statistical difference between both groups (p=0,17).Conclusion:.The result of this study has shown that no significant difference of serum IL-6 level in HELLP syndrome women compared to normotensive pregnant women.

Rubella virus infections and immune status among pregnant women before the introduction of rubella vaccine in Amhara Regional State, Ethiopia

BackgroundRubella and its associated congenital anomalies have been greatly reduced in most developed countries through use of the rubella vaccine. However, the magnitude of the problem is underestimated and there are no well-established rubella/congenital rubella syndrome prevention and control strategies in many developing countries, including Ethiopia. The aim of this study was to determine the prevalence of rubella virus infections among pregnant women and their immune status before the introduction of rubella vaccine in Amhara Regional State, Ethiopia. MethodsA prospective cross-sectional study was conducted among pregnant women in Dessie, Felege-Hiwot, and University of Gondar referral hospitals, from December 2015 to February 2017. After obtaining written informed consent, socio-demographic data, reproductive history, clinical manifestations, and the possible risk factors for rubella virus infections were collected using a structured questionnaire. The laboratory analysis of rubella-specific antibodies was done using an enzyme-linked immunoassay method on venous blood samples. Data were entered and analyzed using IBM SPSS Statistics version 20. Binary logistic regression was used to determine the strength of association between the dependent variables and covariates. ResultsA total of 600 pregnant women were included in the study. Their mean age was 26.4±5years (range 16–40 years). The overall seroprevalence of rubella infection was 89%. Of the total study participants, 9.5% were positive for rubella-specific IgM antibody, which indicates acute/recent rubella virus infection. In contrast, 79.5% of them had protective levels of rubella-specific IgG antibody and were immune as a result of previous wild-type rubella infection. However, 11% of the pregnant women were negative for both rubella-specific antibodies; these women represent the susceptible group. ConclusionsA large number of pregnant women had acute/recent rubella virus infections at the time of data collection, indicating that the virus is endemic in the study area. More than a tenth of pregnant women were found to be susceptible to acquiring the infection in future pregnancies, with the possible risk of rubella-associated congenital anomalies. Hence screening of all women of child-bearing age before conception and during pregnancy might reduce the devastating effects of the virus on the developing fetus.

Efficient and selective separation of metronidazole from human serum by using molecularly imprinted magnetic nanoparticles.

Magnetic molecularly imprinted nanoparticles were prepared through surface-initiated reversible addition fragmentation chain transfer polymerization by using metronidazole as a template. The molecularly imprinted magnetic nanoparticles were characterized by attenuated total reflection Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, transmission electron microscopy, X-ray diffraction, and vibrating sample magnetometry. The adsorption characteristics were also investigated and the kinetics of the adsorption of metronidazole on the imprinted nanoparticles were described by the second-order kinetic model with the short equilibrium adsorption time (30min). The adsorption isotherm was well matched with the Langmuir isotherm in which the maximum adsorption capacity was calculated to be 40.1mg/g. Furthermore, the imprinted magnetic nanoparticles showed good selectivity as well as reusability even after six adsorption-desorption cycles. The imprinted magnetic nanoparticles were used as a sorbent for the selective separation of metronidazole from human serum. The recoveries of metronidazole from human serum changed between 97.5 and 99.8% and showed similar sensitivity as an enzyme-linked immunoassay method. Therefore, the molecularly imprinted magnetic nanoparticles might have potential application for the selective and reliable separation of metronidazole from biological fluids in clinical applications.

Anti-Mullerian Hormone: Establishing the Ovarian Reserve Range with Age in Rivers State Women, Niger Delta Region of Nigeria

Background: The measurement of circulating anti-Mullerian hormone (AMH) in the plasma of adult women has been used as a tool in the assessment of ovarian reserve. This is based on its ability to reflect the number of growing follicles in the ovaries and knowing the level of AMH in a woman’s blood is generally a good indicator of her ovarian reserve and this can be achieved by making reference to a decision values. Objective: The purpose of this study was to measure the level of this hormone in normal, apparently healthy subjects in Rivers State, Nigeria with respect to age. The percentage of women, at their reproductive age with infertility problems in the state is in the increase and the essence was to establish a reference range for the hormone because of its clinical application in women fertility. Materials/method: A total of one thousand and sixty two apparently healthy women divided into four age groups were recruited for this study; this comprised of 300 each in age group of 20 - 30, 31 - 40, 41 - 50 respectively and 162 in 51 - 60 years age group from May 2014 to June 2017. They were randomly selected after filling a questionnaire form that has the information of age, tribe, last date of menses and whether on medication for any infertility problems. Excluded from the study were subjects identified with different endocrine dysfunction and/or on drugs for different types of anovulatory dysfunction. The Enzyme linked Immunoassay method was used in the measurement of the AMH. Results: The result from the measurement of plasma AMH levels showed a Mean ± SD value of 3.193 ± 0.943, 1.644 ± 0.691, 0.516 ± 0.276 and 0.135 ± 0.081 ng/ml respectively for the 1st, 2nd, 3rd and 4th age groups respectively. The mean value for the AMH decreases with increasing age and was statistically significant at the different age group levels (P < 0.05). The distribution pattern of the AMH, showed that a greater percentage of the women, about 46%, in age group 41 - 50 and 51 - 60 had a significant reduced level of the hormone from the group mean value, when compared with age group 20 - 30 and 31 - 40 years with about 15%. Conclusion: The reference, established range for the AMH (that is, 95% prediction interval), with respect to age in Rivers State women of Niger Delta region, Nigeria is as follows: −2.250 - 4.136; 0.953 - 2.350; 0.240 - 0.792 and 0.054 - 0.216 respectively for the age groups. This study summarizes the findings concerning AMH and its role as a marker for the quantitative aspect of ovarian reserve, ovarian aging, as well as ovarian dysfunction in this region of the country.

Designing a novel ELISA method based on CagA, NapA recombinant antigens to increase sensitivity and specificity of Helicobacter pylori whole cell antigen detection.

In this research, we designed a direct Enzyme Linked Immunoassay method to detect Helicobacter pylori antigens in stool specimens. Helicobacter pylori infection as the worldwide problem is related to many gastrointestinal disorders such as gastritis, gastric cancer, non-ulcer disease, peptic ulcer disease and duodenal ulcer. We produced and purified recombinant CagA and NapA antigens in Escherichia coli and extracted their antibodies from a panel of positive sera specimens. We designed a novel enzyme linked immunoassay direct method in combination with the whole cell for the qualitative and quantitative detection of Helicobacter pylori antigens in human stool. Assay performance was evaluated by histopathology staining and urease activity. The sensitivity and specificity of assay was determined as 91.7 [95% confidence interval: 89.3-95.6%] and 93.1% [95% CI: 91.2-96.4%], respectively. Novel ELISA exhibits enhanced sensitivity and specificity of Helicobacter pylori detection in comparison with another commercially available kit. Combination of the recombinant antigens and whole cell of Helicobacter pylori in immunoassay designing is a new approach about early diagnosis, treatment and fallowing up of the Helicobacter pylori infected patients, especially in peptic cancer cases.

Th2 axis-related cytokines in patients with neuromyelitis optica spectrum disorders.

Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory autoimmune disease of the central nervous system. Increasing evidence indicates that NMOSD is a Th2- and Th17-dominant disease. IL-25, IL-31, and IL-33 are three newly found Th2-related cytokines, and their roles in the pathogenesis of NMOSD have not been studied. This study aimed to measure the serum levels of IL-25, IL-31, and IL-33 in patients with NMOSD and evaluate their clinical implications. Serum was collected from patients with NMOSD (n=48) and healthy controls (HC, n=28). Serum level measurements of IL-25, IL-31, IL-33, IL-17A, and IL-6 were performed using enzyme-linked immunoassay (ELISA) method. The serum levels of IL-25, IL-31, and IL-33 were significantly higher in patients with NMOSD as compared to HC. The serum level of IL-31 was significantly correlated with IL-17A (r=0.382,P=0.009) in patients with NMOSD; the latter is a critical cytokine in the pathogenesis of NMOSD. The serum level of IL-33 was higher in patients with characteristic brain lesions than patients without (307pg/mL vs 166pg/mL, P=0.028). Furthermore, the serum level of IL-33 in the acute phase of the disease was positively correlated with annualized relapse rate (r=0.364, P=0.04). We found higher serum levels of IL-25, IL-31, and IL-33 in patient with NMOSD as compared to healthy controls. The serum level of IL-33 during acute phase was associated with more past attacks in patients with NMOSD.

CEREBROSPINAL FLUID LIPIDOMIC PROFILE IN ALZHEIMER’S DISEASE

Lipid dysregulation is suggested to participate in the pathogenesis of Alzheimer's disease (AD). However, alterations in individual lipid species in the cerebrospinal fluid (CSF) have not been examined. CSF samples were obtained from 57 AD patients and 68 control, cognitively normal individuals. b-amyloid 1–42 (Ab1–42), phosphorylated tau (P-tau) and total tau (T-tau) proteins were determined by Enzyme-Linked Immunoassay (ELISA) method. Lipid identification and quantification were performed by multi-dimensional mass spectrometry-based Shotgun Lipidomic (MDMS-SL) technique. Levels of long chain sphingomyelin, sulfatide and alkyl phosphatidyl choline were significantly lower for AD than in case of controls, while the level of short chain ceramid, phosphatidyl ethanolamine and phosphatidyl serine were significantly increased in AD. Sphingomyelin and alkyl phosphatidyl choline negatively, while phosphatidyl etanolamine positively correlated with the P-tau level of CFS. These new AD related lipidomic results provide a better understanding of lipid perturbations in AD pathogenesis and may be utilized in the searching for CSF biomarkers. Acknowledgement: This work was supported by the Hungarian Research and Technology Innovation Fund through the Hungarian Brain Research Program (KTIA_13_NAP-A-II/16).

Enzyme-linked immunoassay based on imprinted microspheres for the detection of sulfamethazine residue

This study attempts to develop an enzyme-linked immunoassay (ELISA) using a molecularly imprinted polymer (MIP) as an artificial recognition element. The MIP microspheres were prepared using precipitation polymerization with SM2 as the template molecule, methacrylic acid as the functional monomer and ethylene glycol dimethacrylate as the cross-linker. After the microspheres were coated in microtiter plate wells, the molecular imprinting ELISA (MI-ELISA) method was established based on the direct competition between free SM2 and horseradish peroxidase (HRP)-labelled SM2 in heterogeneous mode. The linear regression analysis data for the calibration curve showed a good linear relationship with a regression coefficient of 0.999 in the concentration range of 100μgL−1–3200μgL−1. Furthermore, following the selective solid-phase extraction (SPE) with bulk SM2 MIPs as the sorbent and MI-ELISA detection, the limits of detection and quantification were 6.8μgkg−1 and 20.4μgkg−1, respectively, for SM2 in swine muscle. For the first time, MI-ELISA combined with molecular imprinting SPE was developed to determine trace SM2 in real samples, and the results show that it can be a useful analytical tool for quick detection in residue analysis.

Detection of sulfonamide and tetracycline antibiotic residues in Turkish pine honey

ABSTRACTPine honey is a honeydew honey that can be produced endemically in eastern Mediterranean. Turkey is the world’s largest producer of pine honey with around 95% export share. Antibiotic residue in honey is still an important problem worldwide. The aim of this study was to determine sulfonamide and tetracycline group antibiotics in pine honey collected from Aegean Region of Turkey. For this purpose, 59 natural pine honey samples were detected by competitive enzyme-linked immunoassay method. Tetracycline group antibiotics were found in 35 honey samples out of 59 between 6 and 42 ppb, whereas in 24 of the samples the levels were below the detection level (< 4 ppb). Sulfonamide group antibiotics were found in 31 honey samples out of 59 between 3 and 32 ppb, while 28 samples were below the detection limit (< 2 ppb). There are no maximum residue limits established for antibiotics in honey according to the European Community regulations, which means honeys should not contain antibiotics.

Detection of environmentally hazardous pesticide in fruit and vegetable samples using gold nanoparticles

A new approach for simple, competitive and sensitive, dipstick immunoassay (DIA) was developed to detect organophosphorus pesticide. Residual pesticides such as kitazine in fruits and vegetables are one of the major food safety concerns for consumers in the world. It is the alternative method to nitrocellulose strip (NC) for the detection of harmful pesticides. The matrix effects of different food samples (tomato, cucumber, grapes and orange) were studied in flow through ELISA (Enzyme Linked Immunoassay) and DIA method. Different size morphology was synthesized from gold nanoparticles and bioconjugated to Ig G, which served as the detecting agent confirmed by AFM. The different concentration of organophosphorus in the food sample was detected by the development of purple color in the membrane strips. Among four fruit juices, the grape juice exhibits increased absorbance resulted that the bioconjugated gold nanoparticle is actively detected the kitazine present in the grape juice evidenced by ELISA method has good reproducibility and satisfactory accuracy. The interpretation of the results was visual, and these ELISAs can be used as convenient qualitative tools for the rapid screening ofkitazine residues in food samples.