Enzyme-linked Fluorescent Assay Method Research Articles

The Relationship Between D-Dimer Levels and the Value of Prothrombin Time and Activated Partial Thromboplastin Time in Patients with COVID-19 At Rsup Dr. M. Djamil Padang

Covid-19 spread widely resulting in a global pandemic that continues to this day. People who are confirmed to have severe COVID-19 will cause thrombosis which results in increased levels of D-dimer prolonging Prothrombin Time or Activated Partial Thromboplastin Time, increased fibrinogen, and thrombocytopenia. one of the parameters that determine the presence of coagulation can be seen from the levels of D-dimer, PT and APTT. The purpose of this study was to determine the relationship between D-dimer levels and the value of prothrombin time and activated partial thromboplastin time in Covid-19 patients. This research is analytic observational with a cross sectional study design. The sample in this study amounted to 30 respondents. The examination carried out was D-dimer levels using the ELFA (Enzyme Linked Fluorescent Assay) method using Vidas, Prothrombin Time and Activated Partial Thromboplastin Time using the Flouroscence Flowcytometry method using the Sysmex Cs 2500 tool. The data were analyzed using the Spearman’s rho correlation test. The results of the study on Covid-19 average levels of D-dimer 1696.23 g/ml ±1567.448 SD, the value of Prothrombin Time 11.103±1.1898 seconds and Activated Partial Thromboplastin Time 32.643±17.3796 seconds. Conclusion There is a significant relationship between D-dimer levels with the Prothrombin Time P value of 0.003 and with Prothrombin Time and Activated Partial Tromboplastin Time P value of 0.031. There was no significant relationship between D-dimer levels and Activated Partial Tromboplastin Time P value 0.110. High D-dimer levels are accompanied by a prolonged prothrombin time.

Performance of the Enzyme Linked Fluorescent Assay in Diagnosis of HIV Infection; Comparative Evaluation with other Methods.

Our study aimed to investigate the role of the enzyme linked fluorescent assay (ELFA) method in the diagnosis of Human Immunodeficiency Virus (HIV) infection by comparing it with enzyme-linked immunosorbent assay (ELISA) and chemiluminescent microplate immunoassay (CMIA) methods and its role in the HIV diagnostic algorithm and to update the recommended algorithm for HIV testing. We evaluated 101 HIV-reactive and 101 HIV-negative specimens. All samples were studied with the methods of anti HIV1/2 test micro-ELISA, ELFA, and CMIA. At the same time, HIV RNA PCR and western blot (WB)/rapid immunochromatographic test (RICT) were also studied with the same samples. All HIV RNA and WB positive samples (n = 101) were positive with micro-ELISA, CMIA and ELFA. Twenty-five negative samples of HIV RNA and WB were positive with micro-ELISA and CMIA, while just 6 samples were positive with ELFA. When all samples were evaluated together, the false positivity rate of the ELFA method was found to be 5.9%, and the false positivity rates of the micro-ELISA and CMIA methods were determined to be 31.7% and 30.7%, respectively. It was determined that there is a high level of agreement between the ELFA method and confirmation tests. It was thought that it might take place in the preconfirmation stage. As can be seen from the results obtained, the false positive rate by ELFA method was found to be about five times lower than that of micro-ELISA and CMIA methods. Considering that antigen (p24) and antibody positivity can be given separately with this aspect, it can be considered that there is a confirmation place in HIV diagnosis algorithm.

Correlation between Cytomegalovirus (CMV) Infection in Children with Cerebral Palsy (CP) and Future Anxiety among Parents

One of the causes of Cerebral Palsy (CP) is a viral infection in the mother during pregnancy. The most recorded infections were Cytomegalovirus (CMV) infections. Inappropriate parenting can affect the psychological condition of CP children. Psychological conditions that are not good can cause the child's immune system to decrease and increase the risk of CMV reactivation. This is one factor in the emergence of parental anxiety about the future sustainability of their CP children. It was to determine the relationship of anti-cytomegalovirus (CMV) Immunoglobulin G in children with CP with future anxiety among parents. The design of this study used a cross-sectional approach that using CP children and their parents. Serological examination used the ELFA (Enzyme-Linked Fluorescent Assay) method for 40 CP children and a questionnaire on the future anxiety of parents of children with special needs were answered directly by the parents of CP children, 40 fathers and 40 mothers. Analysis of the research data used Spearman Rank. The results of this study found an anti-CMV IgG seropositive relationship in CP children with future anxiety in fathers (p-value = 0.022; a 0.05) and mothers (p-value = 0.011; a 0.05). If anxiety in parents has not good treatment, it will impact the care of CP children, the developmental progress of CP children will not optimal.

Immunization Against Hepatitis A in Migrant Children: Three Vaccination Strategies, A Retrospective Study.

Hepatitis A is endemic in many countries. Swiss guidelines recommend vaccinating patients native from endemic areas. In Geneva's Children's hospital, migrant children are screened and vaccinated if seronegative. Because hepatitis A's prevalence is decreasing worldwide, more children are seronegative at arrival, highlighting the need for immunization in medical centers and refugee camps and questioning the benefits of systematic serology. Other Swiss hospitals vaccinate regardless of serostatus. This study's aim is to assess migrant children's immunity according to origin and age, and the cost-effectiveness of different immunization strategies. We retrospectively analyzed 329 children's serostatus (1-16 years of age) between 2012 and 2015, using enzyme-linked fluorescent assay method. Serology and vaccine costs were based on local prices. Groups were compared with χ test and the age-seropositivity relationship was studied with linear regression. The predominant regions were the Eastern Mediterranean and European Regions with mostly negative serologies (71% and 83%) and the African Region with mostly positive serologies (79%). Immunity varied depending on birth country. Regardless of region, seropositivity increased with age (P < 0.001). The most cost-effective vaccination strategy was an individualized approach based on age and origin, reducing costs by 2% compared with serology-guided immunization and by 17% compared with systematic vaccination. Many migrant children >5 years old are seronegative and at risk of clinical infection. They need to be immunized. New guidelines according to age and origin should be defined to reduce immunization costs. We recommend systematic vaccination for patients <5 years old or native from low endemicity areas (≤25.7% of seropositivity). For the others, we propose serology-based vaccination.

Investigation of Hepatitis E Seroprevalence in HIV Positive Patients by a Novel Enzyme Linked Fluorescent Assay Test.

Hepatitis E virus (HEV) infection is usually an acute self-limiting disease, which causes rapidly progressive cirrhosis and chronic infection in patients with hematological malignancies, patients requiring chemotherapy, and HIV-infected patients. The aim of this study was to investigate the positivity of hepatitis E IgM and IgG in HIV positive patients with the recently introduced Enzyme Linked Fluorescent Assay (ELFA) commercial kits. The study included 126 patients who were followed up by the Infectious Diseases and Clinical Microbiology Clinic of Sakarya University Training and Research Hospital between October 2017 and December 2018 for HIV positivity. Serum samples of the patients were evaluated for anti-HEV IgG and IgM positivity with a novel commercially available kit using the ELFA method (bioMerieux, France). The study group consisted of 126 patients with HIV infection. Anti-HEV IgG antibodies were studied primarily from plasma samples. Anti-HEV IgM positivity was also investigated in patients with anti-HEV IgG positivity. The study group consisted of 114 (90.5%) males and 12 (9.5%) females with a mean age of 38.11 ± 13.32 (min: 18, max: 80) years. Anti-HEV IgG was positive in 5 (4.0%) HIV-positive patients. One of the anti-HEV IgG positive patients was newly diagnosed with HIV and the other four patients were being followed up for HIV positivity. Anti-HEV IgM was negative in all patients. None of the patients with anti-HEV IgG positivity had anti-HCV and HBsAg positivity. In the study, anti-HEV IgG positivity was found to be 4% in HIV-positive individuals, and no HCV and HBV co-infection was detected in any patients with HIV and HEV coexistence. HEV infections do not emerge as a priority among HIV-infected people, but HEV should also be investigated in HIV-infected individuals with liver abnormalities of uncertain etiology.

Evaluation and validation of an alternative method to detect Campylobacter spp. in dairy products.

Foods implicated in human campylobacteriosis include raw or undercooked poultry and raw dairy products. Because Campylobacter spp. are the most frequently reported cause of bacterial infection in the European Union and because conventional methods are cumbersome, rapid methods for Campylobacter detection and quantification in food are needed. With this study we sought to validate, according to the standard procedure (UNI EN ISO 16140:2003), an alternative to the reference analytical method (UNI EN ISO 10272-1:2006) for official controls of Campylobacter spp. in raw milk and dairy products. Milk samples collected from 16 milk vending machines located throughout the Genoa metropolitan area were analyzed using two different methods, an enzyme-linked fluorescent assay (ELFA) and a real-time PCR assay, and evaluated in parallel against the reference method. In addition, a total of 460 samples of raw milk collected from milk vending machines were analyzed by ELFA. Results obtained with ELFA showed it was compliant with UNI EN ISO 10272-1:2006 criteria and that the immunoassay had 100% sensitivity, specificity, and accuracy. Regarding samples of milk vending machines, 5.0% (23/460) tested positive at ELFA screening and were subsequently confirmed as C. jejuni. Validation according to UNI EN ISO 16140:2003 of the ELFA method suggests it may be a useful alternative to conventional methods for detecting Campylobacter spp. in official controls.

Detection of Escherichia coli O157 in raw and cooked meat: comparison of conventional direct culture method and Enzyme Linked Fluorescent Assay (ELFA)

Background: Verocytotoxin Escherichia coli is a frequent and important cause of diarrhea and haemolyticuremic syndrome all over the world. Consumption of ground beef, lettuce, and other kinds of food havebeen associated with outbreaks,The aim of this study was to detect the presence of E. coli O157 in meat products collected from hospitalfood catering services in Rome, using a rapid detection method in comparison with a standard culturemethod to verify the effectiveness of HACCP system.Methods: Three hundred and ten food samples (80 of cooked and 230 of raw meat) were screenedfor E.coli O157 by ISO culture method and by enzyme-linked-fluorescent-assay (ELFA)-based methods(VIDAS®system, bioMérieux). All isolates obtained were tested for VT1 and VT2 genes by PCR. Thestatistical analysis considered absolute frequencies and percentages. The K statistic was applied to assessagreement between direct culture method and the VIDAS system.Results: A total of 6 (1,9%) E.coli O157 isolates were recovered from raw meat samples by the culturemethod; of these only four were identified by PCR as VTEC producers. A total of 9 (2,9%) E.coli O157isolates were recovered from raw meat samples by the VIDAS® system. No E.coli O157 was detectedin cooked products. All comparisons between the direct culture method and the VIDAS system werestatistically significant (K= 0,795; p&lt;0.001).Conclusions: The present study showed how ELFA-based methods are highly specific and rapid for thedetection of E.coli O157 in food samples compared with the direct culture method. ELFA method is usefulto verify the effectiveness of the HACCP system in the risk management of potential contaminating hazardsduring the preparation of foods for susceptible persons

Comprehensive Survey of Pasteurized Fluid Milk Produced in the United States Reveals a Low Prevalence of Listeria monocytogenes

A comprehensive survey was undertaken to generate contemporary data on the prevalence of Listeria monocytogenes in pasteurized fluid milk produced in the United States. Samples (5,519) near the sell-by expiration date were purchased at retail outlets over a 5-week period and analyzed for presence of L. monocytogenes. Products consisted of whole milk, nonfat milk, and chocolate milk packaged in gallon, half gallon, quart, pint, and half-pint containers. Samples were collected from both large and small retail stores in urban and suburban locations in four FoodNet cities (Baltimore, Md., Atlanta, Ga., St. Paul/Minneapolis, Minn., and San Francisco, Calif.). Samples were prescreened for L. monocytogenes by the AOAC-approved rapid Vitek immunodiagnostic assay system, enzyme-linked fluorescent assay method. Positive prescreening samples were cultured according to the Bacteriological Analytical Manual, enumerated for L. monocytogenes with a nine-tube most-probable-number (MPN) procedure, and confirmed by biochemical characterization. The frequency of isolation of L. monocytogenes in these products was 0% (0 of 1,897) in whole milk, 0.05% (1 of 1,846) in nonfat milk, 0% (0 of 1,669) in chocolate milk, and 0% (0 of 107) in other (reduced fat and low fat) milk samples. Overall, L. monocytogenes was confirmed in only 0.018% of pasteurized milk samples (1 of 5,519). Enumeration of the single confirmed positive nonfat milk sample revealed low-level contamination (<0.3 MPN/g), even when sampled 5 days past the expiration of the sell-by date. The results confirm the low frequency of contamination of pasteurized fluid milk products by L. monocytogenes for products sold in the United States and reaffirm the reduction of contamination frequency of fluid milk by L. monocytogenes when compared with earlier estimates from the U.S. Food and Drug Administration Dairy Safety Initiatives Program.

The development of an efficient and rapid enzyme linked fluorescent assay method for the detection of Listeria spp. from foods

Conventional isolation methods, including the Health Products and Food Branch (HPFB), Health Canada method used for the isolation and identification of Listeria species and Listeria monocytogenes from foods and environmental samples, can take a week or more to complete and are usually labor-intensive. This has led to the development of various rapid methods which attempt to generate results comparable to standard methods but in a reduced time-frame with less hands-on operation. Our previous work with rapid detection systems indicated that the recommended enrichment protocols failed to grow the Listeria to detectable levels in a reliable and consistent manner. In the present study, a novel enrichment protocol is described and consists of samples being pre-enriched in Palcam broth (incubated at 35 °C for 26 h), enriched in UVM 2 (30 °C for 26 h) and then plated and analysed by a rapid detection kit, with results being generated after 52 h of incubation. In total, 200 naturally contaminated samples were analysed by both the HPFB standard method and the Palcam method. The results showed that the Palcam method is comparable to the HPFB method. Further analysis involved a rapid detection system, which applies ELISA techniques and automation in an enzyme-linked fluorescent assay (ELFA) system. This system, referred to as the Vitek Immuno Diagnostic Assay System or VIDAS, can identify Listeria to the genus or species ( L. monocytogenes) level. In this comparison, an additional 324 naturally contaminated samples were analysed by both the Palcam and ELFA methods. The sensitivity and specificity of the ELFA method were 98.1% and 97.0%, respectively, while the efficiency was 97.5%. False-negative and false-positive rates were 1.9% and 3.0%, respectively. These results show that the ELFA method (when using the Palcam method for pre- and secondary enrichment) was efficient and gave reliable results after 52 h of incubation, and met Health Canada's criteria for approval as a rapid method.

A COMPARISON OF THE HEALTH PROTECTION BRANCH AND THE ENZYME LINKED FLUORESCENT ASSAY METHODS FOR THE ISOLATION AND IDENTIFICATION OF LISTERIA SPP. AND LISTERIA MONOCYTOGENES FROM FOODS

Abstract The conventional Health Protection Branch method (MFHPB‐30) for the detection of Listeria monocytogenes is labor intensive and may require seven days to complete, depending on the initial levels of the organism in the food sample. Thus, there is a definite need for a rapid isolation and detection method. There are two commercially available versions of the enzyme linked fluorescent assay (ELFA) method for identifying Listeria from foods. One identifies Listeria to the genus level and the other to the species (L. monocytogenes) level, with both giving results within 50 h. In the present study, the two ELFA methods were compared to the HPB method to determine their efficiency.For the genus‐specific test, 326 uninoculated foods were compared by both the HPB and ELFA methods. When comparing the HPB method to the genus‐specific ELFA method, the latter had a false‐negative rate of 4.6%. There were no false positives. For the species‐specific ELFA assay, of the 186 foods compared by the two methods, a false‐negative rate of 8.1% and a false‐positive rate of 3.8% was observed in the ELFA method as compared to the HPB method.

Immunodiagnosis of angiostrongyliasis with monoclonal antibodies recognizing a circulating antigen of mol.wt 91,000 from Angiostrongylus cantonensis

In the analysis of excretory-secretory (ES) antigens from infective third-stage larvae (L3) of Angiostrongylus cantonensis, one major component of mol.wt 91,000 was not precipitated by pooled sera of patients with eosinophilic meningoencephalitis. Monoclonal antibodies (Mc Ab) secreted from two hybridoma cell lines, established against somatic antigens of L3, recognized this molecule but with different epitope specificities indicated by an additivity index (A.I.) of 83%. The 2 Mc Ab (TD2 and 3A5) belonged to IgG2a and IgM classes, respectively. Combinations of TD2 and 3A5 were used in a sensitive enzyme-linked fluorescent assay (ELFA) for the immunodiagnosis of human angio-strongyliasis. The double-antibody sandwich ELFA method was applied firstly to sera from experimentally infected rats using either TD2 or 3A5 to coat the assay plates. Two fluorescence unit (F.U.) peaks appeared in sera from infected rats collected 18 and 44 days after infection. Specimens from 35 patients were tested, all cerebrospinal fluids (CSF) and most sera (88%) showed positive reactions and the average F.U. of CSF was greater than that of serum.