scholarly journals Comparison of nonstructural protein-1 antigen detection by rapid and enzyme-linked immunosorbent assay test and its correlation with polymerase chain reaction for early diagnosis of dengue

2017 ◽  
Vol 9 (03) ◽  
pp. 177-181 ◽  
Author(s):  
Seema Gaikwad ◽  
Sandhya S. Sawant ◽  
Jayanthi S. Shastri
Keyword(s):  
Polymerase Chain Reaction ◽  
Nonstructural Protein ◽  
Antigen Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Nonstructural Protein 1 ◽  
Ns1 Antigen ◽  
Polymerase Chain ◽  
Sera Samples

Abstract INTRODUCTION: Early diagnosis of dengue is important for appropriate clinical management and vector control. Different serological tests based on the principle of immunochromatography and enzyme-linked immunosorbent assay (ELISA) are commonly used for detection of antigen and antibodies of dengue virus. The performance of these tests depends on the sensitivity and specificity. Hence, the study was undertaken to compare nonstructural protein-1 (NS1) antigen detection by rapid and ELISA with real-time polymerase chain reaction (RT-PCR) for diagnosis of dengue. MATERIALS AND METHODS: Prospective laboratory study was carried out on sera samples (n = 200) from clinically suspected cases of dengue. The sera samples were subjected for NS1 antigen detection test by rapid test, NS1 ELISA, and RT-PCR. The results of rapid and ELISA tests were compared with real Time PCR. RESULTS: The sensitivity, specificity, positive, and negative predictive value of rapid dengue NS1 antigen test were 81.5%, 66.7%, 78.2%, and 71.1%, respectively whereas that of NS1 ELISA were 89.9%, 100%, 100%, and 94%, respectively. Concordance of Rapid NS1 and NS1 ELISA with PCR was 75.5% and 94%. DISCUSSION AND CONCLUSION: NS1 antigen ELISA can be implemented in diagnostic laboratories for diagnosis of dengue in the acute phase of illness. The test also has great potential value for use in epidemic situations, as it could facilitate the early screening of patients and limit disease expansion.

scholarly journals Evaluation of Sandwich Enzyme-Linked Immunosorbent Assay and Reverse Transcription Polymerase Chain Reaction for the Diagnosis of Foot-and-Mouth Disease

Intervirology ◽  
2021 ◽  
pp. 1-6
Author(s):  
Salman Khan ◽  
Syed Asad Ali Shah ◽  
Syed Muhammad Jamal
Keyword(s):  
Polymerase Chain Reaction ◽  
Viral Genome ◽  
Foot And Mouth Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Kappa Value ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Level Of Agreement

<b><i>Background:</i></b> Foot-and-mouth disease (FMD) is an infectious and highly contagious disease of cloven-hoofed domestic and wild animals, causing heavy economic losses to the livestock industry. Rapid and reliable diagnosis of the disease is essential for the implementation of effective control measures. This study compared sandwich enzyme-linked immunosorbent assay (S-ELISA) and conventional reverse transcription polymerase chain reaction (RT-PCR) for the diagnosis of FMD. <b><i>Methods:</i></b> A total of 60 epithelial samples from suspected cases of FMD were tested using both S-ELISA and RT-PCR assays. The level of agreement between the assays was assessed by calculating the Kappa value. <b><i>Results:</i></b> S-ELISA detected 38 (63%) samples positive for FMD virus (FMDV). Being predominant, serotype O was detected in 22 (57.9%) of the total samples tested positive, whereas 9 (23.7%) and 7 (18.4%) samples were found positive for serotypes A and Asia-1, respectively. RT-PCR detected viral genome in 51 (85%) of the samples using pan-FMDV primers set, 1F/1R. Thirty-six samples were found positive and 7 negative by both the tests. The level of agreement between the tests was assessed by calculating the Kappa value, which was found to be fair (Kappa value = 0.303 and 95% CI = 0.089; 0.517) and significant (<i>p</i> = 0.009). However, 2 samples, which were found positive on S-ELISA tested negative on RT-PCR. This may be attributed to the presence of nucleotide mismatch(es) in the primer-binding sites that may have resulted in failure of amplification of the viral genome. The serotype-specific RT-PCR assays not only confirmed serotyping results of S-ELISA but were also able to establish serotype in 9 S-ELISA-negative but pan-FMDV RT-PCR-positive samples. <b><i>Conclusions:</i></b> The RT-PCR assay contributes significantly to establishing a quick, sensitive, and definitive diagnosis of FMD in resource-constrained countries. Samples giving negative results in S-ELISA should be tested in RT-PCR for the disease detection and virus typing.

Serum based polymerase chain reaction and enzyme linked immunosorbent assays for diagnosis of bovine brucellosis

2018 ◽  
Author(s):  
V. Naveen Kumar ◽  
M. Vijaya Bharathi ◽  
G. Selvaraju ◽  
K. Porteen ◽  
K. Vijayarani
Keyword(s):  
Polymerase Chain Reaction ◽  
Clinical Specimen ◽  
Enzyme Linked Immunosorbent Assay ◽  
Bovine Brucellosis ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Molecular Assays ◽  
Specific Pcr ◽  
Polymerase Chain ◽  
Sera Samples

Brucellosis is one of the economically important diseases in India and diagnosis of brucellosis using single test is cumbersome due to variation in sensitivity and specificity among the different test. The present study was aimed to assess the suitability of serum as clinical specimen in molecular diagnosis and evaluate the serology and molecular assays as in diagnosis of bovine brucellosis. A total of 821 bovine sera samples were subjected to indirect Enzyme Linked Immunosorbent Assay (i-ELISA) and serum based Polymerase Chain Reaction (PCR) assay. On serology 6.70 per cent positivity of brucellosis were reported and on PCR assay, 47 and 29 sera samples were positive for bcsp 31 genus specific and IS711 species specific PCR assay respectively with per cent positivity of 5.72 and 3.53. In comparison between serology and molecular test, 44 samples were positive for both assays and 11 and 3 samples were positive for serology and molecular assays individually. This study suggests that serum sample can be utilised as the choice of clinical specimen for both PCR assay and i-ELISA will be a future choice for the diagnosis of bovine brucellosis.

Detection of small round structured viruses in clinical and environmental samples by polymerase chain reaction

1995 ◽  
Vol 31 (5-6) ◽  
pp. 375-382 ◽  
Author(s):  
M. Wolfaardt ◽  
C. L. Moe ◽  
W. O. K. Grabow
Keyword(s):  
Polymerase Chain Reaction ◽  
Tap Water ◽  
Practical Method ◽  
Glass Wool ◽  
Polluted Water ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Elution Method

Norwalk (NV) and other small round structured viruses (SRSVs) have been identified as common causes of gastroenteritis. Outbreaks of Norwalk gastroenteritis have been associated with contaminated drinking water and food such as oysters and salads. The cloning and sequencing of the NV genome has made it possible to detect NV and related viruses by the reverse transcription-polymerase chain reaction (RT-PCR). We applied RT-PCR to detect SRSVs in faecal specimens from two gastroenteritis outbreaks in South Africa, designated “Christmas” and “Grootbrak” and were able to detect SRSVs in all of the three specimens from the Christmas outbreak and in two of 16 specimens from the Grootbrak outbreak. The RT-PCR procedure used appeared to be more sensitive for the detection of SRSVs in patient stool specimens than immune electron microscopy and NV antigen detection by enzyme linked immunosorbent assay. The RT-PCR procedure proved suitable for the detection of SRSVs in seeded samples of sewage, sewage sludge, river water, and tap water. However, sensitivity was lower for seeded samples of sewage and sludge than for tap water, which indicates interference by high levels of organic matter. The RT-PCR procedure was also used to show that small numbers of SRSVs can successfully be recovered from large volumes of water by means of a glass wool adsorption-elution method. Since no practical method is available for quantitation of the small numbers of SRSVs concerned, it was not possible to evaluate the efficiency of recovery. Although no SRSVs have been detected by direct testing of sewage and sludge samples, the results obtained in this study show that RT-PCR detection of SRSVs in sewage and polluted water environments is feasible, and that small numbers of the viruses can, like many other enteric viruses, successfully be recovered by means of a glass wool adsorption-elution method.

scholarly journals Head-to-Head Comparison of Rapid and Automated Antigen Detection Tests for the Diagnosis of SARS-CoV-2 Infection

2021 ◽  
Vol 10 (2) ◽  
pp. 265
Author(s):  
Julien Favresse ◽  
Constant Gillot ◽  
Maxime Oliveira ◽  
Julie Cadrobbi ◽  
Marc Elsen ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Clinical Performance ◽  
Antigen Detection ◽  
Clinical Validation ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Viral Loads ◽  
Automated Assay ◽  
Polymerase Chain ◽  
Asymptomatic Subjects

(1) Background: The detection of SARS-CoV-2 RNA in nasopharyngeal samples through real-time reverse transcription-polymerase chain reaction (RT-PCR) is considered the standard gold method for the diagnosis of SARS-CoV-2 infection. Antigen detection (AD) tests are more rapid, less laborious, and less expensive alternatives but still require clinical validation. (2) Methods: This study compared the clinical performance of five AD tests, including four rapid AD (RAD) tests (biotical, Panbio, Healgen, and Roche) and one automated AD test (VITROS). For that purpose, 118 (62.8%) symptomatic patients and 70 (37.2%) asymptomatic subjects were tested, and results were compared to RT-PCR. (3) Results: The performance of the RAD tests was modest and allowed us to identify RT-PCR positive patients with higher viral loads. For Ct values ≤25, the sensitivity ranged from 93.1% (95% CI: 83.3–98.1%) to 96.6% (95% CI: 88.1–99.6%), meaning that some samples with high viral loads were missed. Considering the Ct value proposed by the CDC for contagiousness (i.e., Ct values ≤33) sensitivities ranged from 76.2% (95% CI: 65.4–85.1%) to 88.8% (95% CI: 79.7–94.7%) while the specificity ranged from 96.3% (95% CI: 90.8–99.0%) to 99.1% (95% CI: 95.0–100%). The VITROS automated assay showed a 100% (95% CI: 95.5–100%) sensitivity for Ct values ≤33, and had a specificity of 100% (95% CI: 96.6–100%); (4) Conclusions: Compared to RAD tests, the VITROS assay fully aligned with RT-PCR for Ct values up to 33, which might allow a faster, easier and cheaper identification of SARS-CoV-2 contagious patients.

scholarly journals IDENTIFICATION OF DENGUE VIRUS SEROTYPES AT THE DR. SOETOMO HOSPITAL SURABAYA IN 2016 AND ITS CORRELATION WITH NS1 ANTIGEN DETECTION

2018 ◽  
Vol 23 (2) ◽  
pp. 151
Author(s):  
Jeine Stela Akualing ◽  
Aryati Aryati ◽  
Puspa Wardhani ◽  
Usman Hadi
Keyword(s):  
Polymerase Chain Reaction ◽  
Dengue Virus ◽  
Reverse Transcriptase ◽  
Ribonucleic Acid ◽  
Rna Virus ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Ns1 Antigen ◽  
Dengue Virus Serotypes ◽  
Polymerase Chain

Serotipe virus dengue yang beredar terus mengalami perubahan dan berbeda di setiap daerah. Pergeseran serotipe maupun genotipedi dalamnya, mempengaruhi terjadinya wabah dengue di berbagai negara. Perbedaan serotipe diduga bernasab dengan deteksi antigen(Ag) non-structural 1 (NS1), namun belum banyak penelitian yang mendukung hal tersebut. Penelitian potong lintang dikerjakan sejakFebruari-Agustus 2016 dan didapatkan 60 subjek infeksi virus dengue (IVD) dan 25 non-IVD. Ribonucleic acid (RNA) virus denguediperiksa di semua subjek menggunakan Simplexa Dengue Real-Time Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)termasuk identifikasi serotipe virus dengue dan pemeriksaan NS1 menggunakan uji cepat NS1 Panbio. Perbedaan perbandingan variabelkategorikal dianalisis dengan uji Fisher Exact. Kenasaban antara serotipe dengan deteksi Ag NS1 dianalisis dengan Chi-Kuadrat. RNAvirus dengue terdeteksi di 43 dari 60 subjek IVD (71,7%). Serotipe terbanyak adalah DENV-3 (62,8%). Pergeseran dominasi serotipetelah terjadi di Surabaya, sebelumnya dari DENV-2 ke DENV-1 dan sekarang DENV-3, kemungkinan akibat mobilitas pejamu, transporvirus dan faktor geografis. Kepekaan uji cepat NS1 75% dan kekhasan 100%. Persentase deteksi NS1 antar serotipe berbeda bermakna(p=0,002). Deteksi NS1 lebih rendah pada DENV-1 dibandingkan DENV-2 (p=0,007) ataupun DENV-3 (p=0,003). Serotipe virusdengue bernasab dengan deteksi NS1 (p=0,005). Ciri serotipe maupun genotipe virus dengue kemungkinan mempengaruhi sekresiNS1. Telah terjadi pergeseran serotipe virus dengue di pasien IVD di Surabaya sehingga diperlukan surveillance berkesinambunganuntuk memperkirakan terjadinya wabah. Serotipe bernasab dengan deteksi NS1. Salah satu penyebab hasil negatif palsu NS1 adalahperbedaan serotipe.

scholarly journals Combination of ELISA and RT-PCR tests in the diagnosis of toxoplasmic infection in aborted women and congenitally infected infants.

2014 ◽  
Vol 8 (3) ◽  
pp. 44-47
Author(s):  
Khitam Y. Obaid AL-Dujaily ◽  
Noor SH. Abdul-Amir
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Good Combination ◽  
Igg Antibodies ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Toxoplasmic Infection ◽  
Sera Samples

The diagnosis of toxoplasmic infection in aborted women and congenitally infected infants suspected to have toxoplasmosis infection can be difficult due to similarity symptoms with other diseases. A combination of symptoms, serology and polymerase chain reaction (PCR)may facilitate diagnosis of toxoplasmosis in some patients. The present study compare the detection of toxoplasmosis infection by ELISA IgA and IgG antibodies with Real Time polymerase chain reaction (RT-PCR)in the study subjects. A total of 81 sera samples, 57(70.3%) samples from aborted women and 24(29.7%)samples from congenitally infants have been studied. 49(86%) samples from the aborted women were positive and 8(14%) samples were negative as diagnosed by one or two of ELISA markers (IgAand IgG).The ELISA results indicated that 15(62.5%) samples from infants were positive and 9(37.5%) samples of them were negative. RT-PCR tests indicated that 33(67.3%) from the mothers and 6(40%) from the infants were agreed with ELISA positive samples. For ELISA negative samples, RT-PCR detected toxoplasmosis DNA in 4 (50%) and 2 (22.2%) for the mothers and infants respectively. Therefore, ELISA and RT-PCR can make a good combination tests in detection toxoplasmopsis infection.

Interlaboratory Evaluation of a New Reverse Transcriptase Polymerase Chain Reaction–Based Enzyme-Linked Immunosorbent Assay for the Detection of Circulating Melanoma Cells: A Multicenter Study of the Dermatologic Cooperative Oncology Group

2001 ◽  
Vol 19 (6) ◽  
pp. 1723-1727 ◽  
Author(s):  
U. Reinhold ◽  
C. Berkin ◽  
A.-K. Bosserhoff ◽  
A. Deutschmann ◽  
C. Garbe ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Tumor Cells ◽  
High Sensitivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Blood Samples ◽  
Melanoma Patients ◽  
Polymerase Chain ◽  
Interlaboratory Reproducibility

PURPOSE: Reverse transcription-polymerase chain reaction (RT-PCR)–based detection of tyrosinase mRNA is the most frequently used laboratory method for the detection of circulating tumor cells in melanoma patients. However, previously published results showed considerable variability in the PCR positivity rates. MATERIALS AND METHODS: We designed a collaborative study to assess the sensitivity, specificity, and clinical relevance of a new standardized RT-PCR–based enzyme-linked immunosorbent assay (ELISA) for the detection of circulating melanoma cells. Blood samples of healthy donors mixed with cells of a melanoma cell line were prepared in a blinded fashion, and aliquots were sent to seven participating laboratories experienced in RT-PCR. RESULTS: The results demonstrate a high sensitivity (1 melanoma cell/mL blood) and specificity (no false-negatives and 7.4% [2 of 28] false-positives) of the assay and a satisfactory rate of interlaboratory reproducibility. The analysis of aliquots of blinded samples derived from 60 melanoma patients identified tyrosinase mRNA in 17 of 60 (28.3%): three (20%) of 15 stage I patients, two (13.3%) of 15 stage II patients, five (35.7%) of 14 stage III patients, and seven (43.8%) of 16 stage IV patients. The interlaboratory reproducibility of positive samples, however, was extremely low and indicates the presence of low amounts of target mRNA. CONCLUSION: Reverse transcriptase-PCR ELISA has a high sensitivity and specificity for the detection of tyrosinase mRNA in peripheral blood cells. The low interlaboratory reproducibility for the detection of tumor cells in blood samples of melanoma patients, however, raises the question of relevance of this assay for clinical use.

scholarly journals Interference Between D and M Types of Plum pox virus in Japanese Plum Assessed by Specific Monoclonal Antibodies and Quantitative Real-Time Reverse Transcription-Polymerase Chain Reaction

2006 ◽  
Vol 96 (3) ◽  
pp. 320-325 ◽  
Author(s):  
Nieves Capote ◽  
M. Teresa Gorris ◽  
M. Carmen Martínez ◽  
Margarita Asensio ◽  
Antonio Olmos ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Monoclonal Antibodies ◽  
Real Time ◽  
Reverse Transcription ◽  
Enzyme Linked Immunosorbent Assay ◽  
Plum Pox Virus ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Japanese Plum ◽  
Polymerase Chain

The dynamics of virus interference between two isolates of Plum pox virus (PPV) belonging to the main PPV types, D and M, were analyzed in Japanese plum (Prunus salicina) by challenge inoculations. To assess the consequences of a PPV-M infection on plum already infected with PPV-D, and vice versa (predominance of one of the strains, recombination, synergism, symptoms aggravation, and so on), 30 Japanese plum trees were graft inoculated with PPV-D or PPV-M isolates in quarantine conditions. One year postinoculation, in the event that the inoculated isolates were detected in the whole plant, a second challenge inoculation (PPV-M or PPV-D, respectively) was performed by grafting. The presence of PPV-D, PPV-M, or both was monitored for 7 years by double-antibody sandwich indirect enzyme-linked immunosorbent assay using specific monoclonal antibodies. Reverse transcription-polymerase chain reaction (RT-PCR) with D- and M-specific primers confirmed the serological typing. Real-time RT-PCR assays were performed using D- and M-specific fluorescent 3′ minor groove binder-DNA probes, which were able to detect and quantify PPV populations in the inoculated plants with greater precision. The presence of PPV-D in Japanese plum did not cross-protect the trees against PPV-M infection. In PPV-D-infected plants, the PPV-M strain used as challenge inoculum behaved differently depending on the plum cultivar assayed. In cv. Black Diamond, PPV-M invaded the plant progressively, displacing the previous PPV-D population; whereas, in cv. Sun Gold, both PPV isolates coexisted in the plant. In contrast, the PPV-D isolate used was unable to infect plants of both cultivars in which a PPV-M population already was established. After 7 years, no synergism was observed and no recombination event between PPV-D and PPV-M genomes was detected.

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