scholarly journals Low Sensitivity of Real Time PCRs Targeting Retrotransposon Sequences for the Detection of Schistosoma japonicum Complex DNA in Human Serum

Pathogens ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1067
Author(s):  
Hagen Frickmann ◽  
Ulrike Loderstädt ◽  
Beatrice Nickel ◽  
Sven Poppert ◽  
Peter Odermatt ◽  
...  
Keyword(s):  
Human Serum ◽  
Real Time ◽  
Genome Sequences ◽  
Serum Samples ◽  
Consensus Primer ◽  
Hybridization Probe ◽  
Low Sensitivity ◽  
Positive Results ◽  
Multiple Variants ◽  
Potential Use

While hybridization probe-based real-time PCR assays targeting highly repetitive multi-copy genome sequences for the diagnosis of S. mansoni complex or S. haematobium complex from human serum are well established, reports on the evaluation of respective assays for the identification of S. japonicum complex DNA in human serum are scarce. Here, we assessed the potential use of the retrotransposon sequences SjR2 and SjCHGCS19 from S. japonicum, S. mekongi and S. malayensis for the diagnosis of Asian Schistosoma infections. Based on available S. japonicum sequences and newly provided S. mekongi and S. malayensis sequences, hybridization probe-based real-time PCRs targeting SjR2 and SjCHGCS19 of the S. japonicum complex were designed both as consensus primer assays as well as multi-primer assays for the coverage of multiple variants of the target sequences. The assays were established using plasmids and S. mekongi DNA. While the consensus primer assays failed to detect S. mekongi DNA in human serum samples, the multi-primer assays showed positive or borderline positive results but only in 9.8% (6/61) of serum samples from patients with confirmed S. mekongi infections. Some cross-reactions with samples positive for S. mansoni or S. haematobium were observed but with the SjCHGCS19-PCR only. In spite of the low sensitivity, the presented experience may guide future evaluations of S. japonicum-complex-specific PCRs from human serum.

scholarly journals Usefulness of real-time PCR assay targeting lipL32 gene for diagnosis of human leptospirosis in Uruguay

2013 ◽  
Vol 7 (12) ◽  
pp. 941-945 ◽  
Author(s):  
Sabina González ◽  
Juan Pablo Geymonat ◽  
Elba Hernández ◽  
Juan Martín Marqués ◽  
Felipe Schelotto ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Dna Amplification ◽  
Diagnostic Sensitivity ◽  
Analytical Sensitivity ◽  
Serum Samples ◽  
Initial Management ◽  
Negative Results ◽  
Acute Infections ◽  
Positive Results

Introduction: Assays based on DNA amplification can provide information that contributes to the initial management of patients with leptospirosis. However, these have not been adopted in Uruguay. Our aim was to evaluate the performance of the lipL32 real-time PCR (qPCR) for diagnosis of leptospirosis. Methodology: We analyzed by microscopic agglutination test (MAT) and lipL32 qPCR serum samples from 183 patients with suspected leptospirosis. To establish the analytical sensitivity of the qPCR, experimentally spiked samples with known amounts of Leptospira interrogans were analyzed. Results: The analytical sensitivity of the qPCR was 102 leptospires/mL. In 98 patients MAT results were negative meanwhile 85 showed positive reactions, revealing acute infections. Twenty six acute-phase sera of these 85 patients showed a positive signal by qPCR (diagnostic sensitivity 30%). In these patients the average time between onset of symptoms and collection of the first sample was 8 days. In patients with negative results for qPCR and positive MAT results (n=59) the average interval between onset of symptoms and collection of the first sample was 13 days. The qPCR did not yield false positive results. Conclusions: The qPCR had a lower diagnostic sensitivity than MAT and a higher cost. However, it allowed to make an early diagnosis in 26 patients. In patients with confirmed acute infections and negative results by qPCR, more than 8 days had elapsed between the onset of the illness and extraction of the first serum sample. Our data support that the qPCR from sera have clinical utility within the first week of illness.

Detection of Colorado Tick Fever viral RNA in acute human serum samples by a quantitative real-time RT-PCR assay

2007 ◽  
Vol 140 (1-2) ◽  
pp. 43-48 ◽  
Author(s):  
Amy J. Lambert ◽  
Olga Kosoy ◽  
Jason O. Velez ◽  
Brandy J. Russell ◽  
Robert S. Lanciotti
Keyword(s):  
Human Serum ◽  
Real Time ◽  
Viral Rna ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Serum Samples ◽  
Human Serum Samples

Counterimmunoelectrophoresis in determination of prostatic acid phosphatase in human serum.

1978 ◽  
Vol 24 (1) ◽  
pp. 140-142 ◽  
Author(s):  
A G Foti ◽  
J F Cooper ◽  
H Herschman
Keyword(s):  
Acid Phosphatase ◽  
Human Serum ◽  
Prostatic Cancer ◽  
Serum Proteins ◽  
Prostatic Acid Phosphatase ◽  
Serum Samples ◽  
Increased Activity ◽  
Positive Results

Abstract We evaluated counterimmunoelectrophoresis for use in measuring prostatic acid phosphatase in detection of prostatic cancer. After staining for acid phosphatase, we could detect as little as 0.3 ng of purified enzyme standard complexed with antibody by this technique. However, when serum samples were used as antigen, the method was less sensitive (1.5-2.0 ng) because some of the serum proteins migrate with the phosphatase and decrease the intensity of the stain for acid phosphatase. For this reason we could not detect the phosphatase in serum samples of normal persons; only patients with moderately (or greater) increased activity in their serum showed positive results. In contrast, by radioimmunoassay as little as 1.0 ng of the phosphatase can be detected in serum.

scholarly journals Evaluation of rapid, cassette immunochromatographic tests in the serological diagnosis of COVID-19

2021 ◽  
pp. 25-37
Author(s):  
Waldemar Rastawicki ◽  
Klaudia Płaza ◽  
Adam Pietrusiński
Keyword(s):  
Low Cost ◽  
Lateral Flow ◽  
Serological Diagnosis ◽  
Igg Antibodies ◽  
Serum Samples ◽  
Serological Tests ◽  
Negative Results ◽  
Lateral Flow Assays ◽  
Low Sensitivity ◽  
Positive Results

Introduction: Lateral flow assays (LFIA) are the technology behind low-cost, simple, rapid and portable detection devices popular in biomedicine. Lately, they are very common used in serodiagnosis of SARS-CoV-2 infections. The aim of the presented study was to assess the usefulness of selected LFIA in serological diagnosis of COVID-19. Methods: The usefulness of seven lateral flow assays in the serodiagnosis of COVID-19 was evaluated (VAZYME, DIAGNOSIS, PCL, INGEZIM, BIOSENSOR, ACCU-TELL, NOVAtest). The study used 107 serum samples obtained from 74 individuals with current SARS-CoV-2 infection confirmed by RT-PCR. The ELISA-IgG (Euroimmun) was used as the reference assay for sensitivity and specificity testing. Results: The highest percentage of positive results was obtained when searching for IgG antibodies with the NOVAtest (40.6%) and DIAGNOSIS (39.2%) sets and the lowest detection for the PCL set - 25.5%. In the case of searching for IgM antibodies in all sets, significantly lower percentages of positive results compared to the IgG class were recorded. In general, all lateral flow assays showed low sensitivity in relation to the Euroimmun ELISA-IgG. The DIAGNOSIS kit (64.5%) was characterized by the highest sensitivity, and the PCL kit was the lowest (38.7%). On the other hand, the specificity of all kits was very high, almost 100% in almost all cases. Conclusions: Lateral flow assays due to their low sensitivity are not suitable for quick diagnosis of the current SARS-CoV-2 infections and cannot be an alternative to genetic or even antigen tests. They may be used only to retrospectively test the presence of IgG antibodies. However, a negative results of LFIA in suspected disease or after vaccination should be confirmed by more sensitive serological tests.

scholarly journals Single-step label-free nanowell immunoassay accurately quantifies serum stress hormones within minutes

2021 ◽  
Vol 7 (27) ◽  
pp. eabf4401
Author(s):  
S. Reza Mahmoodi ◽  
Pengfei Xie ◽  
Daniel P. Zachs ◽  
Erik J. Peterson ◽  
Rachel S. Graham ◽  
...  
Keyword(s):  
Human Serum ◽  
Real Time ◽  
Limit Of Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Label Free ◽  
Serum Samples ◽  
Electrode Gap ◽  
Buffer Solution ◽  
Human Serum Samples ◽  
Free Cortisol

A non-faradaic label-free cortisol sensing platform is presented using a nanowell array design, in which the two probe electrodes are integrated within the nanowell structure. Rapid and low volume (≤5 μl) sensing was realized through functionalizing nanoscale volume wells with antibodies and monitoring the real-time binding events. A 28-well plate biochip was built on a glass substrate by sequential deposition, patterning, and etching steps to create a stack nanowell array sensor with an electrode gap of 40 nm. Sensor response for cortisol concentrations between 1 and 15 μg/dl in buffer solution was recorded, and a limit of detection of 0.5 μg/dl was achieved. Last, 65 human serum samples were collected to compare the response from human serum samples with results from the standard enzyme-linked immunosorbent assay (ELISA). These results confirm that nanowell array sensors could be a promising platform for point-of-care testing, where real-time, laboratory-quality diagnostic results are essential.

Rapid detection of Chlamydia/Chlamydophila group in samples collected from swine herds with and without reproductive disorders

2014 ◽  
Vol 17 (2) ◽  
pp. 367-369 ◽  
Author(s):  
K. Rypula ◽  
A. Kumala ◽  
P. Lis ◽  
K. Niemczuk ◽  
K. Płoneczka-Janeczko ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Rapid Detection ◽  
Complement Fixation Test ◽  
Complement Fixation ◽  
Fixation Test ◽  
Reproductive Disorders ◽  
Serum Samples ◽  
Swine Herds

Abstract The study was carried out in seven reproductive herds of pigs. In three of them reproductive disorders were observed. Three herds consisted of 10-50 and four consisted of 120-500 adult sows and they were called small and medium, respectively. Fifty-seven adult sows were randomly selected from herds. Serum samples were tested using the complement fixation test and swabs from both eyes and from the vaginal vestibule were examined using real-time PCR. All serum samples were negative. Infected sows were present in each of the study herds. In total, there were 28 positive samples (53%, 28/48) in real-time PCR in sows with reproductive disorders and 35 (53%, 35/66) in sows selected from herds without problems in reproduction. One isolate proved to be Chlamydophila pecorum, whereas all the remaining were Chamydia suis

Isolation and identification of Marek’s disease virus (MDV) from feather follicle epithelium and internal organs of diseased chickens in Dakahlia Governorate, Egypt

2019 ◽  
Vol 20 (2) ◽  
pp. 6-11
Author(s):  
Aly El-Kenawy ◽  
Mohamed El-Tholoth ◽  
Emad A
Keyword(s):  
Real Time ◽  
Disease Virus ◽  
Real Time Pcr ◽  
Marek's Disease Virus ◽  
Marek's Disease ◽  
Marek’S Disease Virus ◽  
Marek’S Disease ◽  
Field Samples ◽  
Feather Follicle ◽  
Positive Results

In the present study, a total of 16 samples including feather follicle epithelium, ovary, spleen and kidney (4 samples for each organ) were collected from diseased chicken flocks suspected to be infected with Marek’s disease virus (MDV) at Dakahlia Governorate, Egypt during the period from October 2016 to October 2017. Each sample was pooled randomly from three to five birds (90 to 360 days old). The isolation of the suspected virus from the collected samples was carried out via chorioallantoic membranes (CAMs) of 12 days old embryonated chicken eggs (ECEs). Three egg passages were carried out for each sample. Hyperimmune serum was prepared against standard MDV. MDV in both field and egg passaged samples (after 3rd passage) was identified by agar gel precipitation test (AGPT) and indirect fluorescence antibody test (IFAT). Molecular identification of virus was carried out by conventional polymerase chain reaction (PCR) and real- time PCR in four selected samples. The results revealed that 14 samples (87.5%) including 4 (100%) samples from feather follicle epithelium, ovary and kidney and 2 (50%) samples from spleen, showed positive results in virus isolation after 3rd passage. The positive results percentage by AGPT for field samples were 50% (8 out of 16 samples), while after the 3rd passage in ECEs were 37.5% (6 out of 16 samples) and the positive results percentage by IFAT for field samples were 62.5% (10 out of 16 samples), while after the 3rd passage in ECEs were 81.25 % (13 out of 16 samples). Viral nucleic acid was detected in all selected samples by conventional and real- time PCR. The results indicate that feather follicle epithelium is the best organ for MDV detection. IFAT is superior over AGPT in virus detection. Conventional and real - time PCR could be efficiently used for molecular detection of the virus.

Simultaneous Monitoring of Febuxostat and Uric Acid in Human Serum Samples Using the Direct Square-Wave Voltammetric Method

2019 ◽  
Vol 15 (6) ◽  
pp. 678-684
Author(s):  
Biljana Nigović ◽  
Jakov Vlak
Keyword(s):  
Uric Acid ◽  
Human Serum ◽  
Oxidase Inhibitor ◽  
Serum Samples ◽  
Boron Doped Diamond ◽  
Fast Analysis ◽  
Xanthine Oxidase Inhibitor ◽  
Voltammetric Method ◽  
Square Wave ◽  
Human Serum Samples

Background: High uric acid serum level, hyperuricemia, is now associated with many diseases such as gout, chronic kidney disease, hypertension, coronary artery disease and diabetes. Febuxostat is a novel selective xanthine oxidase inhibitor approved for the treatment of hyperuricemia. Objective: The aim of this study was to develop a first analytical method for the simultaneous determination of febuxostat and uric acid. Methods: An unmodified boron-doped diamond electrode provided concurrent quantitation of drug at low levels and uric acid, which has clinical significance in the diagnosis and therapy of hyperuricemia, at relatively high concentrations. The direct square-wave voltammetric method was applied to the analysis of both analytes in human serum samples. Results: Under the optimized conditions, the linear response of peak current on febuxostat concentration was achieved in the range from 7.5 × 10-7 to 3 × 10-5 M, while uric acid showed two linear ranges of 5 × 10-6 - 5 × 10-5 M and 5 × 10-5 - 2 × 10-4 M. The method was successfully utilised for quantification of both analytes in human serum samples. Good recoveries were obtained without interference from common inorganic cations and anions as well as glucose, dopamine, ascorbic and folic acids at concentrations expected in physiological conditions. Conclusion: The great benefits of developed method are fast analysis (only 7.5 s for run), low cost and simplicity of performance.

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