scholarly journals False-Positive Results of Plasma PCR for Cytomegalovirus DNA due to Delayed Sample Preparation

2000 ◽  
Vol 38 (9) ◽  
pp. 3249-3253 ◽  
Author(s):  
Peter Schäfer ◽  
Werner Tenschert ◽  
Matthias Schröter ◽  
Kai Gutensohn ◽  
Rainer Laufs
Keyword(s):  
False Positive ◽  
Room Temperature ◽  
Significant Risk ◽  
Median Number ◽  
Plasma Samples ◽  
Cmv Infection ◽  
Latently Infected ◽  
Positive Results ◽  
Native Plasma ◽  
Cmv Dnaemia

Positive results by cytomegalovirus (CMV) PCR of plasma are considered predictive of active CMV infection in kidney allograft recipients. To assess whether contamination with leukocyte-derived CMV DNA can distort the results, aliquots of whole-blood samples from 60 CMV immunoglobulin G-positive patients with leukocyte CMV DNAemia were stored for up to 24 h at room temperature (RT) and at 4°C before plasma preparation. Native and ultrafiltered plasma samples were tested by CMV and β-globin PCRs. Among 30 latently infected patients (negative for CMV pp65 antigens), low baseline rates (10%) and levels (median number of copies, 10 [per 10 μl]) of CMV plasma DNAemia in native plasma samples increased significantly over time (after 4 h at RT, 37% [P < 0.001]; median number of copies, 45 [P < 0.001]). Similar effects were found during storage at 4°C. Ultrafiltration reduced the levels of CMV plasma DNAemia, but by 6 h of storage the levels were significantly elevated as well. CMV and β-globin DNA kinetics in plasma were parallel. In contrast, 30 actively infected patients (pp65 positive) had high baseline rates (87% in native samples) and levels (median number of copies, 75) of CMV plasma DNAemia. No significant effects of storage or ultrafiltration and no concordance with β-globin DNA kinetics were seen. In conclusion, delayed preparation of plasma samples bears a significant risk of false-positive CMV PCR results, probably due to leukocyte lysis. This has important implications in the clinical setting and for PCR standardization.

Solving the problem of antibody interference in commercial "sandwich"-type immunoassays of carcinoembryonic antigen.

1989 ◽  
Vol 35 (1) ◽  
pp. 146-151 ◽  
Author(s):  
H J Hansen ◽  
G LaFontaine ◽  
E S Newman ◽  
M K Schwartz ◽  
A Malkin ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Carcinoembryonic Antigen ◽  
False Positive ◽  
Room Temperature ◽  
Murine Monoclonal Antibody ◽  
Heat Extraction ◽  
Normal Individuals ◽  
Sandwich Type ◽  
Commercial Enzyme Immunoassay ◽  
Positive Results

Abstract We evaluated the effect of human anti-murine antibodies (HAMA) on apparent concentrations of carcinoembryonic antigen (CEA) as measured in serum with commercial enzyme immunoassay (EIA) kits manufactured by Abbot ("two-step" double monoclonal antibody assay), Roche, and Hybritech (room-temperature protocol). In sera from patients given parenteral murine monoclonal antibody for experimental diagnostic and immunotherapy studies, HAMA titers were determined with Immunomedics' "ImmuSTRIP HAMA-EIA" kit reagents. "True" CEA titers were established by using the ImmuCEA/MA-EIA and heat-extraction to destroy HAMA before assay for CEA. The concordance of the ImmuCEA/MA assay with the Abbott and Roche CEA EIAs was established with sera from normal individuals and from patients who had not received parenteral injections of murine monoclonal antibody. At high (100 mg/L) concentrations of HAMA, false-positive results were observed with all three kits. The Hybritech and Roche assays were more sensitive to interference by HAMA than was the Abbott CEA-EIA, false-positive results being observed at HAMA concentrations between 1 and 10 mg/L. Similar sensitivity of the three kits to interference by primate anti-MAb sera was demonstrated. Use of primate anti-MAb sera to create controls with HAMA activity and of analyte is recommended to evaluate MAb assays for potential HAMA interference and for use to devise methods to eliminate HAMA interference.

scholarly journals Utility of Major Leukocyte Subpopulations for Monitoring Secondary Cytomegalovirus Infections in Renal-Allograft Recipients by PCR

1998 ◽  
Vol 36 (4) ◽  
pp. 1008-1014 ◽  
Author(s):  
Peter Schäfer ◽  
Werner Tenschert ◽  
Liana Cremaschi ◽  
Kai Gutensohn ◽  
Rainer Laufs
Keyword(s):  
Renal Allograft ◽  
Antiviral Treatment ◽  
Plasma Samples ◽  
Cmv Infection ◽  
Active Infection ◽  
Ganciclovir Therapy ◽  
Copy Numbers ◽  
Pp65 Antigen ◽  
Dna Copy Numbers ◽  
Positive Results

The feasibility of the major peripheral blood leukocyte (PBL) subsets for use in qualitative and quantitative PCR to monitor secondary cytomegalovirus (CMV) infection and ganciclovir therapy was assessed with 188 blood samples derived from 40 CMV immunoglobulin G-positive renal-allograft recipients. In pp65 antigen-positive patients all leukocyte fractions, but only 79.5% of plasma preparations, were PCR positive. In pp65 antigen-negative samples from patients after antiviral treatment only 7.3% of polymorphonuclear cell (PMNL) samples, but 81.8% of peripheral blood mononuclear cells (PBMC), and 10.9% of plasma samples remained PCR positive. Similarly, in patients with latent infections only 5.0% of PMNL, but 51.7% of PBMC preparations, and 8.0% of plasma samples were PCR positive. Regarding patients with active CMV infection, CMV DNA copy numbers in PMNL correlated significantly with pp65 antigen-positive cell counts before and after onset of ganciclovir therapy. Significant differences in CMV DNA copy numbers in PMNL and plasma were observed (i) between patients with symptomatic infection and those with asymptomatic infection and (ii) between patients with active infection and those with latent infection. In contrast, PBMC harbored equally low CMV DNA levels both in patients with active infection and those with latent infections, and no decline of CMV DNA load in PBMC was observed during antiviral treatment. We conclude that detection of CMV DNA in PMNL, not in PBMC, is associated with active infections and is more sensitive than detection of CMV DNA in plasma. Negative PCR results for PMNL after antiviral therapy indicate recovery, and fewer unwanted positive results occur compared to PBMC and plasma. Therefore, purified PMNL should be preferred for analysis by qualitative CMV PCR to avoid unwanted positive results. The CMV DNA load in PBMC compared with that in PMNL is negligible during active infection, so mixed PBL are sufficient for use in quantitative PCR.

scholarly journals The Risk of False-Positive Serological Results for Paratuberculosis in Mycobacterium bovis-Infected Cattle

Pathogens ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1054
Author(s):  
Anna Didkowska ◽  
Monika Krajewska-Wędzina ◽  
Daniel Klich ◽  
Kinga Prolejko ◽  
Blanka Orłowska ◽  
...  
Keyword(s):  
Mycobacterium Bovis ◽  
False Positive ◽  
Significant Risk ◽  
Elisa Test ◽  
Cross Reactivity ◽  
Economic Losses ◽  
Infected Cattle ◽  
Increased Risk ◽  
Gross Lesions ◽  
Positive Results

Both bovine tuberculosis (BTB) and paratuberculosis (paraTB) continue to cause significant economic losses in cattle breeding; in addition, their etiological agents have zoonotic potential. Although the diagnostics of both diseases are still being improved, problems still remain, such as the potential for cross-reactivity to the antigens used in tests. The aim of the present study was to confirm whether animals known to harbor Mycobacterium bovis antibodies are at increased risk of yielding positive results in paraTB serotesting and, additionally, to verify the accuracy of three commonly used methods for confirming M. bovis infection: ELISA, the tuberculin skin test (TST), and the presence of gross lesions. Material was collected from 98 dairy cattle suspected of BTB due to TST-positive results. During postmortem examination, gross lesions were assessed visually. Blood, lymph nodes, and TB-suspected organs were collected. Serum was obtained from the collected blood and tested serologically for TB and paraTB. The tissues underwent standard microbiological testing for M. tuberculosis complex. Among the 98 TST-positive individuals, tuberculous gross lesions were detected in 57 (58.1%), MTBC were isolated in 83 (84.7%), and the ELISA test was positive for 21 (21.4%). None of the lesions characteristic for paraTB were detected. The chance of obtaining a positive TB result by ELISA was seven times higher using the ELISA-paraTB method; hence, there is a significant risk of obtaining false-positive serological results for paraTB in M. bovis-infected cattle. However, the hypothesis that infection of M. bovis or prior TST performance may have boosted the host immune response and therefore increased the sensitivity of the paraTB-ELISA cannot be excluded.

Critical Evaluation of Impedance Phlebography for the Diagnosis of Deep Vein Thrombosis

1974 ◽  
Vol 31 (02) ◽  
pp. 273-278
Author(s):  
Kenneth K Wu ◽  
John C Hoak ◽  
Robert W Barnes ◽  
Stuart L Frankel
Keyword(s):  
Deep Vein Thrombosis ◽  
False Positive ◽  
False Negative ◽  
Critical Evaluation ◽  
Original Method ◽  
Vein Thrombosis ◽  
Negative Results ◽  
False Negative Results ◽  
Deep Vein ◽  
Positive Results

SummaryIn order to evaluate its daily variability and reliability, impedance phlebography was performed daily or on alternate days on 61 patients with deep vein thrombosis, of whom 47 also had 125I-fibrinogen uptake tests and 22 had radiographic venography. The results showed that impedance phlebography was highly variable and poorly reliable. False positive results were noted in 8 limbs (18%) and false negative results in 3 limbs (7%). Despite its being simple, rapid and noninvasive, its clinical usefulness is doubtful when performed according to the original method.

Comparison of two rapid tests for the detection of legionellaceae in water: a microbiological-immunological method and a commercial gene-probe testkit

1995 ◽  
Vol 31 (5-6) ◽  
pp. 403-406 ◽  
Author(s):  
E. Frahm ◽  
U. Obst
Keyword(s):  
Monoclonal Antibodies ◽  
Conventional Method ◽  
Standard Method ◽  
False Positive ◽  
Gene Probe ◽  
Immunological Method ◽  
Probe Test ◽  
Rapid Tests ◽  
Positive Results

Two recently developed Legionella detection tests, a microbiological-immunological method based on monoclonal antibodies (carried out as a colony-blot assay) and a commercial gene-probe testkit (the EnvironAmp Legionella Kit), are compared with the standard method. The colony-blot assay is faster than the conventional method; the gene-probe test is much faster still and is the most sensitive, but in consequence is at greater risk of false-positive results.

False-Positive Ethanol Level in Urine and Plasma Samples of a Resuscitated Infant

2020 ◽  
Author(s):  
B Lefrère ◽  
D Wohrer ◽  
C Godefroy ◽  
M Soichot ◽  
A Mihoubi ◽  
...  
Keyword(s):  
False Positive ◽  
Ethanol Concentration ◽  
Enzymatic Assay ◽  
Complex Congenital Heart Disease ◽  
Plasma Samples ◽  
Limit Of Quantification ◽  
Chromatography Method ◽  
Ethanol Level

Abstract We report the case of an 11-month-old male infant with a complex congenital heart disease who was admitted in the intensive care unit following cardiorespiratory arrest at home. Toxicological urine screening reported an ethanol concentration of 0.65 g/L using an enzymatic assay, without suspicion of alcohol intake; a significant amount of ethanol concentration was found in two plasma samples using the same enzymatic assay. Plasma and urine ethanol concentrations were below the limit of quantification (LOQ) when tested using a gas chromatography method. Urine ethanol level was also below the LOQ when tested by enzymatic assay after an initial urine ultrafiltration. These results confirmed our suspicion of matrix interference due to elevated lactate and lactate dehydrogenase levels interfering in the enzymatic assay. This analytical interference, well-known in postmortem samples, extensively studied in vitro, has been rarely reported in vivo, especially in children. To the best of our knowledge, this case is only the sixth one reported in an infant’s plasma and the first initially discovered from urine. Indeed, as for ethanol, this last matrix has not been studied in the context of this artifact that may induce false-positive ethanol results while seeking a diagnosis in life-threatening or fatal situations that are potentially subject to forensic scrutiny. In parallel to a synthetic literature review, we propose a simple, informative decision tree, in order to help health professionals suspecting a false-positive result when performing an ethanol assay.

scholarly journals The Effect of Genomic DNA Contamination on the Detection of Circulating Long Non-Coding RNAs: The Paradigm of MALAT1

Diagnostics ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1160
Author(s):  
Athina N. Markou ◽  
Stavroula Smilkou ◽  
Emilia Tsaroucha ◽  
Evi Lianidou
Keyword(s):  
False Positive ◽  
Tissue Samples ◽  
Dna Contamination ◽  
Dnase Treatment ◽  
Circular Dnas ◽  
Non Coding Rnas ◽  
Nsclc Patients ◽  
Treatment Step ◽  
Primary Tissue ◽  
Positive Results

The presence of contaminating gDNA in RNA preparations is a frequent cause of false positives in RT-PCR-based analysis. However, in some cases, this cannot be avoided, especially when there are no exons–intron junctions in the lncRNA sequences. Due to the lack of exons in few of long noncoding RNAs (lncRNAs) and the lack of DNAse treatment step in most studies reported so far, serious questions are raised about the specificity of lncRNA detection and the potential of reporting false-positive results. We hypothesized that minute amounts of gDNA usually co-extracted with RNA could give false-positive signals since primers would specifically bind to gDNA due to the lack of junction. In the current study, we evaluated the effect of gDNA and other forms of DNA like extrachromosomal circular DNAs (eccDNAs) contamination and the importance of including a DNAse treatment step on lncRNAsexpression.As a model, we have chosen as one of the most widely studied lncRNAs in cancer namely MALAT1, which lacks exons. When we tested this hypothesis in plasma and primary tissue samples from NSCLC patients, our findings clearly indicated that results on MALAT1 expression are highly affected by the presence of DNA contamination and that the DNAse treatment step is absolutely necessary to avoid false positive results.

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