scholarly journals T2 Magnetic Resonance Assay-Based Direct Detection of Three Lyme Disease-Related Borrelia Species in Whole-Blood Samples

2017 ◽  
Vol 55 (8) ◽  
pp. 2453-2461 ◽  
Author(s):  
Jessica L. Snyder ◽  
Heidi Giese ◽  
Cheryl Bandoski-Gralinski ◽  
Jessica Townsend ◽  
Beck E. Jacobson ◽  
...  
Keyword(s):  
Magnetic Resonance ◽  
Real Time ◽  
Whole Blood ◽  
Limit Of Detection ◽  
Direct Detection ◽  
Erythema Migrans ◽  
Clinical Samples ◽  
Blood Samples ◽  
Content Type ◽  
Whole Blood Samples

ABSTRACTIn early Lyme disease (LD), serologic testing is insensitive and seroreactivity may reflect active or past infection. In this study, we evaluated a novel assay for the direct detection of three species ofBorreliaspirochetes in whole blood. The T2 magnetic resonance (T2MR) assay platform was used to amplifyBorreliaDNA released from intact spirochetes and to detect amplicon. Analytical sensitivity was determined from blood spiked with known concentrations of spirochetes, and the assay's limit of detection was found to be in the single-cell-per-milliliter range: 5 cells/ml forB. afzeliiand 8 cells/ml forBorrelia burgdorferiandBorrelia garinii. Clinical samples (n= 66) from confirmed or suspected early LD patients were also analyzed.B. burgdorferiwas detected using T2MR in 2/2 (100%) of blood samples from patients with confirmed early LD, based on the presence of erythema migrans and documentation of seroconversion or a positive real-time blood PCR. T2MR detectedB. burgdorferiin blood samples from 17/54 (31%) of patients with probable LD, based on the presence of erythema migrans without documented seroconversion or of documented seroconversion in patients with a compatible clinical syndrome but without erythema migrans. Out of 21 clinical samples tested by real-time PCR, only 1 was positive and 13 were negative with agreement with T2MR. An additional 7 samples that were negative by real-time PCR were positive with T2MR. Therefore, T2MR enables a low limit of detection (LoD) forBorreliaspp. in whole blood samples and is able to detectB. burgdorferiin clinical samples.

Surface acoustic wave immunosensor for real-time detection of hepatitis B surface antibodies in whole blood samples

2009 ◽  
Vol 24 (10) ◽  
pp. 3120-3125 ◽  
Author(s):  
Hun Joo Lee ◽  
Kak Namkoong ◽  
Eun Chol Cho ◽  
Christopher Ko ◽  
Jae Chan Park ◽  
...  
Keyword(s):  
Hepatitis B ◽  
Acoustic Wave ◽  
Real Time ◽  
Surface Acoustic Wave ◽  
Whole Blood ◽  
Blood Samples ◽  
Whole Blood Samples ◽  
Real Time Detection ◽  
Surface Antibodies

scholarly journals Flexible Micro Spring Array Device for High-Throughput Enrichment of Viable Circulating Tumor Cells

2014 ◽  
Vol 60 (2) ◽  
pp. 323-333 ◽  
Author(s):  
Ramdane A Harouaka ◽  
Ming-Da Zhou ◽  
Yin-Ting Yeh ◽  
Waleed J Khan ◽  
Avisnata Das ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
Whole Blood ◽  
Ex Vivo ◽  
Cell Damage ◽  
Flexible Structures ◽  
Capture Efficiency ◽  
Clinical Samples ◽  
Blood Samples ◽  
Whole Blood Samples

Abstract BACKGROUND The dissemination of circulating tumor cells (CTCs) that cause metastases in distant organs accounts for the majority of cancer-related deaths. CTCs have been established as a cancer biomarker of known prognostic value. The enrichment of viable CTCs for ex vivo analysis could further improve cancer diagnosis and guide treatment selection. We designed a new flexible micro spring array (FMSA) device for the enrichment of viable CTCs independent of antigen expression. METHODS Unlike previous microfiltration devices, flexible structures at the micro scale minimize cell damage to preserve viability, while maximizing throughput to allow rapid enrichment directly from whole blood with no need for sample preprocessing. Device performance with respect to capture efficiency, enrichment against leukocytes, viability, and proliferability was characterized. CTCs and CTC microclusters were enriched from clinical samples obtained from breast, lung, and colorectal cancer patients. RESULTS The FMSA device enriched tumor cells with 90% capture efficiency, higher than 104 enrichment, and better than 80% viability from 7.5-mL whole blood samples in <10 min on a 0.5-cm2 device. The FMSA detected at least 1 CTC in 16 out of 21 clinical samples (approximately 76%) compared to 4 out of 18 (approximately 22%) detected with the commercial CellSearch® system. There was no incidence of clogging in over 100 tested fresh whole blood samples. CONCLUSIONS The FMSA device provides a versatile platform capable of viable enrichment and analysis of CTCs from clinically relevant volumes of whole blood.

scholarly journals Development of a Broad-Range 23S rDNA Real-Time PCR Assay for the Detection and Quantification of Pathogenic Bacteria in Human Whole Blood and Plasma Specimens

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Paolo Gaibani ◽  
Mara Mariconti ◽  
Gloria Bua ◽  
Sonia Bonora ◽  
Davide Sassera ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Whole Blood ◽  
Extraction Method ◽  
Sample Type ◽  
Blood Samples ◽  
Human Whole Blood ◽  
Rdna Gene ◽  
Whole Blood Samples ◽  
Pcr Targeting

Molecular methods are important tools in the diagnosis of bloodstream bacterial infections, in particular in patients treated with antimicrobial therapy, due to their quick turn-around time. Here we describe a new broad-range real-time PCR targeting the 23S rDNA gene and capable to detect as low as 10 plasmid copies per reaction of targeted bacterial 23S rDNA gene. Two commercially available DNA extraction kits were evaluated to assess their efficiency for the extraction of plasma and whole blood samples spiked with different amount of eitherStaphylococcus aureusorEscherichia coli, in order to find the optimal extraction method to be used. Manual QIAmp extraction method with enzyme pre-treatment resulted the most sensitive for detection of bacterial load. Sensitivity of this novel assay ranged between 10 and 103 CFU per PCR reaction forE. coliandS. aureusin human whole blood samples depending on the extraction methods used. Analysis of plasma samples showed a 10- to 100-fold reduction of bacterial 23S rDNA in comparison to the corresponding whole blood specimens, thus indicating that whole blood is the preferential sample type to be used in this real-time PCR protocol. Our results thus show that the 23S rDNA gene represents an optimal target for bacteria quantification in human whole blood.

scholarly journals Detection of Genetically Modified Corn (Bt176) in Spiked Cow Blood Samples by Polymerase Chain Reaction and Immunoassay Methods

2005 ◽  
Vol 88 (2) ◽  
pp. 654-664 ◽  
Author(s):  
Laetitia Petit ◽  
Fabienne Baraige ◽  
Yves Bertheau ◽  
Philippe Brunschwig ◽  
Annick Diolez ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Whole Blood ◽  
Limit Of Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rrna Gene ◽  
Chain Reaction ◽  
Cry1ab Protein ◽  
Blood Samples ◽  
Polymerase Chain ◽  
Whole Blood Samples

Abstract The fate of DNA and protein transgenic sequences in products derived from animals fed transgenic crops has recently raised public interest. Sensitive molecular tests targeting the Bt176 genetic construct and the transgenic Cry1Ab protein were developed to determine whether plant sequences, especially transgenic sequences, are present in animal products. A protocol for total DNA extraction and purification from cow whole blood samples was first drawn up and assessed by spiking with known amounts of DNA from Bt176 maize. The limit of detection for transgenic sequences (35S promoter and Bt176-specific junction sequence) was determined by both the polymerase chain reaction–enzyme-linked immunosorbent assay (PCR–ELISA) and the 5′-nuclease PCR assay. Four additional PCR systems were built to substantiate the results. The first detects a mono-copy maize-specific sequence (ADH promoter). Two others target multi-copy sequences from plant nucleus (26S rRNA gene) and chloroplast (psaB gene). The last one, used as a positive control, targets a mono-copy animal sequence (αs1-casein gene). Both methods detected a minimum spiking at 25 copies of Bt176 maize/mL in 10 mL whole blood samples. The sandwich ELISA kit used detected down to 1 ng transgenic Cry1Ab protein/mL spiked whole blood.

Evaluation of an automated and highly sensitive real-time PCR assay for CMV DNA in whole blood samples

2006 ◽  
Vol 36 ◽  
pp. S26
Author(s):  
C. Deback ◽  
S. Akhavan ◽  
S. Blanc-Perrel ◽  
F. Dupuis ◽  
S. Schaffer ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Whole Blood ◽  
Pcr Assay ◽  
Blood Samples ◽  
Highly Sensitive ◽  
Whole Blood Samples

scholarly journals Stimulation of Bovine Whole-Blood Samples Cultured in Media Supplemented with Recombinant Interleukin-7 (IL-7) and IL-12 Extends the Life Span of the Gamma Interferon Assay To Detect Mycobacterium bovis-Infected Cattle

2016 ◽  
Vol 54 (9) ◽  
pp. 2315-2320 ◽  
Author(s):  
E. Gerace ◽  
P. Pasquali ◽  
B. Oesch ◽  
M. Falduto ◽  
F. Mandanici ◽  
...  
Keyword(s):  
Mycobacterium Bovis ◽  
Whole Blood ◽  
Gamma Interferon ◽  
Infected Cattle ◽  
Blood Samples ◽  
Content Type ◽  
Rpmi Medium ◽  
Interleukin 7 ◽  
Ifn Γ ◽  
Whole Blood Samples

The gamma interferon (IFN-γ) assay is widely used to measure cell-mediated immune (CMI) response for the early detection of tuberculosis infection. Processing whole-blood samples for CMI-based diagnostics is time sensitive and usually must occur within 8 h of collection to ensure optimal assay performance. In this study, we developed and tested a modified protocol, in which whole-blood samples fromMycobacterium bovis-infected cattle were diluted 1:1 in RPMI medium containing 0.3% fetal bovine serum (FBS) added or not to recombinant mouse interleukin-7 (rmIL-7) or rmIL-12, alone or in combination, and stored at 4°C. At 3 and 6 days postcollection, the diluted blood samples were adjusted to 10% FBS, dispensed into culture trays, stimulated with a bovine purified protein derivative fromM. bovis, and incubated at 37°C in 5% CO2in air. Plasma was removed and assayed for an IFN-γ response using bovine IFN-γ enzyme-linked immunosorbent assay (ELISA) (Bovigam). The results were then compared with those obtained from the conventional procedure. The IFN-γ responses of the samples stored up to 6 days postcollection in the supplemented RPMI medium were similar to those observed in the samples processed within 8 h after sampling, indicating that lymphocyte vitality and response were preserved. The addition of rmIL-7 and rmIL-12, alone or in combination, to culture medium can enhance lymphocyte survival and thus extends the time limit within which the IFN-γ assay can be applied as a diagnostic tool in bovine tuberculosis surveillance and eradication.

PX-13 Detection and quantitation of cytomegalovirus(CMV)DNA in EDTA whole blood samples by automated extraction and real-time PCR

2009 ◽  
Vol 46 ◽  
pp. S50
Author(s):  
B.D.A. Michelin ◽  
M. Bozic ◽  
S. Hammerschmidt ◽  
C. Homberg ◽  
D. Violan ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Whole Blood ◽  
Automated Extraction ◽  
Blood Samples ◽  
Whole Blood Samples
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