scholarly journals Recombinase polymerase amplification (RPA) with lateral flow detection for three Anaplasma species of importance to livestock health

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Andrea Salazar ◽  
Francisco M. Ochoa-Corona ◽  
Justin L. Talley ◽  
Bruce H. Noden
Keyword(s):  
Internal Control ◽  
Limit Of Detection ◽  
Lateral Flow ◽  
Serum Samples ◽  
Laboratory Equipment ◽  
Blood Samples ◽  
Gapdh Gene ◽  
Granulocytic Anaplasmosis ◽  
Positive Serum ◽  
Species Specific

AbstractAnaplasma marginale, A. ovis, and A. phagocytophilum are the causative agents of bovine anaplasmosis, ovine anaplasmosis, and granulocytic anaplasmosis, respectively. The gold standard for diagnosis of post-acute and long-term persistent infections is the serological cELISA, which does not discriminate between Anaplasma species and requires highly equipped laboratories and trained personnel. This study addresses the development of a rapid, isothermal, sensitive, species-specific RPA assays to detect three Anaplasma species in blood and cELISA A. marginale-positive serum samples. Three RPA primer and probe sets were designed targeting msp4 genes of each Anaplasma species and the internal control (GAPDH gene) for each assay. The limit of detection of gel-based or RPA-basic assays is 8.99 × 104 copies/µl = A. marginale, 5.04 × 106 copies/µl = A. ovis, and 4.58 × 103 copies/µl = A. phagocytophilum, and for each multiplex lateral flow or RPA-nfo assays is 8.99 × 103 copies/µl of A. marginale, 5.04 × 103 copies/µl of A. ovis, 4.58 × 103 copies/µl of A. phagocytophilum, and 5.51 × 103 copies/µl of internal control (GAPDH). Although none of the 80 blood samples collected from Oklahoma cattle were positive, the RPA-nfo assays detected all A. marginale cattle blood samples with varying prevalence rates of infection, 83% of the 24 cELISA A. marginale-positive serum samples, and all A. phagocytophilum cell culture samples. Overall, although early detection of three Anaplasma species was not specifically addressed, the described RPA technique represents an improvement for detection of three Anaplasma in regions where access to laboratory equipment is limited.

scholarly journals Detection of Candida albicans DNA from blood samples using a novel electrochemical assay

2011 ◽  
Vol 60 (4) ◽  
pp. 467-471 ◽  
Author(s):  
Alastair Muir ◽  
Gordon Forrest ◽  
John Clarkson ◽  
Alan Wheals
Keyword(s):  
Genomic Dna ◽  
Limit Of Detection ◽  
Rapid Identification ◽  
Clinical Samples ◽  
Electrochemical Assay ◽  
Blood Samples ◽  
Resistant Species ◽  
Life Threatening ◽  
Appropriate Therapy ◽  
Species Specific

The genus Candida contains a number of yeast species which are opportunistic pathogens and are associated with life-threatening infections in immunocompromised individuals. Provision of appropriate therapy relies on the rapid identification of the infecting species, and existing methods of identifying Candida species in clinical samples are time and resource intensive and are not always specific enough to differentiate between drug-susceptible and drug-resistant species. We have previously developed a system for the rapid detection of yeast pathogens in clinical samples using PCR followed by hybridization with a suite of five species-specific, electrochemically labelled DNA probes. The limit of detection of the assay was shown to be 37 fg (∼1 genome) per reaction using extracted genomic DNA. We carried out a study to test the limit of detection of one of the probes, CA PR3, using blood samples from a healthy donor that were spiked with genomic DNA or with C. albicans cells. Our results demonstrated a limit of detection of 37 fg (ml blood)−1 (∼1 genome ml−1) using extracted DNA or 10 c.f.u. (ml blood)−1 using C. albicans cells, indicating that the assay is capable of detecting C. albicans nucleic acid at levels that are encountered in clinical samples.

scholarly journals Lateral Flow Immunoassay for Visible Detection of Human Brucellosis Based on Blue Silica Nanoparticles

2021 ◽  
Vol 8 ◽  
Author(s):  
Lirui Ge ◽  
Dan Wang ◽  
Fengnan Lian ◽  
Jinbin Zhao ◽  
Yue Wang ◽  
...  
Keyword(s):  
Silica Nanoparticles ◽  
Covalent Bond ◽  
Lateral Flow ◽  
Clinical Diagnostics ◽  
Lateral Flow Immunoassay ◽  
Great Promise ◽  
Human Brucellosis ◽  
Serum Samples ◽  
Positive Serum ◽  
Health And Economic Development

Brucellosis is a highly contagious zoonosis chronic infectious disease with a strong latent capability to endanger human health and economic development via direct or indirect ways. However, the existing methods for brucellosis diagnosis are time-consuming and expensive as they require a tedious experimental procedure and a sophisticated experimental device and performance. To overcome these defects, it is truly necessary to establish a real-time, on-site, and rapid detection method for human brucellosis. Here, a lateral flow immunoassay (LFIA) with a rapid, sensitive, and alternative diagnostic procedure for human brucellosis with a high degree of accuracy was developed based on blue silica nanoparticles (SiNPs), Staphylococcal protein A (SPA), and surface Lipopolysaccharide of Brucella spp. (LPS), which can be applied for rapid and feasible detection of human brucellosis. To our knowledge, this is the first report that uses blue SiNPs as a signal probe of LFIA for the rapid diagnosis of human brucellosis. The precursor of blue SiNPs@SPA such as colorless SiNPs and blue SiNPs was synthesized at first and then coupled with SPA onto the surface of blue SiNPs by covalent bond to prepare blue SiNPs@SPA as a capture signal to catch the antibody in the brucellosis-positive serum. When SPA was combined with the antibodies in the brucellosis-positive serum, it was captured by LPS on the test line, forming an antigen–antibody sandwich structure, resulting in the T line turning blue. Finally, the results showed that it is acceptable to use blue SiNPs as visible labels of LFIA, and standard brucellosis serum (containing Brucella spp. antibody at 1,000 IU/ml) could be detected at a dilution of 10−5 and the detection limit of this method was 0.01 IU/ml. Moreover, it also demonstrated good specificity and accuracy for the detection of real human serum samples. Above all, the blue SiNPs-based LFIA that we developed provides a rapid, highly accurate, and inexpensive on-site diagnosis of human brucellosis, and shows great promise in clinical diagnostics for other diseases.

scholarly journals Archived dengue serum samples produced false-positive results in SARS-CoV-2 lateral flow-based rapid antibody tests

2021 ◽  
Vol 70 (6) ◽  
Author(s):  
Himadri Nath ◽  
Abinash Mallick ◽  
Subrata Roy ◽  
Soumi Sukla ◽  
Keya Basu ◽  
...  
Keyword(s):  
Dengue Virus ◽  
False Positive ◽  
Lateral Flow ◽  
Accurate Estimation ◽  
Serum Samples ◽  
Positive Serum ◽  
Rapid Tests ◽  
Antibody Tests ◽  
Positive Results ◽  
Rapid Antibody

Co-endemicity of SARS-CoV-2 and dengue virus (DV) infection is becoming a matter of serious concern as it has been already reported that antibodies (Ab) elicited by SARS-CoV-2 infection can produce false-positive results in dengue IgG and IgM rapid tests and vice versa. Here we communicate that five of thirteen DV antibody-positive serum samples from Kolkata, archived in 2017 (predating the COVID-19 outbreak), produced false-positive results in SARS-CoV-2 IgG/IgM lateral flow-based rapid tests. Our results emphasize the importance of implementing tests with higher specificity to conduct sero-surveillance for accurate estimation of SARS-CoV-2/DV prevalence in regions where both viruses now co-exist.

scholarly journals Rapid Detection of Hepatitis B Virus in Blood Samples Using a Combination of Polymerase Spiral Reaction With Nanoparticles Lateral-Flow Biosensor

2021 ◽  
Vol 7 ◽  
Author(s):  
Lin Lin ◽  
Jinshuai Guo ◽  
Haiyang Liu ◽  
Xiaofeng Jiang
Keyword(s):  
Limit Of Detection ◽  
Point Of Care ◽  
Diagnostic Technique ◽  
Lamp Assay ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Nucleic Acid Amplification ◽  
Analytical Sensitivity ◽  
Blood Samples ◽  
Nucleic Acid Amplification Technology

A rapid, highly sensitive, and robust diagnostic technique for point-of-care (PoC) testing can be developed using the combination of the nanoparticle-based lateral flow biosensors (LFB) and isothermal nucleic acid amplification technology. Here, we developed a polymerase spiral reaction (PSR) containing FITC-labeled DNA probes coupled with the nanoparticle-based LFB assay (PSR-LFB) to detect the amplified products to detect HBV visually. Under the optimized conditions, the PSR assay involved incubation of the reaction mixture for 20 min at 63°C, followed by visual detection of positive amplicons using LFB, which would generate a red test line based on the biotin/streptavidin interaction and immunoreactions, within 5 min. A cross-reactivity test revealed that the developed PSR-LFB assay showed good specificity for HBV and could distinguish HBV from other pathogenic microorganisms. For the analytical sensitivity, the limit of detection (LoD) of PSR-LFB assay was recorded as 5.4 copies/mL of HBV genomic DNA, which was ten-times more sensitive than qPCR and loop-mediated isothermal amplification (LAMP). Additionally, all the HBV-positive (29/82) samples, identified using ELISA, were also successfully detected by the PSR-LFB assay. We found that the true positive rate of the PSR-LFB assay was higher than that of qPCR (100 vs. 89.66%, respectively), as well as the LAMP assay (100 vs. 96.55%, respectively). Furthermore, the integrated procedure could be completed in 60 min, including the processing of the blood samples (30 min), an isothermal reaction (20 min), and result visualization (5 min). Thus, this PSR-LFB assay could be a potentially useful technique for PoC diagnosis of HBV in resource-limited countries.

scholarly journals Rapid, Sensitive, and Specific Lateral-Flow Immunochromatographic Point-of-Care Device for Detection of Herpes Simplex Virus Type 2-Specific Immunoglobulin G Antibodies in Serum and Whole Blood

2007 ◽  
Vol 15 (1) ◽  
pp. 159-163 ◽  
Author(s):  
Elisabeth I. Laderman ◽  
Emma Whitworth ◽  
Erickson Dumaual ◽  
Mark Jones ◽  
Andrew Hudak ◽  
...  
Keyword(s):  
Whole Blood ◽  
Point Of Care ◽  
Cross Reactivity ◽  
Serum Samples ◽  
Blood Samples ◽  
Positive Serum ◽  
Whole Blood Samples ◽  
Simplex Virus ◽  
Sensitivity Specificity

ABSTRACT Herpes simplex virus type 2 (HSV-2) is a common human pathogen that can cause a variety of clinical manifestations in humans. In order to provide near-patient results to allow for faster counseling and treatment, a rapid point-of-care test that is accurate and simple to use is desirable. Here, we describe the development and evaluation of an HSV-2 immunoglobulin G (IgG)-specific antibody lateral-flow immunochromatographic assay (LFIA) based on colloidal gold nanoparticles. A total of 359 serum samples and 100 whole-blood samples were tested in the newly developed HSV-2 LFIA. Serum results were compared to those from the HerpeSelect HSV-2 enzyme-linked immunosorbent assay (ELISA), and whole-blood sample results were compared to those of both ELISA and HerpeSelect HSV-1 and -2 immunoblotting (IB). The sensitivity of the HSV-2 LFIA compared to that of the HerpeSelect ELISA was 100% (89/89), and the specificity was 97.3% (257/264). Cross-reactivity with HSV-1 IgG-positive serum samples was observed in 2.6% (5/196) of samples, 2.9% (1/34) for rubella virus, and 6.2% (1/16) for Epstein-Barr virus. No cross-reactivity in varicella-zoster virus or cytomegalovirus IgG-positive serum samples was observed. No interference was observed from bilirubin-, triglyceride-, albumin-, or hemoglobin-spiked samples. The concordance of the LFIA results between capillary whole blood, EDTA-treated venous whole blood, heparin-treated venous whole blood, and serum was 99% (99/100). In conclusion, the LFIA for HSV-2 IgG-specific antibodies demonstrated excellent sensitivity, specificity, and concordance for both serum and whole-blood samples compared to the sensitivity, specificity, and concordance of both HSV-2 ELISA and IB.

scholarly journals Rapid, Sensitive, and Specific Lateral-Flow Immunochromatographic Device To Measure Anti-Anthrax Protective Antigen Immunoglobulin G in Serum and Whole Blood

2006 ◽  
Vol 13 (5) ◽  
pp. 541-546 ◽  
Author(s):  
Raymond E. Biagini ◽  
Deborah L. Sammons ◽  
Jerome P. Smith ◽  
Barbara A. MacKenzie ◽  
Cynthia A. F. Striley ◽  
...  
Keyword(s):  
Internal Control ◽  
Immunoglobulin G ◽  
Whole Blood ◽  
Protective Antigen ◽  
Lateral Flow ◽  
Enzyme Linked Immunosorbent Assay ◽  
Blood Samples ◽  
Test Kit ◽  
Anthrax Protective Antigen ◽  
Whole Blood Samples

ABSTRACT Evidence from animals suggests that anti-anthrax protective antigen (PA) immunoglobulin G (IgG) from vaccination with anthrax vaccine adsorbed (AVA) is protective against Bacillus anthracis infection. Measurement of anti-PA IgG in human sera can be performed using either enzyme-linked immunosorbent assay or fluorescent covalent microsphere immunoassay (ELISA) (R. E. Biagini, D. L. Sammons, J. P. Smith, B. A. MacKenzie, C. A. Striley, V. Semenova, E. Steward-Clark, K. Stamey, A. E. Freeman, C. P. Quinn, and J. E. Snawder, Clin. Diagn. Lab. Immunol. 11:50-55, 2004). Both these methods are laboratory based. We describe the development of a rapid lateral-flow immunochromatographic assay (LFIA) test kit for the measurement of anti-PA IgG in serum or whole-blood samples (30-μl samples) using colloidal gold nanoparticles as the detection reagent and an internal control. Using sera from 19 anthrax AVA vaccinees (anti-PA IgG range, 2.4 to 340 μg/ml) and 10 controls and PA-supplemented whole-blood samples, we demonstrated that the LFIA had a sensitivity of approximately 3 μg/ml anti-PA IgG in serum and ∼14 μg/ml anti-PA IgG in whole blood. Preabsorption of sera with PA yielded negative anti-PA LFIAs. The diagnostic sensitivity and specificity of the assay were 100% using ELISA-measured anti-PA IgG as the standard. This kit has utility in determining anti-PA antibody reactivity in the sera of individuals vaccinated with AVA or individuals with clinical anthrax.

Ferromagnetic Resonance Biosensor for Homogeneous and Volumetric Detection of DNA

2017 ◽  
Author(s):  
Bo Tian ◽  
Peter Svedlindh ◽  
Mattias Strömberg ◽  
Erik Wetterskog
Keyword(s):  
Magnetic Nanoparticles ◽  
Ferromagnetic Resonance ◽  
Limit Of Detection ◽  
Point Of Care ◽  
Rolling Circle Amplification ◽  
Resonance Field ◽  
Isothermal Amplification ◽  
Detection Sensitivity ◽  
Serum Samples ◽  
Rolling Circle

In this work, we demonstrate for the first time, a ferromagnetic resonance (FMR) based homogeneous and volumetric biosensor for magnetic label detection. Two different isothermal amplification methods, <i>i.e.</i>, rolling circle amplification (RCA) and loop-mediated isothermal amplification (LAMP) are adopted and combined with a standard electron paramagnetic resonance (EPR) spectrometer for FMR biosensing. For RCA-based FMR biosensor, binding of RCA products of a synthetic Vibrio cholerae target DNA sequence gives rise to the formation of aggregates of magnetic nanoparticles. Immobilization of nanoparticles within the aggregates leads to a decrease of the net anisotropy of the system and a concomitant increase of the resonance field. A limit of detection of 1 pM is obtained with an average coefficient of variation of 0.16%, which is superior to the performance of other reported RCA-based magnetic biosensors. For LAMP-based sensing, a synthetic Zika virus target oligonucleotide is amplified and detected in 20% serum samples. Immobilization of magnetic nanoparticles is induced by their co-precipitation with Mg<sub>2</sub>P<sub>2</sub>O<sub>7</sub> (a by-product of LAMP) and provides a detection sensitivity of 100 aM. The fast measurement, high sensitivity and miniaturization potential of the proposed FMR biosensing technology makes it a promising candidate for designing future point-of-care devices.<br>

A Point of Care Lateral Flow Assay for Rapid and Colorimetric Detection of Interleukin 6 and Perspectives in Bedside Diagnostics

2020 ◽  
Author(s):  
Carla Eiras
Keyword(s):  
Interleukin 6 ◽  
Limit Of Detection ◽  
Inflammatory Diseases ◽  
Point Of Care ◽  
Colorimetric Detection ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Aggregation Assay ◽  
Before And After ◽  
Bedside Diagnosis

Interleukin-6 (IL-6) is a multifunctional cytokine and high bloodstream levels of which have been associated with severe inflammatory diseases, such as dengue fever, sepsis, various cancers, and visceral leishmaniasis (VL). Rapid tests for the quantification of IL-6 would be of great assistance for the bedside diagnosis and treatment of diseases such as VL. We have developed a lateral flow assay (LFA) for rapid and colorimetric IL-6 detection, consisting of anti-IL-6 antibodies conjugated to gold nanoparticles (AuNPs). The optimal concentration of anti-IL-6 used in the conjugate was determined to be 800.0 μg/mL, based on an aggregation assay using LFA. A linear relationship between IL-6 standard concentration and color intensity was observed after 20 min, with a linear range between 1.25 ng/mL and 9,000 ng/mL. The limit of detection for this method was estimated a t0.38 ng/mL. The concentration of IL-6 in five patients with severe VL was measured using LFA, and the results were consistent with those obtained using the cytometric bead array (CBA) method. A thorough analysis of the LFA membranes’ surface morphology, before and after sample contact, was performed using atomic force microscopy (AFM).The prototype described here is still being tested and improved, but this LFA will undoubtedly be of great help in the clinical quantification of IL-6.

A Point of Care Lateral Flow Assay for Rapid and Colorimetric Detection of Interleukin 6 and Perspectives in Bedside Diagnostics

2020 ◽  
Author(s):  
Carla Eiras
Keyword(s):  
Interleukin 6 ◽  
Limit Of Detection ◽  
Inflammatory Diseases ◽  
Point Of Care ◽  
Colorimetric Detection ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Aggregation Assay ◽  
Before And After ◽  
Bedside Diagnosis

Interleukin-6 (IL-6) is a multifunctional cytokine and high bloodstream levels of which have been associated with severe inflammatory diseases, such as dengue fever, sepsis, various cancers, and visceral leishmaniasis (VL). Rapid tests for the quantification of IL-6 would be of great assistance for the bedside diagnosis and treatment of diseases such as VL. We have developed a lateral flow assay (LFA) for rapid and colorimetric IL-6 detection, consisting of anti-IL-6 antibodies conjugated to gold nanoparticles (AuNPs). The optimal concentration of anti-IL-6 used in the conjugate was determined to be 800.0 μg/mL, based on an aggregation assay using LFA. A linear relationship between IL-6 standard concentration and color intensity was observed after 20 min, with a linear range between 1.25 ng/mL and 9,000 ng/mL. The limit of detection for this method was estimated at a t0.38 ng/mL. The concentration of IL-6 in five patients with severe VL was measured using LFA, and the results were consistent with those obtained using the cytometric bead array (CBA) method. A thorough analysis of the LFA membranes’ surface morphology, before and after sample contact, was performed using atomic force microscopy (AFM). The prototype described here is still being tested and improved, but this LFA will undoubtedly be of great help in the clinical quantification of IL-6.

scholarly journals PRRSV detection by qPCR in processing fluids and serum samples collected in a positive stable breeding herd following mass vaccination of sows with a modified live vaccine

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
A. Lebret ◽  
P. Berton ◽  
V. Normand ◽  
I. Messager ◽  
N. Robert ◽  
...  
Keyword(s):  
The United States ◽  
Live Vaccine ◽  
Mass Vaccination ◽  
Respiratory Syndrome Virus ◽  
Serum Samples ◽  
Blood Samples ◽  
Stability Monitoring ◽  
Breeding Herd ◽  
Modified Live Vaccine ◽  
Processing Fluid

AbstractIn the last two decades, in France, Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) stabilization protocols have been implemented using mass vaccination with a modified live vaccine (MLV), herd closure and biosecurity measures. Efficient surveillance for PRRSV is essential for generating evidence of absence of viral replication and transmission in pigs. The use of processing fluid (PF) was first described in 2018 in the United States and was demonstrated to provide a higher herd-level sensitivity compared with blood samples (BS) for PRRSV monitoring. In the meantime, data on vertical transmission of MLV viruses are rare even as it is a major concern. Therefore, veterinarians usually wait for several weeks after a sow mass vaccination before starting a stability monitoring. This clinical study was conducted in a PRRSV-stable commercial 1000-sow breed-to-wean farm. This farm suffered from a PRRS outbreak in January 2018. After implementing a stabilisation protocol, this farm was controlled as stable for more than 9 months before the beginning of the study. PF and BS at weaning were collected in four consecutive batches born after a booster sow mass MLV vaccination. We failed to detect PRRSV by qPCR on PF and BS collected in a positive-stable breeding herd after vaccination with ReproCyc® PRRS EU (Boehringer Ingelheim, Ingelheim, Germany).

Sign in / Sign up

Export Citation Format

Share Document