Focal adhesions (FAs) and associated actin stress fibers (SFs) form a complex mechanical system that mediates bidirectional interactions between cells and their environment.
This linked network is essential for mechanosensing, force production and force transduction, thus directly governing cellular processes like polarization, migration and extracellular matrix remodeling.
We introduce a tool for fast and robust coupled analysis of both FAs and SFs named the Focal Adhesion Filament Cross-correlation Kit (FAFCK). Our software can detect and record location, axes lengths, area, orientation, and aspect ratio of focal adhesion structures as well as the location, length, width and orientation of actin stress fibers.
This enables users to automate analysis of the correlation of FAs and SFs and study the stress fiber system in a higher degree, pivotal to accurately evaluate transmission of mechanocellular forces between a cell and its surroundings.
The FAFCK is particularly suited for unbiased and systematic quantitative analysis of FAs and SFs necessary for novel approaches of traction force microscopy that uses the additional data from the cellular side to calculate the stress distribution in the substrate.
For validation and comparison with other tools, we provide datasets of cells of varying quality that are labelled by a human expert.
Datasets and FAFCK are freely available as open source under the GNU General Public License.
Comparison of performance between Fast Track Diagnostics Respiratory Kit and the CDC global reference laboratory for influenza rRT-PCR panel for detection of influenza A and influenza B
Background: Reliable diagnostics are a key to identifying influenza infections.
Objectives: Our objectives were to describe the detection of influenza among severe acute respiratory infection (SARI) cases, to compare test results from the Fast Track Diagnostics (FTD) Kit for influenza detection to the Centers for Disease Control (CDC) human influenza virus detection and characterization panel, and to assess seasonality of influenza in Burkina Faso.
Methods: Nasopharyngeal and oropharyngeal specimens from SARI cases (hospitalized patients with fever, cough, and onset in the previous 10 days) were tested using the FTD-33 Kit and the CDC rRT-PCR influenza assays. We assessed sensitivity and specificity of the FTD-33 Kit for detecting influenza A, influenza B, and the influenza A(H1N1)pdm09 strain using the CDC human influenza rRT-PCR panel as the gold standard.
Results: From December 2016 to February 2019, 1706 SARI cases were identified, 1511 specimens were tested, and 211 were positive for influenza A (14.0%) and 100 for influenza B (6.6%) by either assay.
Higher influenza circulation occurred between November and April with varying peaks of influenza A and influenza B. Sensitivity of the FTD-33 assay was 91.9% for influenza A, 95.7% for influenza B, and 93.8% for A(H1N1)pdm09 subtype. Specificity was over 99% for all three tests.
Conclusions: Our study indicates that Burkina Faso has one peak of influenza each year which is similar to the Northern Hemisphere and differs from other countries in West Africa. We found high concordance of influenza results between the two assays indicating FTD-33 can be used to reliably detect influenza among SARI cases.
Keywords: Burkina Faso; West Africa; diagnostics; influenza; severe acute respiratory infections.
Sustainable and fast saliva-based COVID-19 virus diagnosis kit using a novel GO-decorated Au/FBG sensor
Monitoring the COVID-19 virus through patients’ saliva is a favorable non-invasive specimen for diagnosis and infection control. In this study, salivary samples of COVID-19 patients collected from 6 patients with the median age of 58.5 years, ranging from 34 to 72 years (2 females and 4 males) were analyzed using an Au/fiber Bragg grating (FBG) probe decorated with GO. The probe measures the prevalence of positivity in saliva and the association between the virus density and changes to sensing elements.
When the probe is immersed in patients’ saliva, deviation of the detected light wavelength and intensity from healthy saliva indicate the presence of the virus and confirms infection.
For a patient in the hyperinflammatory phase of desease, who has virus density of 1.2 × 108 copies/mL in saliva, the maximum wavelength shift and intensity changes after 1600 s were shown to be 1.12 nm and 2.01 dB, respectively. While for a patient in the early infection phase with 1.6 × 103 copies/mL, these values were 0.98 nm and 1.32 dB.
The precise and highly sensitive FBG probe proposed in this study was found a reliable tool for quick detection of the COVID-19 virus within 10 s after exposure to patients’ saliva in any stage of the disease.
Comparison of the Luminex xTAG Respiratory Viral Panel Fast v2 Assay With Anyplex II RV16 Detection Kit and AdvanSure RV Real-Time RT-PCR Assay for the Detection of Respiratory Viruses.
The accurate and rapid identification of the causative viruses is important for the timely diagnosis and management of respiratory infections.
Multiplex molecular diagnostic techniques have been widely adopted to detect respiratory viruses.
We compared the results of a newly upgraded, multiplex, molecular bead-based respiratory viral panel (RVP) assay with the results of Anyplex II RV16 detection kit and AdvanSure RV real-time RT-PCR assay.
We tested 254 respiratory specimens and cultured viral strains using the Luminex xTAG RVP Fast v2 assay (Luminex Molecular Diagnostics, Canada) and Anyplex II RV16 detection kit and compared the results. Specimens showing discordant results between the two assays were tested with a AdvanSure RV real-time RT-PCR assay.
Of the 254 respiratory specimens, there was total agreement in the results between the xTAG RVP Fast v2 assay and the other real-time PCR assay in 94.1-100% of the specimens. The agreement levels were relatively low (94.1-97.6%) for specimens of adenovirus, coronavirus NL63, and parainfluenza type 3. In comparison to the other assay, the xTAG RVP Fast v2 assay detected a higher number of parainfluenza type 3 (4 cases) and metapneumovirus (9 cases).
The xTAG RVP Fast v2 assay showed comparable capabilities compared with the other assays; it will be useful for identifying respiratory viral infections in patients with respiratory symptoms. Clinicians should be aware of the characteristics of the assays they use, since different assays show different detectability for each virus.
Validation of the Applied Biosystems 7500 Fast Instrument for the Detection of Salmonellae with SureTect Salmonella Species PCR Kit.
The Thermo Scientific SureTect™ Salmonella species real-time PCR assay is a rapid alternative method designed for the detection of salmonellae in a wide range of foods, animal feeds, and production-environment samples.
The assay has previously been validated according to the AOAC Research Institute Performance Tested Methods(SM) program using Thermo Scientific PikoReal™ PCR cycler and Thermo Scientific SureTect Software Performance Tested Method 051303).
This report details the method-modification study performed to validate an updated assay format, utilizing a reduced target probe concentration and an extension of the PCR cycler platform to enable the use of the kit with a Applied Biosystems 7500 Fast PCR cycler and Applied Biosystems RapidFinder™ Express 2.0 software.
During this validation study, a matrix study was conducted on a subset of the method’s claimed matrixes, comparing the performance of the modified SureTect Salmonella species kit (a reduced target probe concentration with a 7500 Fast platform) to the reference method detailed in ISO 6579:2002.
No significant difference by probability of detection statistical analysis was found between SureTect or International Organization for Standardization methods for any of the matrixes analyzed during the study. Inclusivity and exclusivity studies using the modified method demonstrated accurate results for the 117 Salmonella and 36 non-Salmonella strains tested.
Multiple production lots of the newly formatted kit were evaluated and found to be consistent with the current assay.
Robustness studies confirmed that the change to the kit had no impact on the assay’s performance when alterations were made to method parameters having the greatest potential impact on assay performance.
Method modification of the Legipid® Legionella fast detection test kit.
Legipid(®) Legionella Fast Detection is a test based on combined magnetic immunocapture and enzyme-immunoassay (CEIA) for the detection of Legionella in water. The test is based on the use of anti-Legionella antibodies immobilized on magnetic microspheres.
Target microorganism is preconcentrated by filtration. Immunomagnetic analysis is applied on these preconcentrated water samples in a final test portion of 9 mL.
The test kit was certified by the AOAC Research Institute as Performance Tested Method(SM) (PTM) No. 111101 in a PTM validation which certifies the performance claims of the test method in comparison to the ISO reference method 11731-1998 and the revision 11731-2004 “Water Quality: Detection and Enumeration of Legionella pneumophila” in potable water, industrial water, and waste water.
The modification of this test kit has been approved. The modification includes increasing the target analyte from L. pneumophila to Legionella species and adding an optical reader to the test method. In this study, 71 strains of Legionella spp. other than L. pneumophila were tested to determine its reactivity with the kit based on CEIA.
All the strains of Legionella spp. tested by the CEIA test were confirmed positive by reference standard method ISO 11731. This test (PTM 111101) has been modified to include a final optical reading.
A methods comparison study was conducted to demonstrate the equivalence of this modification to the reference culture method.
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Two water matrixes were analyzed. Results show no statistically detectable difference between the test method and the reference culture method for the enumeration of Legionella spp.
The relative level of detection was 93 CFU/volume examined (LOD50). For optical reading, the LOD was 40 CFU/volume examined and the LOQ was 60 CFU/volume examined.
Results showed that the test Legipid Legionella Fast Detection is equivalent to the reference culture method for the enumeration of Legionella spp.