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. 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.
Development of an in-house fast real-time PCR method for detection of fish allergen in foods and comparison with a commercial kit.
Food allergy is recognised as an important human health problem. Fish represent one of the most important causes of food hypersensitivity reaction. Small amounts of the allergen can cause severe reactions in sensitive individuals, so correct labelling is essential to ensure the protection of consumers. The objective of the present work was to develop a reliable, sensitive and specific real-time PCR method for the detection of fish and traces of fish in all kind of products included those that have undergone aggressive treatments such as high temperature or pressure.
This methodology was validated simulating products likely to contain this allergen and spiking them with fish cooking water. In addition, a comparison between the performance of in-house methodology and a commercial kit, both of them based on real-time PCR, was carried out. This work is relevant because it is the first, rapid real-time PCR method developed to date for the detection of fish in processed food products. The results obtained confirm the present assay is a useful tool in detecting fish and, therefore, minimising exposure and reducing incidences of allergic reaction to fish in contaminated products.
Sample-to-SNP kit: a reliable, easy and fast tool for the detection of HFE p.H63D and p.C282Y variations associated to hereditary hemochromatosis.
Classical hereditary hemochromatosis involves the HFE-gene and diagnostic analysis of the DNA variants HFE p.C282Y (c.845G>A; rs1800562) and HFE p.H63D (c.187C>G; rs1799945). The affected protein alters the iron homeostasis resulting in iron overload in various tissues. The aim of this study was to validate the TaqMan-based Sample-to-SNP protocol for the analysis of the HFE-p.C282Y and p.H63D variants with regard to accuracy, usefulness and reproducibility compared to an existing SNP protocol. The Sample-to-SNP protocol uses an approach where the DNA template is made accessible from a cell lysate followed by TaqMan analysis. Besides the HFE-SNPs other eight SNPs were used as well. These SNPs were: Coagulation factor II-gene F2 c.20210G>A, Coagulation factor V-gene F5 p.R506Q (c.1517G>A; rs121917732), Mitochondria SNP: mt7028 G>A, Mitochondria SNP: mt12308 A>G, Proprotein convertase subtilisin/kexin type 9-gene PCSK9 p.R46L (c.137G>T), Plutathione S-transferase pi 1-gene GSTP1 p.I105V (c313A>G; rs1695), LXR g.-171 A>G, ZNF202 g.-118 G>T. In conclusion the Sample-to-SNP kit proved to be an accurate, reliable, robust, easy to use and rapid TaqMan-based SNP detection protocol, which could be quickly implemented in a routine diagnostic or research facility.
Performance Analysis of Novel Nucleic Acid Detection Kit for Mycoplasma pneumoniae
Respiratory tract infections caused by Mycoplasma pneumoniae is a serious risk for child health. It has been difficult to prevent and control for a variety of reasons; therefore, timely diagnosis is particularly important for treatment of patients. At present, the rapid M pneumoniae test kits based on nucleic acid amplification have been commercialized and used as primary diagnostic tools for M pneumoniae infection, but current kits are time-consuming, which is difficult to meet the requirement for accurate and rapid diagnosis of M pneumoniae during epidemics.
EpiQuik Total Histone H3 Acetylation Detection Fast Kit (Colorimetric) | |||
| P-4030 | EpiGentek |
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EpiQuik Total Histone H3 Acetylation Detection Fast Kit (Fluorometric) | |||
| P-4031 | EpiGentek |
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EpiQuik Total Histone H4 Acetylation Detection Fast Kit (Colorimetric) | |||
| P-4032 | EpiGentek |
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EpiQuik Total Histone H4 Acetylation Detection Fast Kit (Fluorometric) | |||
| P-4033 | EpiGentek |
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Rat Acetylation Histone H3 ELISA kit | |||
| E01A10254 | BlueGene | 96T | 700 EUR |
Rat Acetylation Histone H3 ELISA kit | |||
| E02A2149-192T | BlueGene | 192 tests | 1524 EUR |
Rat Acetylation Histone H3 ELISA kit | |||
| E02A2149-48 | BlueGene | 1 plate of 48 wells | 624 EUR |
Rat Acetylation Histone H3 ELISA kit | |||
| E02A2149-96 | BlueGene | 1 plate of 96 wells | 822 EUR |
Pig Acetylation Histone H3 ELISA kit | |||
| E07A2149-192T | BlueGene | 192 tests | 1524 EUR |
Pig Acetylation Histone H3 ELISA kit | |||
| E07A2149-48 | BlueGene | 1 plate of 48 wells | 624 EUR |
Pig Acetylation Histone H3 ELISA kit | |||
| E07A2149-96 | BlueGene | 1 plate of 96 wells | 822 EUR |
Dog Acetylation Histone H3 ELISA kit | |||
| E08A2149-192T | BlueGene | 192 tests | 1524 EUR |
Dog Acetylation Histone H3 ELISA kit | |||
| E08A2149-48 | BlueGene | 1 plate of 48 wells | 624 EUR |
Dog Acetylation Histone H3 ELISA kit | |||
| E08A2149-96 | BlueGene | 1 plate of 96 wells | 822 EUR |
Goat Acetylation Histone H3 ELISA kit | |||
| E01A45173 | BlueGene | 96T | 700 EUR |
Goat Acetylation Histone H3 ELISA kit | |||
| E06A2149-192T | BlueGene | 192 tests | 1524 EUR |
Rapid and accurate test kits are urgently required to diagnose M pneumoniae infection. In this article, we evaluated the performance of a novel nucleic acid detection kit (A) for M pneumoniae from feasibility and sensitivity, and compared it with kit B. Results showed this kit has the advantage of being rapid, sensitive, and specific, which meets the demands for the diagnosis of M pneumoniae infection in clinical settings.
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