Cronobacter Support
05-12-2009, 07:07 PM
1st International Conference on Cronobacter Poster Abstract 24
Rapid detection of Enterobacteriaceae including identification of E. sakazakii (Cronobacter)
BIOTECON Diagnostics has developed a real-time PCR system for the detection of all Enterobacteriaceae with a simultaneous identification of E. sakazakii by parallel detection with differently labeled probes. The test is designed to run on all relevant real-time PCR instruments and comprises all necessary reagents. The method is thoroughly validated for the use in infant formulae with and without probiotic bacteria as well as raw materials and environmental samples. During the validation of the method it was recognized, that most infant formula products contain a background of inactive or non-cultivatable Enterobacteriaceae which leads to positive results in the PCR but cannot be confirmed by cultural methods. To overcome this discrepancy, Reagent D is applied in advance to the DNA extraction, a reagent which inactivates DNA from dead cells for PCR. The kit also includes all necessary controls such as positive and negative and an internal positive control to eliminate false negative results due to inhibition or failures. UNG is used to prevent false positive results by carry-over contamination with "old" amplificates. The presentation will give an overview about the method, its application and results from validation and routine use.
Cordt Grönewald, Matthias Kiehne, Kornelia Berghof-Jäger
BIOTECON Diagnostics GmbH, Hermannswerder Haus 17, 14473 Potsdam, Germany.
Rapid detection of Enterobacteriaceae including identification of E. sakazakii (Cronobacter)
BIOTECON Diagnostics has developed a real-time PCR system for the detection of all Enterobacteriaceae with a simultaneous identification of E. sakazakii by parallel detection with differently labeled probes. The test is designed to run on all relevant real-time PCR instruments and comprises all necessary reagents. The method is thoroughly validated for the use in infant formulae with and without probiotic bacteria as well as raw materials and environmental samples. During the validation of the method it was recognized, that most infant formula products contain a background of inactive or non-cultivatable Enterobacteriaceae which leads to positive results in the PCR but cannot be confirmed by cultural methods. To overcome this discrepancy, Reagent D is applied in advance to the DNA extraction, a reagent which inactivates DNA from dead cells for PCR. The kit also includes all necessary controls such as positive and negative and an internal positive control to eliminate false negative results due to inhibition or failures. UNG is used to prevent false positive results by carry-over contamination with "old" amplificates. The presentation will give an overview about the method, its application and results from validation and routine use.
Cordt Grönewald, Matthias Kiehne, Kornelia Berghof-Jäger
BIOTECON Diagnostics GmbH, Hermannswerder Haus 17, 14473 Potsdam, Germany.