hygiena KIT230134 Kit For The Qualitative Detection Of Salmonella Instructions

: June 28, 2024
: Hygiena

Table of Contents

hygiene KIT230134 Kit For The Qualitative Detection Of Salmonella
What This Product Does
How to Use This Product
Troubleshooting
Additional Information on this Product
Supplementary Information
References
Read User Manual Online (PDF format)
Download This Manual (PDF format)

hygiene KIT230134 Kit For The Qualitative Detection Of Salmonella

foodproof® Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit

Revision A, May 2024
PCR kit for the qualitative detection of Salmonella spp., Salmonella Enteritidis, and Salmonella Typhimurium using real-time PCR instruments.

Product No.
- KIT230134 (LP)
- KIT230135 (RP)
- KIT230136 (DP)

Kit for 96 reactions (lyophilized) for a maximum of 94 samples Store the kits at 2 to 8 °C

What This Product Does

Number of Tests
These kits are designed for 96 reactions with a final reaction volume of 25 µL each. Up to 94 (single sample preparation) plus positive and negative control reactions can be analyzed per run.

Storage and Stability

Store the kit at 2 to 8 °C through the expiration date printed on the label.
Once the kit is opened, store the kit components as described in the following Kit Contents table:

Kit Contents

Component	Label	Contents / Function / Storage

foolproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit

KIT230134-35-36:

Microplate prefilled with 96 reactions (lyophilized)

| Aluminum bags containing an 8-tube strip mat

• KIT230134 with white low-profile* tubes

• KIT230135 with clear regular profile* tubes

• KIT230136 with clear low-profile* tubes

| • 96 prefilled reactions (lyophilized).

• Ready-to-use PCR mix containing primer and hydrolysis probes specific for Salmonella spp., S. Enteritidis (SE) and

S. Typhimurium (STM) DNA and the Internal Control (IC) as well as Taq DNA Polymerase and Uracil-DNA Glycosylase (UNG, heat-labile) for prevention of carry-over contamination.

• For amplification and detection of Salmonella spp. , Salmonella Enteritidis, and Salmonella Typhimurium specific sequences.

• Store at 2 to 8 °C in the aluminum bag (sealed).

• Protect from light and moisture!

Control Template| Vial 1 (purple cap)| • 1 x 300 µL

• Contains a stabilized solution of DNA.

• For use as a PCR run positive control.

• Store at 2 to 8 °C.

H2O PCR-grade| Vial 2 (colorless cap)| • 2 x 1 mL

• Nuclease-free, PCR-grade H2O.

• For use as a PCR run negative control.

Cap strips| Plastic bag containing 8- cap strips| • 12 x 8-cap strip

• For use in real-time PCR after the addition of samples.

Tube profile and instrument compatibility chart are available online: www.hygiena.com/documents

Additional Equipment and Reagents Required

Real-time PCR cycler suitable for detection of FAM-, HEX-, ROX- and Cy5-labeled probes as well as for using low or regular profile strip tubes. If the strip tubes don´t fit the instrument being used, the samples should be transferred to appropriate PCR vessels after resuspension of the lyophilized PCR mix.
DNA Extraction Kit
foolproof StarPrep Three Kit (Product No. KIT230187) or
foolproof StarPrep Three 8-Strip Kit (Product No. KIT230188)

Alternatives:

foolproof Magnetic Preparation Kit I (Product No. KIT230180)
microproof® Suspension Buffer (Product No. KIT230178) is only required for colony screening

Additional requirements:

Internal Amplification Control (Product No. KIT230015) – only required for colony screening
Nuclease-free, aerosol-resistant pipette tips
Pipettes
Vortex-centrifuge Multispin MSC-6000 for PCR strips with SR-32, Rotor for MSC-3000/6000 or Vortex-centrifuge CVP-2 for PCR plates

Applicability Statement
The foolproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit is intended for rapid detection of Salmonella spp. and identification of Salmonella Enteritidis (SE) and Salmonella Typhimurium (STM) DNA from enrichment cultures of all relevant kinds of foods, feeds, environmental samples and primary production stage (PPS) samples that are potentially contaminated with Salmonella. The foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit is intended for the food and feed industry and food testing laboratories. The kit must not be used in diagnostic procedures. The kit described in this Product Instructions document has been developed for real-time PCR instruments with FAM, HEX, ROX, and Cy5 detection channels. The performance of the kit was tested with the following real-time PCR instruments: LightCycler® 480 II, LightCycler 96 (Roche Diagnostics), Mx3005P®, AriaMx (Agilent Technologies), ABI® 7500 fast (Thermo Scientific) PikoRealTM 24, QuantStudioTM 5 (Thermo Scientific) and the CFX96TM Real-Time PCR Cycler (Bio-Rad). The performance of the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit in combination with the foodproof StarPrep Three Kit (Procedure A) and the foodproof StarPrep Three 8-Strip Kit (Procedure A) has been approved in a NordVal International method validation (Certificate No. 055). For this validation study, the following categories were tested: raw and ready-to-cook meat products, raw and ready-to-cook poultry products, and environmental samples. For further information about the enrichment protocols, please refer to ANNEX 1 at the end of the package insert.

How to Use This Product

Before You Begin

Precautions
Detection of Salmonella, Enteritidis, and Typhimurium DNA using the foolproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit requires DNA amplification by PCR. The kit provides all the reagents required for the PCR reaction. However, to achieve reliable results, the entire assay procedure must be performed under nuclease-free conditions. Follow the instructions below to avoid nuclease, carry-over o,r cross-contamination:

Keep the kit components separate from other reagents in the laboratory.
Use nuclease-free labware (e.g., pipettes, pipette tips, reaction vials).
Wear gloves when performing the assay.
To avoid cross-contamination of samples and reagents, use fresh aerosol-preventive pipette tips.
To avoid carry-over contamination, transfer the required solutions for one experiment into a fresh tube, rather than directly pipetting from stock solutions.
Physically separate the workplaces for DNA preparation, PCR setup, and PCR to minimize the risk of carry-over contamination. Use a PCR hood for all pipetting steps.

Keep the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit lyophilized PCR Mix away from light and moisture.

Sample Material
Use any sample material suitable for PCR in terms of purity, concentration, and absence of inhibitors. For the preparation of genomic DNA from various sample enrichments, refer to the corresponding product instructions for a suitable sample preparation kit (see “Additional Equipment and Reagents Required”).

DNA Extraction
Hygiena Diagnostics provides sample preparation kits suitable for all kinds of food, feed, and PPS samples (see “Additional Equipment and Reagents Required”).
For more product information, please refer to www.hygiena.com.

Positive Control
Always run a positive control with the samples. To prepare a positive control, replace the template DNA with the provided control DNA [foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit Control Template (vial 1, purple cap)] or use a positive sample preparation for SE and STM as the positive control.

Negative Control
Always run a negative control with the samples. To prepare a negative control, replace the template DNA with PCR-grade H2O (vial 2, colorless cap). Include a negative control during sample preparation to monitor reaction purity and cross-contamination. This extraction control can be used as an additional negative control reaction.

Confirmation
If required, positive results must be confirmed by appropriate methods (e.g., reference method ISO 6579-1).

Procedure for Qualitative Detection of Salmonella from Enrichment Cultures

Program Setup
The following procedure is optimized for a real-time PCR instrument with a FAM (SE detection), HEX (STM detection), ROX (Salmonella spp. detection) and Cy5 (Internal Control detection) channel. Program the PCR instrument before preparing the samples. Use the following real-time PCR protocol for the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit. For details on how to program the experimental protocol, see the Instrument Operator’s Manual of your real-time PCR cycler:

Pre-incubation 1 cycle

Step 1 : 37 °C for 4 minutes
Step 2 : 95 °C for 5 minutes

Amplification 50 cycles

Step 1: 95 °C for 5 seconds
*Step 2:** 60 °C for 60 seconds

Fluorescence detection in step 2

Note:

For some real-time PCR instruments, the type of probe quencher as well as the usage of a passive reference dye has to be specified. The foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit contains probes with a non-fluorescent (“dark”) quencher and no passive reference dye.
For LightCycler 480 Systems I and II, color compensation is required and will be supplied by Hygiena Diagnostics (Color Compensation Set 3; Product No. KIT230005). Please contact Hygiena Diagnostics for further information.
For users of the Agilent Mx3005P instrument: Click ‘Instrument → Filter Set Gain Settings’ to open the Filter Set Gain Settings dialog box in which the gain settings may be viewed and modified. The Filter Set Gain Setting has to be modified to FAM ‘x2’, HEX to ‘x2’, ROX to ‘x1’, and Cy5 to ‘x4’.
For CFX 96 Analysis Software: ‘Apply Fluorescence Drift Correction’ in ‘Settings’ → ‘Baseline Settings’ has to be activated.

Preparation of the PCR Mix
Proceed as described below to prepare a 25 µL standard reaction. Always wear gloves when handling strips or caps. Use any sample material suitable for PCR in terms of purity, concentration ,and absence of inhibitors.

Note: The PCR strips must be stored in the provided aluminum bag with tilica gel pads to avoid liquid absorption.

Take the needed number of PCR tube strips out of the aluminum bag. Use scissors or a scalpel to cut the strips apart. Tightly seal the bag afterward and store it under the recommended conditions.
Place the PCR tube strips containing the lyophilized reagents in a suitable PCR tube rack. Check that the reagent pellets are at the bottom of the tubes. If not, briefly centrifuge or flick the pellets to the bottom before proceeding.
Carefully uncap the tube strips and discard the cap strips.
Note: Do not leave strips open for extended periods. To avoid unwanted liquid absorption, only open strips shortly before filling.
Pipet 25 µL sample into each PCR vessel:
- For the samples of interest, add 25 µL sample DNA. (If using less volume, add PCR-grade H2O to achieve 25 µL).
- For the negative control, add 25 µL PCR-grade H2O (vial 3, colorless cap).
- For the positive control, add 25 µL foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit Control Template (vial 2, purple cap).
  Note: To reduce the risk of cross-contamination, it is recommended to prepare only one PCR tube strip at a time.
Seal the vessels accurately and tightly with the colorless cap strips.
Mix thoroughly using a vortex centrifuge.
Note: Hygiena Diagnostics recommends vortex centrifuges Multispin MSC-3000 for PCR strips or vortex centrifuge CVP-2 for PCR plates. Dedicated protocols are available for this centrifuge.
Note: Alternatively, resuspend the pellet by manual mixing. This may be achieved by carefully pipetting the sample up and down multiple times during step 4 or flipping the tube strips after sealing while pressing down on the cap strip.
Spin the PCR tube strips for 30 seconds at 150 – 200 x g in a suitable centrifuge.
Note: If your centrifuge exceeds 200 x g, do not centrifuge for more than 5 seconds. Avoid centrifugation at forces exceeding 1,000 x g!
Place the samples in your PCR cycler and run the program as described above.
Note: When using any LightCycler 480 instrument, a special adapter is necessary. For some PCR instruments, the PCR strips should be placed in a balanced order into the cycler block. For example, two strips can be placed in columns 1 and 12.

Procedure for colony identification of Salmonella

Program Setup
The following procedure is optimized for rapid confirmation and identification of presumptive Salmonella colonies obtained in microbiological culture methods (e.g., ISO 6579) for a real-time PCR instrument with a FAM (SE detection), HEX (STM detection), ROX (Salmonella spp. detection) and Cy5 (Internal Amplification Control detection) channel. Program the PCR instrument before preparing the samples. Use the following real-time PCR protocol for the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit. For details on how to program the experimental protocol, see the Instrument Operator’s Manual of your real-time PCR cycler:

Pre-incubation 1 cycle

Step 1: 37 °C for 4 minutes
Step 2: 95 °C for 5 minutes

Amplification 30 cycles

Step 1 : 95 °C for 5 seconds
Step 2 *: 60 °C for 60 seconds

Fluorescence detection in step 2

Note

For some real-time PCR instruments, the type of the probe quencher as well as the usage of a passive reference dye must be specified. The foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit contains probes with a non-fluorescent dark quencher and no passive reference dye.
For LightCycler 480 Systems I and II, color compensation is required and will be supplied by Hygiena Diagnostics (Color Compensation Set 3; Product No. KIT230005). Please contact Hygiena Diagnostics for further information.
For users of the Agilent Mx3005P instrument: Click ‘Instrument → Filter Set Gain Settings’ to open the Filter Set Gain Settings dialog box in which the gain settings may be viewed and modified. For FAM the Filter Set Gain Setting has to be modified to FAM ‘x2’, HEX to ‘x2’, ROX to ‘x1’, Cy5 to ‘x4’

Preparation of the PCR Mix

Proceed as described below to prepare a 25 μL standard reaction. Always wear gloves when handling strips or caps. Use material from single colonies prepared with the microproof® Suspension Buffer (KIT230178).

Note: The PCR strips must be stored in the provided aluminum bag with silica gel pads to avoid liquid absorption.

Take the needed number of PCR tube strips out of the aluminum bag. Use scissors or a scalpel to cut the strips apart. Tightly seal the bag afterward and store it under the recommended conditions.
Place the PCR tube strips containing the lyophilized reagents in a suitable PCR tube rack. Check that the reagent pellets are at the bottom of the tubes. If not, briefly centrifuge or flick the pellets to the bottom before proceeding.
Carefully uncap the tube strips and discard the cap strips.
Note: Do not leave strips open for extended periods. To avoid unwanted liquid absorption, only open strips immediately before filling.
- For CFX 96 Analysis Software: ‘Apply Fluorescence Drift Correction’ in ‘Settings’ → ‘Baseline Settings’ has to be activated.
Pipetting of samples:
- Pipet 20 μL Internal Amplification Control (KIT230015) into each PCR vessel.
- Add 5 μL of suspended colony material to each well.
  Note: Cells in a microproof Suspension Buffer are not inactivated before the pre-incubation step of PCR and are still considered hazardous.
  Note: To reduce the risk of cross-contamination, it is recommended to prepare only one PCR tube strip at a time.
Pipetting of controls:
- For the negative control, pipet 20 μL Internal Amplification Control (KIT230015) into a PCR vessel and add 5 μL PCR-grade H2O (vial 3, colorless cap).
- For the positive control, add 25 μL foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit Control Template (vial 2, purple cap). 6 Seal the vessels accurately and tightly with the colorless cap strips.
Mix thoroughly using a vortex centrifuge.
Note: Hygiena Diagnostics recommends vortex centrifuge Multispin MSC-6000 for PCR strips or vortex centrifuge CVP-2 for PCR plates. Dedicated protocols are available for these centrifuges.
Note: Alternatively, resuspend the pellet by manual mixing. Resuspension may be achieved by cautiously pipetting the sample up and down multiple times during step 4 or flipping the tube strips after sealing while holding down the cap strip.
Spin the PCR tube strips for 30 seconds at 150 – 200 x g in a suitable centrifuge.
Note : If your centrifuge exceeds 200 x g, do not centrifuge for more than 5 seconds. Avoid centrifugation forces exceeding 1,000 x g!
Place the samples in your PCR cycler and run the program as described above.
- Note: When using any LightCycler 480 instrument, a special adapter is necessary. For some PCR instruments, the PCR strips should be placed in a balanced order into the cycler block. For example, two strips can be placed in columns 1 and 12.

Data Interpretation
The amplification of the Salmonella-specific DNA region is analyzed in the fluorescence channel suitable for the ROX-labeled probe detection. The specific DNA region for Salmonella serotype Enteritidis is analyzed in the fluorescence channel suitable for FAM-labeled probe detection and Salmonella Typhimurium in the fluorescence channel suitable for HEX-labeled probe detection. To confirm that negative PCR results are compliant with expected results, an Internal Amplification Control (IC) is included in the assay. To exclude matrix-based PCR inhibition if FAM, HEX, and ROX channels are negative, check the IC in the Cy5 channel. A negative result is valid if the IC has a positive signal in Cy5. Should all channels show a negative result, please refer to Section 3 for troubleshooting. Compare the results from channel FAM (Enteritidis), HEX (Typhimurium), ROX (Salmonella spp.), and channel Cy5 (Internal Control) for each sample, and interpret the results as described in the table below.

Channel FAM| Channel HEX| Channel ROX| Channel Cy5|

Result Interpretation

---|---|---|---|---

Positive

|

Negative

|

Positive

| Positive or Negative| Positive for S. Enteritidis Negative for S. Typhimurium

Negative or Positive for other Salmonella spp.

Negative

|

Positive

|

Positive

| Positive or Negative| Positive for S. Typhimurium Negative for S. Enteritidis

Negative or Positive for other Salmonella spp.

Negative

|

Negative

|

Positive

| Positive or Negative|

Positive for Salmonella spp.

Negative for S. Enteritidis and S. Typhimurium1

Positive

|

Positive

|

Positive

| Positive or Negative|

Positive for S. Enteritidis and S. Typhimurium Negative or Positive for other Salmonella spp.

If the amplification in the ROX channel is very weak (high Cq values), the result in the FAM and/or HEX channels could be negative due to slight differences in the assays’ limit of detection in this multiplex PCR system. In this case, extending the enrichment, performing a second enrichment, or using a larger amount of sample DNA could be used to increase the sensitivity. Due to these slight differences, a negative result in the ROX channel might also occur in the event of very weak amplification for Enteritidis and/or Typhimurium in the FAM/HEX channels.

Note: A prerequisite for the unambiguous discrimination of Salmonella spp., S. Enteritidis, S. Typhimurium, and the Internal Control DNA in this multi-color experiment is a suitable calibration of the PCR instrument for channels FAM, HEX, ROX, and Cy5. Please refer to the operation manual of your real-time PCR cycler for further information.

Troubleshooting

Observation	Possible Reason	Recommendation

No signal increase, even with positive controls.

| An incorrect detection channel has been chosen.| · Set channel settings of FAM, VIC/HEX, ROX, and Cy5.

Pipetting errors.

| · Check for correct reaction setup. Repeat the PCR run.

· Always run a positive control along with your samples.

No data acquisition programmed.| · Check the cycle program.
No signal increase in channel Cy5.| Inhibitory effects of the sample material (e.g., caused by insufficient purification).| · Dilute samples or pipet a lower amount of sample DNA (e.g., 20 µL PCR-grade H2O and 5 µL sample DNA instead of 25 µL sample DNA).
Large amounts of Salmonella spp., Enteritidis or Typhimurium| · If a positive signal in FAM, HEX, or ROX is detected, the PCR run is valid.

Fluorescence intensity is too low.

| ****

Inappropriate storage of kit components.

| · Store the lyophilized PCR mix for the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit at 2 to 8 °C, protected from light and moisture.

The low initial amount of target DNA.

| · Increase the amount of sample DNA. Depending on

the chosen DNA isolation method, inhibitory effects may occur.

Strong decrease of fluorescence baseline.| ****

Resuspension of lyophilized PCR mix not complete.

| · Always resuspend lyophilized PCR mix thoroughly.

· Use a recommended vortex centrifuge with the correct settings.

Negative control samples are positive.

| ****

Carry-over contamination.

| · Exchange all critical solutions.

· Repeat the complete experiment with fresh aliquots of all reagents.

· Always handle samples, kit components, and consumables by commonly accepted practices to prevent carry-over contamination.

· Add positive control after all sample and negative control reaction tubes have been sealed.

Fluorescence intensity varies.

| Insufficient centrifugation of the PCR strips. Resuspension of the PCR mix only in the upper part of the reaction tube.| ****

· Always centrifuge PCR strips.

The outer surface of the vessel or the seal is dirty (e.g., by direct skin contact).| · Always wear gloves when handling the vessels and seal.

Pellets are hard to dissolve.

| ****

The lyophilized PCR mix started to rehydrate.

| · Store the lyophilized PCR mix tightly sealed in the aluminum bag with a silica gel pad.

· Open strips just before filling.

Additional Information on this Product

How this Product Works
The foolproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit contains all necessary reagents (except for template DNA) needed for the detection of Salmonella spp., S. Enteritidis and S. Typhimurium DNA as well as a control template for reliable interpretations of the results. To ensure maximum reliability of the kit and to prevent misinterpretation of negative results due to inhibition of the amplification, an Internal Control (IC) is included. A hydrolysis probe was designed to bind specifically the IC, allowing detection in the Cy5 channel, whereas the specific S. Enteritidis DNA is detected in the FAM channel, the specific S. Typhimurium DNA is detected in the HEX channel, and Salmonella spp. DNA is detected in the ROX channel. In case of a negative result due to inhibition of the amplification by the sample DNA of interest, the amplification of the IC is suppressed as well, whereas a negative result for the sample DNA of interest and amplification of the IC indicates the absence of Salmonella in the sample. The foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit minimizes contamination risk since it includes a heat-labile Uracil DNA N-Glycosylase. Primers and probes provide specific detection of Salmonella spp., S. Enteritidis, and S. Typhimurium DNA in food, feed, and environmental samples, including samples of primary production stages (PPS). The described performance of the kit is guaranteed for use only on the real-time PCR instruments listed above.

Test Principle

Using the kit’s sequence-specific primers in a polymerase chain reaction (PCR), the PCR instrument, in combination with the supplied reagents, amplifies fragments of specific sequences for Salmonella spp., Salmonella Enteritidis, and Salmonella Typhimurium.
The PCR instrument detects these amplified fragments in rehearsal-time rough fluorescence generated by cleavage of the hybridized probe due to the 5´-nuclease activity of the Taq DNA polymerase. The probe is labeled at the 5´-end with a reporter fluorophore and at the 3´-end with a quencher.
During each PCR cycle’s annealing/elongation phase, the probe hybridizes to an internal sequence of the amplicon and is cleaved by the 5’-nuclease activity of Taq DNA polymerase. This cleavage of the probe separates the reporter dye from the quencher dye, increasing the reporter dye signal.
The PCR instrument measures the emitted fluorescence of the reporter dye.

Prevention of Carry-Over Contamination
The heat-labile Uracil DNA N-Glycosylase (UNG) is suitable for preventing carry-over contamination between PCR’s. This technique relies on incorporating deoxyuridine triphosphate (dUTP) during all amplification reactions and the pretreatment of all successive PCR mixtures with the heat-labile UNG. The UNG cleaves DNA at any site where a deoxyuridine residue has been incorporated. The resulting abasic sites are hydrolyzed due to the high temperatures during the initial denaturation step and can no longer serve as PCR templates. The heat-labile UNG is inactivated during the initial denaturation step. Native DNA (e.g., the isolated Salmonella genomic DNA) does not contain uracil and is therefore not degraded by this procedure. Since dTTP is replaced with dUTP and UNG is included in the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit, decontamination can be achieved with the provided reagents.

Background Information
The genus Salmonella, a member of the Enterobacteriaceae family, comprises the two species S. enterica and S. bongori. S. enterica is further divided into six subspecies, of which S. enterica subspecies I enterica is the most clinically significant, causing 99% of Salmonella infections. The subspecies are further subdivided into more than 2,000 serovars defined by somatic and flagellar antigens. S. Enteritidis and S. Typhimurium, both belonging to the subspecies I, are the most frequently reported serovars associated with human cases of Salmonella infection in the EU in 2018 from foodborne outbreaks. S. Enteritidis cases are still increasing and are most commonly associated with the consumption of contaminated eggs and egg products. Due to their importance as human pathogens, the presence of the serovars S. Typhimurium (mono- or diphasic variants) and S. Enteritidis has been tightly regulated in many countries. In the EU, the regulations 2160/2003 and 200/2010 dictate how to deal with Salmonella. Food, feed, and primary production sites must be tested regularly for relevant Salmonellae to prevent human Salmonella infections from being transmitted through contaminated food. The current accepted microbiological method for isolation and detection of Salmonella, according to ISO 6579 in combination with the Kauffman-White serotyping system, takes up to 5 days to complete. Because this method is very time-consuming, PCR has been introduced to the food, feed, and primary production industry as a highly sensitive, specific, and fast alternative detection method [1, 2, 3, 4].

References

European Food Safety Authority. The European Union summary report on trends and sources of zoonoses, zoonotic agents resistance, and foodborne outbreaks in 2017. 10.2903/j. EFSA J. 2018.5500
Mehnert WH et al. (2000) Deutscher Zoonosen-Trendbericht 2000. BgVV.
Scheu PM et al. (1998) Detection of pathogenic and spoilage micro-organisms in food with the polymerase chain reaction. Food Microbiology 15, 13-31.
Scheu P et al. (1998) Evaluation of a PCR ELISA for food testing: Detection of selected Salmonella serovars in confectionery products. Food Biotechnology 12, 1-12.

Quality Control
The foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit is function tested using the LightCycler 480 II System (KIT230134), the Mx3005P (KIT230135) and the Dualo 32 R2 (KIT230136).

Supplementary Information

Ordering Information
Hygiena Diagnostics offers a broad range of reagents and services. For a complete overview and more information, please visit our website at www.hygiena.com.

License Notice
The purchase price of this product includes limited, nontransferable rights under U.S. Patent No. 7,687,247 owned by Life Technologies Corporation to use only this amount of the product to practice the claims in said patent solely for activities of the purchaser for bioburden testing, environmental testing, food testing, or testing for genetically modified organisms (GMO) by the instructions for use accompanying this product. No other rights are conveyed, including no right to use this product for in vitro diagnostic, therapeutic, or prophylactic purposes. Further information on purchasing licenses under the above patent may be obtained by contacting the Licensing Department, Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008.

Email: outlicensing@lifetech.com.

Trademarks
foodproof®, microproof®, vetproof®, ShortPrep®, StarPrep®, RoboPrep® and LyoKit® are registered trademarks of Hygiena Diagnostics GmbH. Hygiena® is a registered trademark of Hygiena. Other brand or product names are trademarks of their respective holders.

Contact and Support
If you have questions or experience problems with this or any other product of Hygiena Diagnostics GmbH, please contact our Technical Support staff (www.hygiena.com/support). Our scientists commit themselves to providing rapid and effective help. We also want you to contact us if you have suggestions for enhancing our product performance or using our products in new or specialized ways. Such customer information has repeatedly proven invaluable to us and the worldwide research community.

Reference Number
The reference number and original Hygiena Diagnostics GmbH article number:
R 602 59-1 (KIT230134), R 602 59-2 (KIT230135), R 602 59-3 (KIT230136)

Change Index

Version 1, June 2020
The first version of the package insert.
Version 2, November 2021
Addition of NordVal International logo.
Additional information in the Applicability Statement.
Addition of ANNEX 1: NordVal #055 Validation Table.
Revision A, May 2024
Rebranding and new layout.
R 602 59 20 -> INS-KIT-230134-35-36-RevA

ANNEX 1 : NordVal #055 Validation Table for the foolproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit

The following table shows the recommended enrichment time for the tested food categories and environmental samples in Buffered Peptone Water (BPW) in combination with foodproof StarPrep Three Kit (Product No. KIT230187) and foodproof StarPrep Three 8 Strip Kit (Product No. KIT230188) that have been analyzed for the NordVal International method validation of the foodproof Salmonella Genus plus Enteritidis & Typhimurium Detection LyoKit (Certificate No. 055). For further information regarding the DNA extraction procedures below, please refer to the appropriate Hygiena Diagnostics package inserts on www.hygiena.com.

DNA Extraction| Enrichment time in BPW at 37 ± 1 °C| DNA extract for PCR
---|---|---
foolproof StarPrep Three Kit Extraction Procedure A STANDARD|

18 h ± 2 h

|

25 µL

foolproof StarPrep Three 8-Strip Kit Extraction Procedure A STANDARD|

18 h ± 2 h

|

25 µL

Tested food categories: