Licensing and Validation

What does fully licensed mean?
Use of real-time PCR technology for food and GMO testing requires licensing from patent holders. BIOTECON Diagnostics’ products are fully licensed. This guarantees the customer absolute legal freedom to use our products. The use of unlicensed products is patent infringement.
Are foodproof® Detection Kits validated?
All foodproof® and microproof® kits are internally validated before being released to the market. This validation includes several criteria that are important for food analysis, such as sensitivity testing, specificity testing, food matrix compatibility, reproducibility and robustness.
Moreover, many foodproof® kits, in particular those for the detection of pathogens, have been independently tested and certified by external institutions such as AOAC-RI, MicroVal, AFNOR or NordVal (link to product quality), which involve a comparison with a reference method (e.g. ISO, USDA, BAM) and interlaboratory comparison of multiple accredited laboratories. These validations are generally carried out as a combination of DNA extraction and real-time PCR detection for all relevant food, feed and environmental samples.
BIOTECON Diagnostics has performed these independent validations of its products for over 10 years, and has many certificates for key parameters, such as Salmonella, Listeria monocytogenes, E. coli O157, Enterobacteriaceae and Cronobacter.

Safety and Benefits

How is cross-contamination in PCR avoided?
BIOTECON Diagnostics has integrated the nucleotide dUTP and the enzyme UNG (uracil-N-glycosylase) into the foodproof real-time PCR kits to prevent cross-contamination from previous reaction runs.
What controls are present in foodproof® Kits?
All ISO mandatory controls are included in all BIOTECON Diagnostics’ kits, which are a positive and negative control as well as an internal amplification control for each sample. Internal amplification controls prevent false-negative results, and are therefore of utmost importance when testing for pathogenic organisms in food, beverages and pharmaceuticals.
Is the highest possible specificity according to ISO 20838 guaranteed?
Yes, BIOTECON Diagnostics uses fluorescently labeled, sequence-specific probes for real-time PCR detection, in contrast to sequence-nonspecific intercalating dyes such as SYBR Green.
How does BIOTECON Diagnostics distinguish between live and dead bacteria when performing PCR analysis?
BIOTECON Diagnostics is the first company to commercially offer a reagent to distinguish the DNA of living and dead cells. Validated according to ISO 16140 (MicroVal), Reagent D was specifically developed to exclude DNA of dead cells and only detect the DNA of living cells within a PCR reaction.
Do foodproof® kits detect all desired species or genera while avoiding false-positives from a related species or genus?
All kits have been tested with a large number of different strains. During inclusivity testing, representative bacteria of the entire desired taxonomic group of organisms are tested and assured to be positively detected. During exclusivity testing, a representative selection of closely related organisms or organisms of the same habitat is examined for negative detection. In both cases, the results must be 100% positive (inclusivity study) or negative (exclusivity study). For the validation of the foodproof® Salmonella kit, more than 700 strains were tested, and even invA-negative strains, such as S. senftenberg, could be detected. This ensures that our foodproof® kits provide the highest level of safety and specificity for the consumer.


At what temperature should I store kits, and how long are they valid?
Liquid foodproof®, foodproof® SL oder microproof® PCR kits are shipped on dry ice and must be stored in the freezer at -15 to -25° C. Lyophilized foodproof® LyoKits are shipped at room temperature and should be stored in a sealed dry bag at 4-8° C. foodproof® DNA isolation kits (ShortPrep, StarPrep, Sample Preparation and Magnetic Preparation) are shipped at room temperature and should be stored at 15 to 25° C. Kit expiration can be found on the packaging of each individual kit. Following production, kits have an expiration date of at least 12 months. Several buffers and reagents must be stored differently after opening. Please note the information in the individual package inserts.
Must the PCR mix (PCR setup) be performed on ice?
No, there is no need to prepare PCR reagents on ice. This can be done at room temperature, as the reagents have been validated and are robust enough. After completing the PCR mix, the remaining reagents should be quickly frozen, and the PCR run started as soon as possible.
Should there be an unexpected interruption, it is recommended that you keep the PCR mix in the refrigerator until able to resume working (do not freeze).

Basics of real-time PCR

Do I need a molecular biologist to perform BIOTECON Diagnostics kits?
There is no need to have a specially trained technician or molecular biologist to be able to use our kits. Our kits are very user friendly and easy to learn and perform. All customers are provided a comprehensive theoretical and practical training. Additionally, regular molecular biology workshops and seminars offered at BIOTECON Diagnostics which allow users to further educate themselves in both techniques and analysis.
What is a melting curve, and what benefits does it provide?
Breaking the base-base hydrogen bonding between two DNA strands requires energy, which depends on the length and the GC content of the particular DNA sequence. At low temperature, the probes bind to their complementary and specific target DNA sequence. While bound, the probes emit a fluorescent signal. If the temperature is then slowly raised, the hydrogen bonds between bases will also break. As enough hydrogen bonds break, the probes detach from the target DNA, and they no longer fluoresce. This loss of fluorescence results in a unique temperature profile, or melting curve. Thus, detachment of a probe at a particular temperature from the target DNA will vary among taxonomic groups. For example, this difference in melting behavior can be used to distinguish the DNA of different species. Melting curves are automatically measured by real-time PCR instruments, allowing clear and easy identification of species.

Real-time PCR Cycler Compatibility

Can I use the foodproof® kits on all real-time PCR cyclers?
BIOTECON Diagnostics offers foodproof® detection, screening and quantification kits for both the Roche LightCycler® probe technology (hybridization probes) as well as TaqMan® probe technology (5’Nuclease / hydrolysis probes). For LightCycler® probe technology, BIOTECON Diagnostics has optimized PCR kits for the carousel based LightCycler® 1.x and 2.0 as well as for the LightCycler® 480. The 5’Nuclease real-time PCR kits run on a variety of real-time PCR cyclers, including the LightCycler® 480 and LightCycler® 96 (Roche Diagnostics), MX 3005P® QPCR System (Agilent) , ABI 7500 Fast and StepOnePlus (Life Technologies), CFX96 Real-Time PCR System (Bio-Rad) and PikoReal24 unit (Thermo Fisher). For more information on which real-time PCR cyclers have been tested and validated with kits from BIOTECON Diagnostics, please refer to the individual package inserts.

Practice Questions

Do I need to repeat my positive / negative results for confirmation of real-time PCR results from BIOTECON Diagnostics kits?
In accordance with the ISO standards ISO 22174 and ISO 20838, confirmation of real-time PCR results obtained using BIOTECON Diagnostics’ kits is not necessary because the integrated primers and probes in the foodproof kits already duplicate and thus verify the results. However, the decision is up to the user if they would like to even further confirm results or not.
How do I dispose of reagents and supplies used for real-time PCR?
An MSDS (Material Safety Data Sheet), which provides information on use and disposal, is available for all BIOTECON Diagnostics’ reagents and consumables. The latest version of each MSDS can be downloaded from BIOTECON Diagnostics’ website on each particular products webpage (under Products).