01. Who has developed BIOFINDER?
BIOFINDER is the result of a joint research project between ITRAM hygiene, UAB (Autonomous University of Barcelona) and the technology centre LEITAT. In the project have been involved experts in chemistry, microbiology and hygiene and food safety.

02. Mechanism of action?
Enzymatic reaction: Biofinder reacts with catalase enzymes from microorganisms.

03. How does the product react?
Biofinder is a biofilm detector that reacts with biofilms and produces foam, which contrasts with the orange color of the product.

04. How does the foam originate?
The foam is caused by an enzymatic reaction that releases oxygen when identifies the presence of catalase enzyme.

05. What is the cause of the foam reaction for 10 min?
Thanks to its surfactant formula and its viscous nature, the production of oxygen bubbles is maintained for a period of time, allowing a visual observation for at least 10 min or longer.

06. What is the minimum amount of microorganisms that should be on a surface to obtain a positive reaction?
For a reaction to be visually positive should be above 4 log units. To lower concentration reacts, but being this a sensorial method, it would give a very weak reaction, being more difficult to visualize.

07. Can there be a false positive reaction?
Yes, and to avoid false positive reactions, we recommend applying the product on surfaces after cleaning and disinfection or alternatively when no visible food waste, i.e. clean.

Some foods, not all, contain catalase and may cause a positive reaction. Anyway, this would indicate low levels of contamination, which is linked to a potential for growth of microorganisms and the consequent formation of biofilms. Finally this would represent a security risk for food. Biofinder in this case would be acting as an indicator of hygiene.

08. Is Biofinder similar to ATP?
Yes, both are enzymatic methods but Biofinder is only qualitative, and detection of ATP by bioluminescence is semi-quantitative. The presence of ATP indirectly assesses the presence of microorganisms or other contaminants that contain the ATP molecule. The enzymatic reaction produces light which is measured on a luminometer. This method uses a scale to interpret the amount of light produced in relative light units (URL). A higher URL readed, the more ATP present in the sample and therefore more contamination.

09. What kind of biofilm does BioFinder detect?
It detects biofilms of microorganisms composed of: bacteria, yeasts and fungi. These last form pseudobiofilms through the growth of their hyphae. Biofilms may be of a single specie (monoespecie) or several species (multispecies). It is worth mentioning that you can also detect some green algae.

10. Which is the list of organisms tested with Biofinder?
- Listeria monocytogenes.
- Escherichia coli.
- Staphylococcus aureus.
- Cronobacter sakazakii
- Salmonella spp.
- Pseudomonas aeruginosa.

11. Does it work with fungi?
Yes, if the microorganisms are aerobics such as Candida albicans, Aspergillus, Penicillium and Saccharomyces cerevisiae, etc. Since this type of micro-organisms are bigger, with the same number of cells (e.g.: 4 log units) would originate a reaction more visible than with bacteria.

12. Is it selective to only one type of bacteria?
Not exactly to only one type of bacteria, but one kind of microorganisms. Our product only reacts against a biofilm composed of one or more micro-organisms that contain the enzyme catalase, i. e. , catalase-positive (bacteria, yeasts and fungi). Let us remember that it detects an enzymatic reaction, a chemical property present in many living microorganisms.

13. Do you have data showing that BioFinder has the ability to detect a wide range of bacteria?
BioFinder has been tested with the most relevant microorganisms in the food industry and is validated by a laboratory accredited by ENAC (National Agency for Accreditation and Certification).

14. Is the method approved by any international regulatory agency?
By now we do not have any international recognition, we are in process to notify to the FDA its uses and applications to be exported to the United States. It is worth mentioning that some components have already FDA approval.

15. Should it be applied on a dry surface or can it also be wet?
It is recommended to be used on surfaces newly sanitized, so if the application is immediate to the sanitizing process, it will be slightly wet. Our product is viscous nature so preferably on dry surfaces the contact is immediate. However, the product contains humectant agents wich allow it to cross a light coating of water, & to contact and react with the microorganisms that meet the above requirements.

16. If I go into a tank and apply Biofinder, to delete it completely, It is enough a rinse for 10 minutes? And in open surfaces?
Yes, It would be enough in CIP systems, and in open surfaces would be enough to do a normal rinse, with abundant water and pressure, such as the one made after the application of a common detergent.

17. Biofinder, could it leave toxic residue after rinsing?
No, if the rinse has been appropriate, since the components in the percentage that are in the formula, and according to the current legislation, are not considered dangerous and with a normal rinse should not remain traces, since the formula is highly soluble in water.

18. List of surfaces tested with BioFinder?
- Stainless steel.
- Polypropylene.
- Epoxy paint-coated surfaces.

This does not mean that Biofinder can not be applied on other surfaces, such as:
- High density polyethylene.
- Polytetrafluoroethylene.
- Polyvinyl chloride.
- Polyurethane.
- Borosilicate glass.
- Ceramic.
- Textiles (polyamide, cotton, polyester, and nylon), etc.

It should not be used in painted or lacquered wood surfaces because it discolours them.

19. Is the surface roughness a factor to consider?
No, the product has been tested on usual surfaces in the food, pharmaceutical or hospital industry. Rougher surfaces are ideal reservoirs for the adherence of microorganisms, which grow in the form of cluster in the cracks. Our product, being a slightly viscous liquid has no difficulty to penetrate the roughness, therefore, the reaction is displayed in the same way.

20. Does it work on textile surfaces with microbial contamination?
Yes, it will react if there is an enough group of microorganisms, a biofilm, above 104 catalase-positive bacterial cells.

21. If we apply the product to a floor and not bubbles (floor of a bathroom that is not clean) what can be the explanation?
That there is no enough amount of catalase-positive microorganisms.

22. In cold areas would it take longer to react?
Enzymes have an optimum temperature of reaction. Below the optimum range its activity slows, and above, as a protein, comes to a point at which is denatured and completely loses its activity on its substrate. In this case the substrate is contained in the formula of BioFinder hydrogen peroxide.
There is a study which mentions that the catalase of Micrococcus luteus (industrial enzyme) is optimal at 10 - 55°C with relative activity of 95-100%. Above 55°C the activity begins to decrease to its total loss as the temperature increases.

23. Should the bubbling be spread throughout all the solution, or would occur at a single point to be considered positive?
Foaming will be observed at the point where the biofilms contamination exists. Therefore, it should not necessarily be throughout the whole area where you applied the product.

24. The spray angle relative to the surface, can be any or is there a suggested one?
You just need to softly spray on the surface, to avoid bubbles by applying it. You can do it on horizontal surfaces with 45° angle, or on vertical surfaces directly, with an angle between 45-135 °.

25. Can I use any spray? I am thinking in the event of damage to the spray of the BioFinder.
We recommend a similar spray, i. e. , apply it in the form of Jet (not spray), having a way to regulate the output of the product and a total seal.

26. Which factors affect the activity of the BioFinder?
An unappropriate storage, as the exposure to high temperatures or sunlight.

27. Since when is the product on the market?
BioFinder was released in Spain on October 18 in the KAUSAL 2012 Congress, wich deals with topics about food safety and self-control.

28. Is the explanation that the reaction is slow because of the metabolism of microorganisms?
The reaction will be slow only if the environment is very cold, it is known that some microorganisms have the ability to grow at refrigeration temperatures. The ideal is to apply BioFinder to ambient temperature, i. e. , the reactant is not very cool (approx. 20 - 25° C).

29. Does Biofinder react with “Beer Stone” (calcium or/and magnesium oxalate)?
If there is “beer stone” and reacts with BioFinder it will be certainly because in it there will be organic salts that have worked as substrates to attach and feed microorganisms forming biofilms. Beer stone, is usually composed of salts of oxalate, which can be of calcium or magnesium, but can also be other organic salts. This causes the growth of microorganisms attached to them.

30. Can we say that Biofinder can be a verifier of the quality of disinfection?
Biofinder is a verifier of the absence of biofilms in a naturally-aerobic environment (common in food industries) . and, by consequence, of the whole procedure of sanitizing.

BioFinder is a complement for the control of hygiene within the programme of cleaning and disinfection. Its use is recommended before starting production (when it has been interrupted during the weekend) to verify the absence of biofilms in the critical control points either to control those critical control points where there may be strains with ease and persistence to develop biofilms.

31. How many applications or analysis can be done with a bottle of BioFinder?
The number will depend on the surface that you want to cover, for example:
5 ml of Biofinder (6 sprays aprox.) can cover a surface of 10 x 10 cm (100 cm2). Therefore, 1 bottle of 500 ml = 100 applications approx. max.

32. On laboratory surfaces, where generally we have contamination of 1 or 2 colonies (bacteria) and a maximum peak of 5 UFC/contact plate, BIOFINDER will react?
Preliminary tests show us that it doesn't work with isolated cells. It is not enough quantity to assume that there is a problem of formation of biofilms, which is what would give us food security problems. However, in a contact plate a colony does not necessarily indicate that there was a unique microorganism on the surface. It can also be originated by a biofilm.

33. In the hospital or clinic area, there are not the same “target” microorganisms than in the food industry, for example: Streptococcus and Enterococcus, which form biofilms in medical instrumentation. Then, BioFinder would not react with biofilms of these catalasa-negative microorganisms?
While there are microorganisms catalasa-negative, the hypothesis is that biofilms developed naturally in an aerobic environment are mixed. Therefore there would be minimally some positive catalase species that would allow us to visualize the reaction. Annex find an example of equipments or machines colonized by mixture of microorganisms, both positive-negative catalases.

34. In the dairy industry environmental microflora is predominantly formed by catalase-negative bacteria, such as the lactic acids (Lactobacillus spp. , Pediococcus SP. , Leuconostoc spp. , Enterococcus spp. , Streptococcus spp. ), BioFinder, would react?
Lactic Acid bacteria are bacteria that require little oxygen to reproduce (facultative anaerobes). The first three produce problems of alteration in the brewing industry, the fourth is more common, and it is indicator of poor hygiene practices (E. fecaelis), the fifth (S. Agalactie) gives mastitis problems. As in the clinical sector (above), these will rarely originate a biofilm, in an aerobic environment, they will usually be mixed with other microorganisms. This way, they could be detected with BioFinder if in the mixture there would be aerobic catalase positive microorganisms.

35. There are rapid methods in market that leave stain on the facilities, would BIOFINDER stain surfaces?
NO, Biofinder, as opposed to other methods based on coloration, does not leave any type of stain.

36. Is BIOFINDER a patented product?
Yes, Biofinder is international patent pending. Currently we have detected some manufacturer seeking to commercialize a similar product, they are trying to copy, fact that we have reported for infringement of our patent. It is important to note that copies and imitations do not provide any guarantee and they lack of scientific studies that support their efficacy.

37. Does the transport and storage of BIOFINDER need any special requirement?
No, Biofinder is not considered hazardous, therefore doesn’t require ADR transport, and storage must be in a dry place, keep it at a temperature environment at 20°C, and avoid direct sunlight on the container.