Lexagene to have prototype of on-site testing device by end of year
LexaGene is working on the automated pathogen detection platform for use at the site of sample collection.
End-users need to collect a sample, load it onto the instrument with a sample preparation cartridge, and press ‘go’. No pipetting or molecular expertise is required.
The patented microfluidic system was invented by company founder and CEO, Dr Jack Regan.
It will process six samples at a time, returning results in about one hour.
Processing of large volumes to see risk
“You look at food safety and, in our opinion at least in the US, there is a great opportunity for new technology to be introduced and we think it is going to be PCR-based,” Dr Regan told FoodQualityNews.
“A platform that can be placed at sites where the food is packaged and our technology is well-suited to that for one reason: it can process large volumes of fluid and this is unique amongst the platforms out there.”
Dr Regan said because of this it can get around a problem which has ‘plagued’ food safety - sub-sampling error.
“This is where the food item may be contaminated but you may miss it because you only analyse a small portion. If you want to minimise the chance of false negative results you need to process more of the sample,” he said.
“We realise that we have an uphill battle in many regards because the food safety industry has been based on determining plain viability so everything has gone through collecting a sample, putting it into an enrichment broth, letting it grow for 24 hours or more and then doing your analysis.
“This is the only way to prove viability but what food safety is all about is mitigating risk and we believe that there is an opportunity to provide a technology which looks for the DNA of certain pathogenic organisms, this is not a test for viability but it is a test for risk assessment.”
Dr Regan said it is up to the end user on how they handle positive results.
“If you detect Salmonella DNA on spinach, you don’t expect to see it there, spinach is treated differently than a beef carcass where they might spray it with decontamination fluid that kills bacteria. We would think the thing to do would be to say ‘ok this is a high risk food item and we are going to send it for confirmation testing’,” he said.
“Our goal is to provide information to allow the food safety officer to make a decision as to what is in the best interest of the company and the customer. The company has to mitigate the risk, they don’t want a recall due to the economic impact and damage to brand name recognition.
“If there is nothing detected DNA-wise you can be pretty sure that culture is not going to grow anything up because PCR is one of the most sensitive detection technologies out there and we would think they would say there is a low risk here so we are going to send to the customer.”
LexaGene’s technology uses single-use disposable cartridges to concentrate and purify genetic material from samples and microfluidics to perform molecular analysis.
Each sample is screened using 12 tri-plex TaqMan reactions. Included in the test are 12 positive and two negative controls to give the operator confidence in results.
Preliminary targets include things like fruit and vegetable wash.
“Our hope is that at the end of a packaging line where spinach is being packaged, you take some of the wash used to wash the spinach and you analyse it and look for E. coli DNA of the pathogenic variety or Salmonella and if you detect a high amount you would flag that as a high risk batch and send it off to culture,” said Regan.
“If you don’t detect any then you would deem the risk to be extremely low and send directly to the customer. This provides a cost advantage for the packaging plant and the producers because they don’t need to keep their food in storage while they await a result back from culture which can take several days.
“They are allowed to deliver a fresher product to the customer which has a longer shelf value and is more valuable than product that is thrown out because it expired before it was purchased.”
Open access approach
The open-access feature will allow end -users to target any pathogen of interest, as they can load their own real-time PCR assays onto the instrument.
“No other platform on the market today is open-access, they are closed-access, because you buy the instrument, you buy the cartridge, the cartridge has reagents in it that detect a set number of pathogens or gene sequences and if you want to detect something different you can’t use their technology,” said Regan.
“In contrast, our technology allows the end user to configure the reagent cartridge the way they want to so if they have a particular assay they have been using for a long time they can take those assays and run them on our equipment and this allows for them to have continuity of data and opens up our technology to a market need which is not addressed.
“We are very interested to see how our technology is received given the fact that the end user has more flexibility because we can create these customized tests to screen for what they are interested in because every customer may have a unique need.”
Regan said there is a niche in food safety that has not yet been filled.
“All the other molecular platforms out there are more designed for the food contract labs where you have expert people to run the tests, in contrast our instrument is designed to be so easy to use, that factory personnel with minimal training can be taught to use it,” he said.
“It only requires collecting the sample, loading the sample onto the instrument, loading a cartridge onto the instrument and pressing go and that is it; just three steps to run the instrument.”
Product development timeline
The first working prototype is expected by November.
“At that point we will evaluate the instrument, see how it has performed - and that is going to be the alpha-prototype,” said Regan.
“We will then spend about five or six months where we will make small tweaks, improve the software and the assays that are running on the instrument and then send the beta-instrument out to customers to evaluate before finally building a production unit that is sold. We expect to be on the market before the end of 2018.”
Dr Regan said feedback from potential customers is important to make sure it is addressing their needs.
“We are going to be looking to identify beta-testers who would test our equipment free of charge, we would ship them the instrument and reagents and they would have free use of that instrument for a period of time with the anticipation they would provide feedback at the end to help improve the product.”
An important thing for food safety is how many different genetic targets can be looked for at once, said Regan.
“Our platform is going to look for 22 different genetic signatures. Bacteria are everywhere in the environment and that is not associated with disease but they are an indicator for how clean the process or equipment is,” he said.
“Because we can look for 22 different genetic signatures at once they will be able to look for indicator species, assays for E. coli, Salmonella and Listeria.
“When you look at detecting Salmonella you might have three or four different assays for Salmonella on the system, each assay would target a different region of the Salmonella genome and so at the end of the test if five of five come up positive you know you have Salmonella.”
Lexagene will develop validated assay panels for targets it believes customers are interested in screening for.
“For customers who want to do fully customized screening what we are going to do is provide this reagent strip. What would be in that strip effectively is reagents for the positive controls but they would add on top of that their own assay that they are interested in so they would add the 22 different assays for the indicator species or pathogens that they are interested in detecting,” said Regan.
“There are 12 tubes in this strip, each tube contains reagents, these are PCR assays looking for particular targets but each tube is what we call triplex – it looks for three different genetic targets.
“Because each tube looks for three, that is three targets times the number of tubes (12) so that is a total of 36. So every time the instrument runs a sample 36 different PCR reactions are being run, of those 36, 22 look for either indicator species or pathogens, 12 are positive controls to make sure the PCR works and two are negative controls.”
Microfluidics and PCR
The microfluidic system is not suited for big particulate matter or dry products.
Regan said down the line it expects to target more challenging food matrices.
“Right now the standard in the industry for looking for E. coli contamination in beef is to take 25g of beef and put that into an enrichment broth, let it grow overnight and from that you analyse it,” he said.
“After an overnight culture, LexaGene’s technology could very easily be used to analyze some of the broth for pathogenic organisms.”
One of the instruments main functions is PCR. The speed of PCR is mostly determined by the volume of fluid being heated and cooled and how quickly it can be done.
“Our instrument is a microfluidic instrument so the actual individual PCR reactions that we are performing are only about 1uL in volume, they are very small, which is beneficial because it reduces the cost of performing a test but it also allows you to thermocycle more quickly to so our goal is to do the PCR step in about 30 minutes. So we would do 40 cycles in about half an hour,” said Regan.
“Then the question is how long does the sample prep take and assembling the assay, which is the upfront portion of it. We anticipate that taking about 30 minutes for a low volume (around 500uL) sample to take that in, concentrate it on the filter, purify the DNA and RNA and then assemble the reactions.”
Lexagene said for a small sample it would be about one hour to get a genetic result and processing a larger volume would add time.
The system will have pressure sensors so when a sample is drawn over the filter, if it is laden with particulate matter and the filter starts to clog the sensor will detect that and the instrument would indicate there is low fluid flow across the filter and would stop drawing in the sample and process the sample as is.
The integrated device in a box performs sample preparation so concentrates and extracts the DNA and assembles and performs the PCR reactions.
“So you can imagine taking a box, placing it in a food packaging plant and having a set of cartridges beside the box. The box has on it some buffers and reagents that would need to be maintained by the end user but they would only have to be changed every couple of months depending on the frequency of use,” said Regan.
“You can anticipate somebody grabbing a sample, loading a sample onto the instrument, grabbing one of the cartridges, which is next to the instrument, loading that onto the instrument and pressing go. There is a touch screen monitor so the data is available about one hour later.
“The instrument will generate trending data places like food packaging plants will want to see, on the levels of indicator species over time and the instrument will plot this data.
“So an auditor can come in and see how many tests were run, when the tests were run and what the level of indicator species has been over time and this is one stop for the auditor to say have they been doing the testing, how clean are their processes, what are they detecting and how did they handle it.
“It is a relatively sensitive instrument so it is not like you can spray it down with the hose at the end of the day. It is designed to be placed near the point of sample collection that might mean it is a 50 foot walk into another room, where they don’t steam clean the equipment.”
