David run the first test in Mexico. We used Red Florescent Protein because it's easy to see the result. The bioreactor ran for 16 hours and it produced that red-orange color that you see. The induction (blue light on )is visible in this video.
click to see the video
There were several issues but none was a showstopper.
The most important was a broken hose in the circulation peristaltic pump. While that came with the pump we are unnecessarily ran the pump at full power about 30% over the max power. The next version of the software will allow up to control the power so it should be fine.
We also had some mechanical problems but we think with a little bit of adjustment and adding some washers to the motor screws will be fine
The software lost internet connectivity at last once so we lost some of the cool graphs. We think we'll might be able to address that in the next software revision. If not we'll find a solution to cache some data.
Hello Everyone! We are very happy to announce we have been selected to receive additional funding from JOGL through the “Proteopresso” project. In recent weeks we have been working, a bit in the background, in order to develop a new automated protein purification system or the “Proteopresso” for short. That device will complement the downstream processing for protein purification generated via the Bioreactor by using different low cost tags including the glass binding tag proposed by Alex in an early discussion at the Bioreactor channel.
In the next days David will send an invite to the key members for a video meeting and of course everyone else is welcome to participate! JOGL page for “Proteopresso” project JOGL page for “Proteopresso” grant application
In recent weeks we have been working, a bit in the background, in order to we recently submitted to the JOGL panel.
We held several conference calls with Vlad B. He will help us with the potentiostat and probes. I received from him a package with material including some different probes that will be used to validate the potentiostat.
I asked DigiKey about how to order individual specialized chips as they come in rolls of hundreds.
Rick provided a new iteration of laser-cut wood rocker parts. They are under quarantine now.
We have five populated boards ready to be used or sent.
Sasha made improvements to the software.
Sasha tested the boards so now they are ready for shipping. We held on the shipping in the hope that we can add a kit for the rocker.
Ordered several more parts and got some more. Among them, the essential air intake filters.
Got the peristaltic pumps, so we have the basics for the rocker.
Sweeta is interested to join and help with UI design and I had an introductory meeting with her.
Alex finalized designing primers for the second round of reagents. He put a new order for these primers. He is looking at testing the Optical Density sensor.
Had discussion with David and started looking at creating a vertical gel box for proteins separation
David is working on lysis solution. He got the parts and is experimenting with the piezoelectric mist generator.
Alexandra re-became available and will be starting on documentation.
Presented our project and team progress at the JOGL weekly call.
I added Narcisse to gitHub so he could add the UI code.
Designed several incremental prototypes for a laser-cut rocker variant. Rick helped with cutting some MDF parts.
Got the cooler part for the PCR machine.
Additional communication too much to mention.
Updated the website.
Previous status:
Adrian Jones populated three PCB with virtually all elements. I got the boards from him, and planted several H-bridges (for optional Peltier elements and optional 2 way control of the peristaltic pumps) so the new 3 new boards look pretty much complete.
I passed the boards to Sacha so he can to test the output signals and actuators and sensors connectivity which he has bee working on for the past two days.
Adrian Jones came up with an improved, solid design for a part of the rocker. It's also better centered so better balance.
Here is a picture of the CNCed rocker module. Great craftsmanship.
I found a box and still looking for a platform for it to unit test it.
Ordered a bunch of high geared motors from Ali to test if they can be used as a replacement for the Nema 17 and allow using Charlotte's design.
Sasha is working on checking the new 3 boards we got from Adrian. This is the first major check with our custom boards.
Discussed and made some slight modifications to the PCBs. They are done manually.
Distributed some of the grant money to cover some expenses
I got a big shipment from Alex with all kind of goodies. Still sorting through it but it has the Blue light inducible organisms and IPTG inducible organisms.
David is working on lysis solution. He ordered and just got both beads and mist piezoelectric parts.
He will start testing them. We need a protocol for testing lysis efficiency so we can all follow it.
David also presented our project and team progress at the JOGL weekly call.
We need to send some protocols to Therese so we can continue work on the paper.
I added Narcisse to gitHub so he could add the UI code.
Scheduled call with Shura to give him more info on the bioreactor.
We need to find something concrete for Alexandra so she can participate in the project
I ordered a piezoelement and board for a sonicator. We need access to a metal lathe to change the shape of the vibrating rod.
Updated the project page.
Modified a file for a laser cut an orbital Shaker that so it can take a Nema17. It's needed for the preparation of the starter culture. I sent it to Rick hoping he can find a laser cutter. I'll probably modify it for a CNC machine as it's very similar to the rocker too.
Some additional communications too many to mention.
Previous Status:
Created a starter culture for PDawn RFP
Reviewed the design of a rocker by Charlotte and made some suggestions
Made a first basic bag with just the hoses for air
Designed, and 3D printed some elements for the exhaust air filter and built a filter using charcoal and some kind of aquarium filter and a conical tube
Doing a test using a substitute bag, air filter, DIY bubble dispersing system and an orbital shaker. The filters seem to work as there is no smell
Discussed with Sasha and Narcisse some UI elements and integration
Discussed with David methods to quantify protein production. Ordered a UV sensor just in case we go UV B-C
Ordered several batches of Nema motors after discovering that one of the distributors increased the price and canceled the shipment
Got one small shipment for PCB parts electronics so hopefully the rest will follow soon
Got a couple of more shipments of barbed connectors including one with elbow connectors
Made a DIY autoclavable air bubble dispersing solution as the usual stones cannot be autoclaved
Started a basic test using a substitute bag, air filter, DIY bubble dispersing system and an orbital shaker. The filters seem to work as there is no smell. So far the air dispersing system also works but we don't know how efficient it is.
Created several injection ports by simply injecting generic silicone in the silicone hoses
Discussed with Sasha about doing a test on the 3D printed rocker this week and prepared the bag for it
Sasha managed to find a better plugin for the ESP and worked on the calibration
Started to prepare a package for Alex in Toronto
Alex is working on the last leg of the cloning. He also managed to order a primer that was somehow dropped from the initial order
Adrian Jones made some suggestion regarding PCR thermal solutions and light sensors. I ordered a cooling unit
Many more discussions with David about strategies for downstream processing. Mexican border control stopped his shipment of glass beads for some unknown reason so we'll need to find another solution for lysis using sonication electrophoresis or something else
I ordered a couple of 1 kg weight sensors as a future potential addition
So things are moving, some faster than others but they are slowly getting the shape
Started the redesign of the OD sensor box as I want to send one to Alex
No progress with the Oxygen sensor or rocker and I discussed with Thomas Landrain the problem with the grant funds that are still not transferred. He said he will check again with the French transferring bank as the systems are different with European banks.
Here is the test running now. It verifies the air including filter and bubbler.
Here is the video
Biorector filter - I printed and I used acetone to strengthen layers adhesion. Then impatiently I put it on the conical tube before the acetone evaporated so you can see fingerprints.
Previous status:
Primers for genetic engineering of the first 3 reagents RT, pfu polymerase, and TAQ received.
Alex is working on the genetic engineering of the three plasmids. He is using a Gibson assembly protocol and he already prepared the inserts. We expect to have the production organisms within one week.
Funds from JOGL still not received yet. They were supposed to be in at the 'end of this week' according to the communication. I sent another message to the JOGL president.
Most essential parts have been ordered. Notable parts missing are temp sensor sleeve, peristaltic pumps (we have one for OD which is enough for the prototype), air pump (not decided yet on the final 12 volts pump as David is trying to determine the min needed throughput). We have a 5 Watt aquarium air pump which is what the other Open Source projects are using. No output air filter so we need to design a solution based on an another project which I know and might require some minor 3D printing.
Looking at a solution for a DIY Oxygen sensor and the only one that is affordable seems to be the one suggested by Cambridge UK/Hackster which is a DIY solution with gold/silver electrodes encased in some autoclavable material.
Narcisse, a developer that joined last week is looking into the Blocky JavaScript library to capture the 'program'.
David is working on the post processing. He researches the list of equipment and reagents to buy for that for all labs. I'm looking of procuring the glass beads to crush open the cells. They seem to be expensive at this point so I might look into alternatives like sonication of elecroporation. Unfortunately they all require extra devices.
Adrian J. finished designing a prototype board for a PCR machine to be included as an optional device to assist in testing. The PCB was sent to China for manufacturing.
Alex will send me the special cutter to make the holes for the tubes in the aluminum block. That is one of the most critical parts.
Alexandra W joined our team. She finished her freshman year majoring in Applied Mathematics & Biology. She volunteered in an electrical and computer engineering lab and was on the build group of a robotics team for four years. She spent a month at Apple's headquarters focusing on a secret product design last summer.
I asked THE-ODIN if the can send us a pipette filter while waiting for our 50 pieces to come from Aliexpress. They said they can. I need to put an order in.
Sasha worked on back end program and now we can change wi-fi settings from web page and they will be stored in flash memory.
Misc additional communication with other projects/stakeholders.
I also made an application with an Ontario program called Ontario together which is a long shot however they send me another form so it was not rejected yet but they give funds to businesses so I don't think it will work.
Updated the website
Previous Status
Primers for genetic engineering of the first 3 reagents RT, pfu polymerase, and TAQ ordered
We got temperature sensors and pH sensors courtesy or Adrian Jones. Many thanks for that.
Funds from JOGL still not received yet. They were supposed to be in at the beginning of this week, I sent in a question. Apparently they can not find the request.
Most essential parts have been ordered. Notable parts missing are temp sensor sleeve, peristaltic pumps (we have one for OD which is enough for the prototype), air pump (not decided yet on the final 12 volts pump as David is trying to determine the min needed throughput). We have a 5 Watt aquarium air pump which is what the other Open Source projects are using. No output air filter so we need to design a solution based on an another project which I know and might require some minor 3D printing.
Sasha integrated the temp sensors and pH sensors.
Here is the video
We have the 3D printed rocker that needs redesign. We tested it at 100 full cycle tilts per second with platform inclination at about 30 degrees. According to information collected by Alex and me, a professional rocker can do max 85 full cycle tilts with a 2 KG (to confirm this weight) silicone bag. The industrial rocker for the wave bioreactor in the comparison spreadsheet works at about 10 degree angle so that should be our target. The rocker ball bearing gets undone in the video but most likely it will be fine at 10 degrees and 85 full cycle a minute.
Here is the video
We still need to redesign the rocker and cut it. I'm still hoping one the many 3D collaborators that promised to help with that will come through. If not I'll have to do it. As the laser printers are not available so we need to look at alternatives.
Alex has started a test batch with a silicone bag and rocker at 85 cycles a minute but without air pump or heating and he got to .4 OD which is about 70% of what a normal manual procedure yield in controlled temperature in terms of bacteria. That is pretty much expected. He is working on extracting the reagents so we can see the actual yield in terms of reagent which is the important indicator of efficiency.
I contacted a number of people for help with UI and CAD design and about 6 were very interested in our project and helping with it and joined or Slack channel. So far, only Serge started a closer discussion so we'll see. Here are some of the names:
Moritz, AdriSolid, Serge, Hammed, Faical.
We started a thread about adding testing and tuning additional capabilities with a small PCR using a similar concept as the main board. Initially we were thinking to just reuse an existing Open Source design as is, but at this point we are looking at existing small/inexpensive Open Source PCRs that can be converted to our ESP/ Client UI driven concept.
Good, long discussion with Nico about a potential submission of a new project potentially using Yeast instead of e-Coli and using large containers. We should still be able to use the same electronics as the wave bioreactor and the same plasmids.
Aggregated the requests from David and Alex for requests on Free Genes collection that I have from Stanford. Poured about 20 Ampicilin plates and several Kanamycyn plates so we can can send to Mexico and Toronto.
More research on the Oxygen sensor. A good paper at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6767127/
Contacted Chiara @Cambridge University which was the lead on the https://www.hackster.io/biomaker/low-cost-oxygen-sensor-for-bioreactors-c7e066 project. She told me there was no additional work on that. I asked for additional info. She sent emails to two other people asking them to contact us. The project requires gold and silver wires and a potentiostat https://hackaday.com/2011/09/14/cheapstat-an-open-source-potentiostat/.
Participated and presented project in the JOGL 1.5 hour long weekly meeting.
Our Slack channel. Ask for an invitation link and then join it and leave an introduction in the introduction channel detailing what part you are interested in. Unfortunately the invite link expires so we cannot post it in here
Look at rows 2, 17 and 19 to understand the principles, the functionality and see different implementations
Open Bioeconomy Lab bioreactor [2]
A re-usable wave bioreactor for protein production in insect cells - very close to what we want [19]
Please read before asking questions as many answers are already present and we want to avoid duplication of information and unnecessary effort.
To understand the project you could either look at our proposal for JOGL project link or read below. The proposal is more complete.
Detecting pathogens like the SARS_COVID2 virus or create treatments like vaccines require reagents (chemicals).
In order to run the reaction you need reagents for each test. Reagents are expensive and many are unavailable due to demand. There is an imperious need to create an alternative cheap source for these reagents. There are several labs working on this, notably the free genes that are doing a great job pioneering this. Thanks to them.
The only way to produce those in usable quantities is to use a bioreactor. Commercial bioreactors cost thousands. That does not reflect manufacturing costs. Existing open source designs are not usable, abandoned prototypes or work in progress.
That's why this this is important. A cheap alternative to commercial bioreactors that would cost less than 200$ would solve this and also make possible the rest of the other initiatives.
In a nutshell,a bioreactor is a container with agitated liquid and control of temperature, Oxygen, PH etc.
Things get a little more complicated when you want to grow mammalian cells like for production of antibodies of some more complicated proteins.
And a little bit more complicate because of the need to be sterile and reusable.
This is Not a One Day Wonder or Hip Vaporware project but a project that was started before the pandemic and will be relevant and offer value to humanity after the pandemic too.
Research and work on modules of this project has been started last year before this pandemic so this is not an opportunistic project nor a project started out of the blue with little experience and little chance of success.
We are basing some of this on great work done by other groups in the academia, DIY and industry.
We will be building a wave bioreactor based on several original ideas.
We are not trying to compete with other Open Source initiative but rather collaborate and provide better solutions and implementation where possible.
We will not spend too much time on great looking web pages, formatting a spelling but try to trim the fat ad focus on delivering. So formatting and typos are not going to be a priority. Sorry, no spell checker in this free editing software.
As you can see when looking at the equipment, this is not our first project and our finished projects are functional and in use.
Our first main target is Reverse Transcriptase. That is the first enzyme needed in the CDC virus detection. A reverse transcriptase (RT) is an enzyme used to generate complementary DNA (cDNA) from an RNA template, a process termed reverse transcription. We will of course run RFP bacteria first to test and optimize.
