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Documentation

This is where the Amybo documentation lives - lets keep all the learning available and usefully organised here. If you’re new you may want to start of with the:

Overview

Other content such as discussions, trial results, case studies, and community updates should live in the Community pages.


1 - Overview

How we hope to democratise food.

What is Amybo?

Amybo is a diverse community of people interested in unlocking the secrets of protein fermentation. Our aim is that anyone will be able to produce tasty, nutritious protein at a lower financial cost and much lower environmental cost than by raising animals to eat.

What are we doing now?

Initially, we have quite a list of questions to answer:

  1. Which single cell protein should we ferment first?
  2. What equipment will we need to ferment it?
  3. How should we share our results to best accelerate open source development?
  4. How can we ensure that the products we make are safe to eat?

You are invited to join us in creating pages on this site addressing these issues (and any other’s we haven’t yet thought of)

Join Us

1.1 - Next steps

Plan of campaign

Next steps

  1. Pick a protein producing microbe to cultivate
  2. Pick a bioreactor to cultivate it with
  3. Conduct initial risk assessment (each subsequent step will start with a risk assesment)
  4. Design/build/procure the bioreactor
  5. Dry commission the bioreactor
  6. Wet commission the bioreactor
  7. Live commision the bioreactor
  8. Cultivate the protein producing microbe
  9. Determine next steps required to produce safe nutricious delicious protein

2 - Proteins

What proteins should we produce?

Step one in protein development is deciding which protein to develop first. Choice of bioreactor, feedstocks and downstream processing are all dependent on the organisms we use and the proteins they produce. This page is very much a work in progress and your input would be hugely welcome.

A note on “closed source” enterprises

We mention a number of commercial “closed source” companies in these sections. We fully support their efforts to bring the benefits of alternative proteins. Commercial competition has brought many excellent innovations to market. Many companies seek to do good, even before seeking profit, however there is a risk that good companies and/or their IP gets acquired by less benevolent enterprises. Shareholder primacy means that some of the world’s poorest, in the most challenging environments may not benefit as much as they could from the innovations that they need most. Open source projects like this serve to benefit all corporations (as they can access our data) while also ensuring our innovations have the potential to benefit all.

Biotech Proteins

Biotechnology is providing amazing advances in many areas of food production including:

  1. Single-cell protein
  2. Precision fermentation
  3. Cultivated meat

These are currently ranked in order that Martin envisages attempting them, but if you are working on open science projects around any of these, please do get in touch.

Meat & Plant Protein

Plants are definitely a more sustainable source of protein than highly inefficient traditional meat. Only a fraction of the protein that livestock eats makes its way into the meat that we eat.

Plant-based protein production still requires significantly more land, and in many cases more water, carbon emissions, fertilisers, pesticides, herbicides, etc. than any of the biotech proteins listed above. Given the inefficiencies of current agribusiness cultivation, this results in significant discharge of nutrients and other environmentally detrimental chemicals into water courses.

There are many improvements that can be made to crop cultivation. For example The Land Institute are doing wonderful work on perennial crops. Many small (and not-so-small) scale farmers and horticulturalists are doing wonderful things around both increasing yields and decreasing environmental impact.

Climate change poses an existential threat, it is essential that we reduce GHG emissions from food production and all other activities. Even if we manage this we still need to adapt to climate change. There are already populated places on earth where natural crop cultivation is impossible. Microbial protein production could be sustainable in all such regions. Our challenge is to ensure that it is economically viable and available to all who need it.

Plant Based Meat

Tofu and Tempeh are examples of ancient meat alternatives. Beanburgers and veggie burgers took the next step in vegetarian fast food. More recently companies like Beyond Meat and Impossible Foods have taken great strides in meat alternatives.

We are inclined to leave plant based meat to well-funded enterprises, as they generally offer no nutritional benefits over the plants they are made from, and naturally requires plants, which have their own downsides.


2.1 - Single-Cell Protein

Which single-cell proteins should we produce?

Single-cell proteins (SCP), sometimes called microbial proteins, are edible unicelular microorganisms containing high amounts of protein. These can be farmed using biomass fermentation:

Biomass fermentation

Biomass fermentation is where the whole microbe (or a dried version) is the product of fermentation.

Given the relative ease of downstream processing when the whole microbe is used, and their gastronomic potential, biomass fermentation may be the preferred route for open source fermentation.

Most types of living microbe could potentially be used for biomass fermentation. We’ll break these down into fungi (yeasts and moulds), bacteria and algae:

Fungi

Fusarium venenatum

Quorn mycoprotein is a well known proprietary product of biomass fermentation. It uses Fusarium venenatum, a filamentous fungi. Quorn’s original patents expired in 2010 but their £30M fermentation towers may prove challenging for open source development.

Sustainable Bioproducts, Inc and 3F Bio Ltd have also submitted Fusarium venenatum GRAS (Generally Recognised As Safe) notices to the US-FDA.

Harrison Lab have published bioinformatic analysis of Fusarium venenatum genomes on GitHub.

Bacteria

Xanthobacter spp.

While we, and many others previously reported that Solar Foods use Cupriavidus necator to produce their proprietary Solein largely from air, their EU Novel Food application indicates that they are using a Xanthobacter species.

Solar Foods take carbon dioxide and water vapour from air. They use solar powered hydrolysis to split the water, providing hydrogen which the bacterium can use as its energy source. Ammonia is used as the nitrogen source.

Cupriavidus necator

Novo-Nutrients and Kiverdi are thought to use Cupriavidus necator. The full genomes of the H16 strain and JMP134 strain have been sequenced.

Algae

Through photosynthesis, algae use sunlight to produce sugar and oxygen from carbon dioxide and water. Nitrogen can be metabolised from urea, meaning all major feedstocks are freely available.

As such (and since a number of algae, containing all essential amino acids, are already sold as ‘superfoods’ for human consumption) algal protein may be the quickest win. That said, it may not be as gastronomically appealing as other proteins on our shortlist.

Arthrospira platensis

Spirulina is commonly used as a food supplement, but Spirulina Gnocchi has been proposed by the European Space Agency for Mars Missions. It has a slightly sweet nutty taste. Spirulina is technically a cyanobateria rather than algae, which means the risk of cyanotoxins (e.g. microcystin, alkaloids and BMAA) need to be mitigated.

Chlorella

Chlorella arguably tastes worse than Spirulina. They also have a lower protein concentration and cellulose walls. In their favour: they are single celled, so may be easier to process, and should not produce cyanotoxins.


2.2 - Precision Fermentation

What is Precision Fermentation and could we implement it?

Precision fermentation is where microbes are used to produce particular chemicals. Insulin and rennet are common cited products of precision fermentation.

Precision fermentation can be used to produce pretty much any chemical. Ingredients that are currently being produced using precision fermentation include:

Casein

Real Vegan Cheese are a non-profit research project using precision fermentation to produce casein - the main protein in milk, cheese, yoghurt and similar dairy products.

The separation processes required to extract the end products of precision fermentation will likely make it a more complex process than biomass fermentation. As such we anticipate that the proteins will be more expensive to make and somewhat harder to democratise.

Precision fermentation also generally involves genetic engineering. We anticipate that corporations who invest in the development of custom strains for precision fermentation may be less inclined to open source them, so we very much welcome the Open Science work of Real Vegan Cheese.

Rennet

Rennet was traditionally obtained from the stomach linings of calves and lambs. A very high proportion of rennet used in cheese making is now produced via commercial precision fermentation.


2.3 - Cultivated Meat

How is cultivated meat made?

The Good Food Institute and New Harvest have useful primers on cultivated meat and cellular argiculture. This may play an important role in mimicking and replacing traditional meat. However, the complexity and cost of doing so, without any significant nutritional benefits over single-cell proteins, mean that it may be best left to well-funded enterprises..?


3 - Equipment

Which equipment should we use?

Commercial competition is an established means of driving innovation. However, as with the microbes and processes, development of either open source or generic equipment may be necessary to ensure that those who most need them can afford them.

Using an open source equipment also enables you to both customise it’s capabilities, and contribute to it’s development.

General purpose lab ware should be relatively easy to procure, and toxicity and chemical analysis can initially be outsourced to commercial labs. There are some more expensive pieces of equipment to focus on:


3.1 - Bioreactors

Which bioreactors should we use?

Once you have determined which protein you wish to cultivate, the next question is what you are going to cultivate it in. There is a need for development of better and more efficient bioreactors across the board - but, given the nature of this project, it makes sense to, at least initially, restrict our scope to open source bioreactors.

Bioreactor Shortlist

Pioreactor

“The Pioreactor is an open-source, affordable, and extensible bioreactor platform.

It currently comes in a cute and cost-effective 20 ml version that can be used in batch, fed batch, continuous, chemostat, turbidostat & PID morbidostat modes. While the software is already open source, we understand from the founder that the hardware and 3D designs are due to be made open source in 2023.

Exciting features for Amybo (as of June 2023):

  • it appears to be more affordable than most automated bioreactors
  • it will soon be fully open source, hence customisable
  • it is clearly under active development
  • it is available for sale with excellent lead times
  • you can add LEDs as a light source for algae
  • the system is compatible with DormantBioLabResources’ open source modular lab racks

Wish list (as of June 2023):

  1. Ability to automatically upload results to a project cloud database
  2. larger vessels - so probes can be added
  3. Easy-open viewport cover (it currently requires 4 screws)
  4. 3D printer files - to minimise shipping cost & carbon, and enable community development
  5. Ability to control temperature below ambient
  6. Ability to wall-mount for bioreactor farms & video production
  7. Fully assembled units (including pre-configured raspberry pi’s) for technophobes
  8. Global stockists - to reduce shipping time, cost and carbon
  9. A way to be certain that reactor overflow isn’t going to fry your raspberry pi

Other Open Source Bioreactors

Hackuarium bioreactor

Watch also Open source bioreactor

Concerns (as of June 2023):

  1. last commit was 7 months ago
  2. It doesn’t appear to be commercially available although full assembly instructions and a detailed bill of materials are given

DIY Wave Bioreactor

Also see resources on Ottawa Bio Science Website)

Concerns (as of October 2023):

  1. last commit was 1 year ago
  2. It doesn’t currently have an active OD reader although plans for it have been articulated

Phenobottle

Watch Photobioreactor (Phenobottle)

Concerns (as of June 2023):

  1. last commit was 2 years ago

EVE-Pi

See also biorxiv article

Concerns (as of June 2023):

  1. last commit was 2 years ago

New Harvest Open Source Bioreactor

See also IRNAS

Concerns (as of June 2023):

  1. last commit was 3 years ago

Biomaker Open Source Bioreactor Project

See also Biomakespace and Hackster

Concerns (as of June 2023):

  1. last commit was 5 years ago.
  2. chat channel is a dead link

Other lists of open-source bioreactors


3.2 - Autoclaves

Which sterilisation systems should we use?

Open Source Autoclave Shortlist

This is a short list, there are a number of open source autoclave control projects:

But the only open source project that claims to deliver a full autoclave, is in our opinion just a thermal steriliser, as it doesn’t appear to pressurise the sterilisation volume.

It may well be that open source autoclaves are too risky. The probability of an explosion with an incorrectly constructed unit is high and the consequences could be lethal.

Given the high risk and low volumes of sales a commercial autoclave would likely be the most expensive item on our equipment list. So we should consider pressure cookers:

Pressure cookers

Electronic

Metallic Black GPC201SS-20 5.5L 1000W Pressure Cooker

Exciting features for Amybo (as of June 2023):

  • Low price
  • Can be used outside the kitchen (no hob required)

Concerns (as of June 2023):

  • Can it accurately monitor and control temperature and pressure?
  • Will condensation be an issue? (Autoclaves use dry steam)

Stovetop

Ikea 365+

Prestige

Other Open Source “Autoclaves”

OpenAutoclave

See also Open Autoclave: Build an open-source off-grid medical instrument sterilizer, Open Source Medical Autoclave for Developing World, Open Autoclave: a Humanitarian Maker Project

Concerns (as of June 2023):

  1. How can this maintain pressure, and if it doesn’t, how can it be called an autoclave?

3.3 - Microscopes

Which microscopes should we use?

Microscopes are third on the list of major open source equipment, because it is very possible to conduct fermentation without a microscope. It would however be much more difficult and less rewarding if you could never see your microbes. Microscopy also plays a valuable role in detecting contamination by unwanted microbes.

Fortunately there are a good number of open source microscopy projects:

Our Microscope Shortlist

Open Flexure

See also:

UC2

See also A versatile and customizable low-cost 3D-printed open standard for microscopic imaging

openFrame

PUMA

See also A Research Grade Open Source Microscope — Made with FreeCAD

Other Open Source Microscopes


4 - Experiments

What experiments have we run so far?

We do all our experiments in the open, and we try to make them as easy to reproduce as possible. We not only share our experimental protocols (like recipes for running your experiments) but also the results.

Here are some experiments we’ve done:

4.1 - Control Testing

A simple control test evaluating results between Pioreactors by Gerrit Niezen

To ensure that the Pioreactors we’re using are working as expected, we ran an experiment where we used the exact same experimental conditions with two Pioreactors, with the hope to produce the same results.

The original experiment is described on the Pioreactor website.

A bunsen burner, small weight scale, three vials and a dropper. Two of the vials contain YPD broth and are covered with aluminium foil. The other vial contains stock solution.

Kit List

  1. 2 x Pioreactor 20 ml - see our thoughts on Pioreactors
  2. 2 x Raspberry Pi Zero 2 W with header
  3. 2 x Raspberry Pi 15W USB-C Power Supply
  4. 2 x Micro SD cards
  5. 3 x 20mL vials
  6. Another computer on the same network as the Raspberry Pi with a web browser
  7. Pressure cooker to ‘autoclave’
  8. Aluminium foil to cover the vessel ports
  9. Distilled water
  10. Baker’s yeast
  11. Digital pocket scale
  12. YPD broth
  13. Measuring cup
  14. Dropper/pipette

Protocol

Preparing and sterilizing the media

  1. Prepare YPD broth media for three 15mL vials.
  2. Pour 300mL tap water into the pressure cooker, place the three vials on the metal trivet in the pressure cooker and turn to a high setting.
  3. Once the water in the cooker starts to boil, steam will come out of the open valve. Put the heaviest weight (15lb) weight on top of the valve.
  4. The steam will lift the weight and start to escape. As soon as the steam starts to escape, start timing the sterilization and turn down the heat so that the steam is only just escaping and not rushing out. Aim to maintain a gentle hissing.
  5. After 20 minutes turn off the heat and leave to cool.
  6. Wait until pressure is completely reduced then lift the weight off the valve allowing any remaining steam to escape. Never open the pressure cooker until the steam valve has been opened to release the pressure.

Using a pressure cooker on a gas stove as an autoclave

Running the experiment

  1. Dilute a small amount of baker’s yeast in 15mL of YPD broth media
  2. Innoculate the three sterile vials with the same amount (just a drop) of culture from the stock solution.
  3. Wipe the vials and place them in the Pioreactors.
  4. Start a new experiment on your Pioreactor dashboard.
  5. Select Manage all Pioreactors
  6. Start Stirring activity
  7. Start Temperature automation activity and set to 30°C
  8. Start OD reading activity
  9. Go back to the graphs and check that an Optical Density (OD) signal is being received.
  10. Select Manage all Pioreators again and start Growth rate.

Two Pioreactors standing side by side, with the one on the right connected to a Raspberry Pi 400

Results

The raw results from the experiment are available as CSV files on GitHub.

A screenshot of the Pioreactor results

Report

Watch Gerrit’s report his results


4.2 - Spirulina Batch

Batch cultivation of Spirulina by Martin Currie

Kit List

  1. Pioreactor 20 ml - see our thoughts on Pioreactors
  2. Super Bright White 5mm LED for photosynthesis
  3. 5mm LED cables to connect the LED
  4. Raspberry Pi 4B 8GB RAM - needn’t be 8GB, that’s just what I had from a prior project
  5. Raspberry Pi 15W USB-C Power Supply
  6. Micro SD - because Tom said
  7. Ethernet cable - just because we use CAT8 in the studio for anything in live production
  8. Another computer on the same network as the Raspberry Pi with a web browser
  9. PH0 Screwdriver
  10. Jewellers scales (to calibrate pipettor and weigh out growth media powder)
  11. 10ml Volumetric flask to make up growth media
  12. 10-100 ul Pipettor and 200 ul tips for inoculation
  13. Weighing boat
  14. Spatula
  15. Pressure cooker to ‘autoclave’
  16. cotton wool and foil to cover the vessel ports.
  17. Distilled water
  18. Spirulina inoculum & growth media - as it was the only Spirulina we could source in time.

5 - Risk management

How do we manage risk?

Amybo and its contributors accept no liability for anyone

If we are proposing potentially new methods of producing food for human consumption, and making those methods available to all, there is a significant risk to human health.

6 - Legal

READ THIS BEFORE TAKING ANY ACTION

DO NOT TRY THIS AT HOME

This website discusses untested and unregulated methods of producing food for human consumption - do not copy them or take any action without at least taking advice from your doctor and a team of experts who are able to review your plans, risk assessments and physical/biological/chemical setup and toxicity/pathogenicity/etc. testing protocols. Please also see our disclaimer.

6.1 - Disclaimer

Amybo is not liable for your actions or inactions

DISCLAIMER

Last updated July 06, 2023

WEBSITE DISCLAIMER

The information provided by Amybo, a trading name of andeye Ltd (‘we’, ‘us’, or ‘our") on amybo.org (the ‘Site’) is for general informational purposes only. All information on the Site is provided in good faith, however we make no representation or warranty of any kind, express or implied, regarding the accuracy, adequacy, validity, reliability, availability, or completeness of any information on the Site. UNDER NO CIRCUMSTANCE SHALL WE HAVE ANY LIABILITY TO YOU FOR ANY LOSS OR DAMAGE OF ANY KIND INCURRED AS A RESULT OF THE USE OF THE SITE OR RELIANCE ON ANY INFORMATION PROVIDED ON THE SITE. YOUR USE OF THE SITE AND YOUR RELIANCE ON ANY INFORMATION ON THE SITE IS SOLELY AT YOUR OWN RISK.

The Site may contain (or you may be sent through the Site) links to other websites or content belonging to or originating from third parties or links to websites and features in banners or other advertising. Such external links are not investigated, monitored, or checked for accuracy, adequacy, validity, reliability, availability, or completeness by us. WE DO NOT WARRANT, ENDORSE, GUARANTEE, OR ASSUME RESPONSIBILITY FOR THE ACCURACY OR RELIABILITY OF ANY INFORMATION OFFERED BY THIRD-PARTY WEBSITES LINKED THROUGH THE SITE OR ANY WEBSITE OR FEATURE LINKED IN ANY BANNER OR OTHER ADVERTISING. WE WILL NOT BE A PARTY TO OR IN ANY WAY BE RESPONSIBLE FOR MONITORING ANY TRANSACTION BETWEEN YOU AND THIRD-PARTY PROVIDERS OF PRODUCTS OR SERVICES.

PROFESSIONAL DISCLAIMER

The Site cannot and does not contain biotechnological/engineering/environmental/food/medical/health/legal advice. The biotechnological/engineering/environmental/food/medical/health/legal information is provided for general informational and educational purposes only and is not a substitute for professional advice. Accordingly, before taking any actions based upon such information, we encourage you to consult with the appropriate professionals. We do not provide any kind of biotechnological/engineering/environmental/food/medical/health/legal advice. THE USE OR RELIANCE OF ANY INFORMATION CONTAINED ON THE SITE IS SOLELY AT YOUR OWN RISK.

This disclaimer was created using Termly’s Disclaimer Generator.

7 - Contribution Guidelines

How to contribute to the Amybo community

Reach out to us

Please add a comment to one of our YouTube videos or drop us a line at hello@amybo.org with any suggestions or questions you may have - or just to say hello, it’s good to know people are reading this.

If you’re comfortable with GitHub (or would like to learn) we’d absolutely love it if you were happy to dive in and edit our pages directly:

Editing Amybo.org

We welcome contributions and improvements to the Amybo.org website. We want this to be as easy as possible so considered a wiki. However, given the controversial nature and risks associated with protein production for human consumption, we decided that an approvals process was required.

Since we’ll be using GitHub for software development, and potentially also for hardware and procedure development, it made sense to use this for community development of the website. If you struggle at all with GitHub development, please get in touch via hello@amybo.org

Web stack

We use Hugo to format and generate our website, the Docsy theme for styling and site structure, and Netlify to manage the deployment of the site. Hugo is an open-source static site generator that provides us with templates, content organisation in a standard directory structure, and a website generation engine. You write the pages in Markdown (or HTML if you want), and Hugo wraps them up into a website.

All submissions, including submissions by project members, require review. We use GitHub pull requests for this purpose. Consult GitHub Help for more information on using pull requests.

Quick start

Here’s a quick guide to updating the docs. It assumes you’re familiar with the GitHub workflow and you’re happy to use the automated preview of your doc updates:

  1. Fork the Amybo pages repo on GitHub.
  2. Make your changes and send a pull request (PR).
  3. If you’re not yet ready for a review, add “WIP” to the PR name to indicate it’s a work in progress. (Don’t add the Hugo property “draft = true” to the page front matter, because that prevents the auto-deployment of the content preview described in the next point.)
  4. Wait for the automated PR workflow to do some checks. When it’s ready, you should see a comment like this: deploy/netlify — Deploy preview ready!
  5. Click Details to the right of “Deploy preview ready” to see a preview of your updates.
  6. Continue updating your doc and pushing your changes until you’re happy with the content.
  7. When you’re ready for a review, add a comment to the PR, and remove any “WIP” markers.

Updating a single page

If you’ve just spotted something you’d like to change while using the docs, Docsy has a shortcut for you:

  1. Click Edit this page in the top right hand corner of the page.
  2. If you don’t already have an up to date fork of the project repo, you are prompted to get one - click Fork this repository and propose changes or Update your Fork to get an up to date version of the project to edit. The appropriate page in your fork is displayed in edit mode.
  3. Follow the rest of the Quick start process above to make, preview, and propose your changes.

Previewing your changes locally

If you want to run your own local Hugo server to preview your changes as you work:

  1. Follow the instructions in Getting started to install Hugo and any other tools you need. You’ll need at least Hugo version 0.45 (we recommend using the most recent available version), and it must be the extended version, which supports SCSS.

  2. Fork the Amybo pages repo repo into your own project, then create a local copy using git clone. Don’t forget to use --recurse-submodules or you won’t pull down some of the code you need to generate a working site.

    git clone --recurse-submodules --depth 1 https://github.com/Amybo-org/pages
    
  3. Run hugo server in the site root directory. By default your site will be available at localhost:1313/. Now that you’re serving your site locally, Hugo will watch for changes to the content and automatically refresh your site.

  4. Continue with the usual GitHub workflow to edit files, commit them, push the changes up to your fork, and create a pull request.

Creating an issue

If you’ve found a problem in the docs, but you’re not sure how to fix it yourself, please create an issue in the Amybo pages repo. You can also create an issue about a specific page by clicking the Create Issue button in the top right hand corner of the page.

Useful resources