Fed-batch setting with sampling

[Bioreactor]

Hi,

I’m working on a project where I would like to characterize a bunch of strains in a fed-batch fermentation mode. Is it possible to use Chi.Bio under these conditions? I’ve been trying to read a bit about the main fermentation settings and seems to be designed to be used as a chemostat/turbidostat.

Also, my experiments would require taking multiple samples for different purposes, meaning that I would need to quickly sample the reactor several times. Is there an easy way to quickly sample the reactors (I would need to know the EXACT volume drawn, very precisely). Can they be coupled to an automated sampling system?

Best,
Bioreactor

[harrison]

Hello,

It is possible to use Chi.Bio as fed batch, but the challenge (as you have identified) is getting liquid addition/removal to be highly accurate. We used low cost pumps in the system to reduce price, so they cannot accurately deliver small volumes. One thing you could do is use some different pump that runs separate to the core platform, and just set it up to remove liquid from the reactor (e.g. with a thin needle through the lid) over time.
As to automated sampling, there are some commercial approaches to this and also DIY ones. See for example this toolbox which works with Chi.Bio
https://www.nature.com/articles/s41467-022-31033-9
Or this approach which uses a liquid handling robot and could be similarly adapted to the Chi.Bio reactors for precise and automated sampling:
https://www.biorxiv.org/content/10.1101/2022.06.13.495893v1

Harrison

[Bioreactor]

Thanks for the reply!

In order to use Chi.Bio in fed-batch mode, what do you think would be the easiest approach?

The idea would be to run a batch phase until glucose is completely consumed (all pumps switched off in batch phase, no feeding at all). We already have some data on the time it takes the strains to consume all the glucose, so probably we could use experiment time as trigger signal to turn on pump1 and start feeding. The goal is to follow an exponential feeding profile (F = F0 * e^(k*t), where F0 is the initial feeding rate, k is a constant and t is the feeding time), but I’m not sure how easy it would be to establish that in a custom program. We would of course need to estimate feeding rates for each of the pump1 as I’ve read quite some variation can be expected, but I’m not sure if we could face additional issues. Maybe if we reach high cell densities oxygen becomes a limitation too?

Regarding sampling we could just stick a syringe through the lid, even though it could be time-consuming for many samples I believe it’s still doable.

[harrison]

Hello,

Yes, what you propose sounds sensible. You would need to calibrate each pump to get the conversion factor between set point (e.g. number from 0.0001 to 1) and what this actually means in terms of mL/minute. Then, it would be fairly straightforward to implement in a cusotm program. In short, make a custom program which when activated turns the pump ON and starts incrementing some counter (i.e. some number that starts at 0 and each minute/algorithm-iteration you add 1 to this). Then, have the Pump1 (or whichever pump) set point update each minute depending on that counter value. Thus, you would have a mechanism which starts counting when you press “go” on the custom program, and counts upward (i.e. increases pump rate) according to whatever formula you desire, until you turn it off.
Note that if you are adding some liquid over time you will also need to make sure you are removing it at a greater rate (i.e. set pump2>>pump1) such that the system does not overflow!

I would see the major potential weakness of this approach being if you wish to add very small amounts of glucose at low concentrations which would mean (because the pumps will not effectively deliver volumes significantly less than ~0.1ml) the batch culture being significantly diluted over time – but maybe this is OK for your application?

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