[harrison]

Hi Timo,
Likely the problem is moisture dried on the moisture sensing tracks (parallel silver lines) on the top of the reactor.
Get a piece of tissue with ethanol/cleaning alcohol and wipe down these tracks thoroughly all around the top of the reactor then try again.

[harrison]

Hi Liliana,
You can turn on the three different FP measurements at the bottom of the user interface to measure the three different proteins.
The selection of filters/LEDs is complicated – it depends on both the emission and excitation spectra of the proteins. You need to ideally select ones that align with their peaks, but also since you are measuring them all together at the same time you will need to ensure that you are selecting filters in such a way to avoid too much overlap between adjacent proteins (i.e. you don’t want to be significantly exciting two at the same time).
This may take some trial and error.
Also, to improve your chances if you are measuring multiple proteins you ideally would use media with low auto-fluroescene (ie. don’t use LB), make sure the FPs are being expressed in fairly large amounts, and finally you would ideally pick proteins with long stokes-shift so that you can maximise the signal-to-noise (ie. reduce the amount of excitation light entering the emission filter).

[harrison]

Hello Albi,

Probably the easiest way would be to de-solder the laser diode that is in there at the moment and attach a laser at approximately the right wavelength you wish for.
In principle this is not so difficult – what you need to do is look at the schematics/part numbers for Chi.Bio (available on thhis site) to get the part number for the laser diode; then look up the datasheet to see what each of the three “pins” on the diode correspond to, and then find a replacement IR laser with the same “pins” and approximately the same output power.
Off the top of my head there should be quite a few options for laser diodes of ~700-800nm with 5mW power maximum (this is all you need, in fact less is fine) which you could be for $10 or less.
You could then solder in the laser onto the circuit board and use the exact same control circuit and all that which would control it (turn on/off etc).
Hope this makes sense!

[harrison]

Hi Katja,

Yes, you can use Filezilla to make copies of the .csv files and transfer them to another computer while it is running to check.
If you are doing a long experiment it might be that it is not showing the pump rate since it was so brief a time – in that case yes I would recommend looking at the csv file to check.

Furthermore I strongly recommend you put the tape in the pump heads as detailed in the troubleshooting manual.

[harrison]

Hello,
Baseband normalisation is essentially calibrating for the excitation intensity as well as OD. Therefore, the baseband-normalised data should stay approximately constant if one was to hold fluorescence fixed and change OD.

If you are allowing BOTH OD and GFP/mScar to change at the same time, then you MIGHT see a reduction in fluroescence essentially because the fluorescence/cell is decreasing. This often happens if you have a system in which it has grown overnight (i.e. to high fluoprescence stationary state) then you set it going back to exponential phase in the morning (i.e. diluted into fresh media), after which the cells which previously had time to achieve massively high fluorescence/cell are now having this diluted out.

I would say in general by far the best results come from keeping OD constant (i.e. in turbidostat mode) then looking at changes in fluorescence.

You could adjust the software to show you non-normalised data if you like. Have you taken a look at the section that discusses normalisation in the Supplementary of the paper?

One other thing – if you use LB you will get generally a TERRIBLE signal (i.e. in terms of signal-noise ratio) due to the fact that it is very auto-fluorescent and generally also has large variability batch-to-batch and within a batch over time. Potentially unless you have VERY fluorescent cells this kind of chaos will completely obscure the actual fluorescence changes youw ant to measure. I would strongly recommend using M9 media at least…

[harrison]

That error can happen if there is a bad connection somewhere in the electronics – or if the system overflows any liquid which lands on the moisture sensing tracks that causes it to crash (deliberately in that case). Note that it could even be that water condenses on the test tube exterior and then ends up on the moisture sensing (silver lines on top surface) and triggers it.
What version of reactors are you using? I assume as above you have the latest version of the Beaglebone software?

[harrison]

Hello,

When the experiment runs it continuously stores your data in a .csv file on the device. You can copy this off the device and open it (directly in excel) using the instructions in the manual. You can then plot it in excel like you would any other data, or read it into some other software program to plot it.

[harrison]

Yes, it does look like it overflowed, likely due to either it being set up in such a way that the in/out media jar liquid levels are equal/above that in the reactor, or, if the output pump was not working effectively (i.e. making a poor seal on its tube).
It seems there is residue on other parts of the heat plate as well – so probably there was more in there than just a drip.
The fact that the liquid is on the chip in the second picture may indicate why the LED is malfunctioning – that chip is responsible for turning it off/on!

I’d recommend cleaning it all very carefully with alcohol, letting it dry, and seeing if that fixes it. Or, you might be better contacting the manufactuer (Labmaker) and asking if there is anything they can do to help.

[harrison]

To be honest I have never used a mac myself and have no idea what rare/unexpected errors might occur. In our lab we have reactors on both windows and linux PCs without such issues.

[harrison]

Hello,

It seems there might be a few different things at play here.

For the Broken Pipe and winch errors – these are errors (If I recall correctly) arising from the actual connection between the Beaglebone Black and your computer controlling it (i.e. PuTTy is unhappy). For example, your computer might be set to de-power or shut down USB devices/connections after some period of inactivity. Hence it is causing issues (which would explain why your colleague is able to run things properly from the python but not the GUI). For this case I would strongly recommend trying another computer, or trying to figure out what it is that yours is unhappy with.

The above one would expect to be similar regardless of any hardware fault (i.e. same error would appear for every reactor).

The second thing you mention (those errors + M1 not working) is indicative of a hardware fault in that particular reactor, potentially due to liquid ingress, or bad hardware assembly in the first place. I would recommend taking the sides off that reactor and looking at the circuit to see if there is any obvious damage. If it is an assembly fault Labmaker might be able to replace it for you

Harrison

[harrison]

Hello,
Yes – I believe I talked to your safety officer via email. It seems most likely there is some hardware fault, either due to a bad connection (i.e. during assembly soldering was not perfect), or if any water/droplets got into the reactor when your team was using it this could similarly cause some such issue.

[harrison]

Hi Nishant,

I am afraid I do not know. I have not tried this before, all mine run of windows or Linux. I know some people have it working on older Macs but I am not sure about M1.
It seems like it is a Beaglebone-specific question, I found this forum thread that might be interesting: https://forum.beagleboard.org/t/is-my-beaglebone-black-busted/29513
But I could not see much more than that online. I suppose it is a fairly old computer (Beaglebone) interfacing with a rather new one (M1) so there may not be a wealth of people trying to do the same thing…

[harrison]

I assume you mean the exterior? The most likely thing to generate a lot of heat would be if the LEDs are on for long periods at high power – is this the case in yours?
Is the reactor working fine function wise?
If it is IS working as expeted the exterior should be at approximately ambient temperature (provided you haven’t set the LEDs to continuously output high power)…

Harrison

[harrison]

Hello,
The minimal volume is ~15ml. If you take the sides off the device you can see why this is – the liquid needs to be high enough to be seen by the sensor/laser/LED.

Harrison

[harrison]

The moisture sensor/s are the parallel silver tracks around the main hole in the device where the test tueb goes, and also around the outside of that top face. They are silver lines approximately 1mm thick and 1mm gap between them. It senses moisture by looking at the conducitivty between those tracks – so if a droplet spans the two it will trigger the sensor.

For the csv, could email it to me at harrison [at] chi.bio

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