Integrated qPCR

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I am looking into options for integrated qPCR machines. We have a BioRad CFX384 that we could use, but we are trying to weigh the pros and cons of what is currently on the market. I have some experience with a QuantStudio 7 Pro and I’m looking into the qTower3 Auto. The most important things to me are robot access and the ability to automate the data analysis as long as the instrument itself performs well.

Ideally, I would like to be able to start the qPCR run, get the output from the instrument as Ct values, perform some minor calculations in a custom script, and generate a worklist for a liquid handler based off the quantification. Is anyone doing this kind of thing already? How does the analysis part work? Is there another instrument I should check out?

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Most folks probably use the QuantStudio 7 Pro. The file it generates is a zip file.

This thread is useful.

Are you using a scheduler or would this handoff be through a liquid handler?

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We would be running with a scheduler. I have played around with the EDS files that the QuantStudio generates before. Thanks for linking to that thread.

In a previous lab we used a QuantStudio as a walk-up instrument, so the biologists always interacted with the data. My understanding was that there was an analysis software that opened the EDS file, performed normalization and Ct calling, then exported as a CSV. Does the EDS file actually contain the analyzed data? If it is just am after of parsing the EDS, then I can handle that.

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Yes it does! Copy any .eds file and then change the ending to .zip., you’ll find that now you can extract a ton of information. Schedulers already have integration drivers for QS7 Pro so if you’re always running the same protocol, you can just create your own data parser.

However if you want to control every granular detail, say modify your QS7 Pro method on the fly… you’ll need to explore the above link.

There are also some data service/integration providers that effectively provide the Thermo Connect Cloud experience via their platform but that’s another level.

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Interesting! I’ll have to read up a bit more to see where to find the processed data in there. You’re right that there is a ton of info available.

Pro tip: if you are working with EDS files from a script (at least in Python), just running “unzip” on the EDS without changing the extension works.

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Hi @DCurrier ,

These are the devices that we’ve found are integration friendly: BioRad CFX, QuantStudio 7, qTower3 Auto, Roche LightCycler, Qiagen dPCR - QIAcuity Eight.

The Thermo QuantStudio 7 can only accept semi-skirted 96 well plates. It can accept fully skirted 384 well plates.

If you need a list of compatible plate types, go to :

qPCR and PCR Plastics Selection Tool | Thermo Fisher Scientific - US

When working with the 96 well semi-skirted plates, you will need this support base:

Applied Biosystems part # 4379590 Description - MicroAmp 96-Well Support Base

MicroAmp™ 96-Well Support Base

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Thanks @JRM! Good to know that the QuantStudio 7 is not compatible with fully-skirted 96-well plates. I think our main use-case would be 384 well anyway, so it should not be a big issue.

Do you have a favorite among those you have worked with? Are they all generally the same as far the core qPCR performance is concerned?

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I’m asking colleagues now. i’ll see what they have to say and let you know.

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The Qiagen QIAcuity dPCR is auto friendly?

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Hi @DCurrier, ok, so the feedback i received was, There is a weakness in the LightCyclers. For certain protocols, it outputs two different files for different components of the “reads”. User will have to manually combine the files together before uploading to their LIMS. because of this, we know people who have switched over to using QuantStudio 7 Pro that outputs a single file with all the relevant data. Hope this helps.

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Thanks for the feedback; this is really helpful.

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I’d be careful with qTower Software. They have a tendency to generate positive curves out of negative raw data.

Do you completely trust the native software for CT calling? In my experience the softwares tend to have issues there in edge cases.

For Quantstudio there are nice 2 component plates available form Azenta. Much better than soft single component plates in terms of plate sealing, data quality and automation friendlyness.

If you are also looking for a plate sealer: Don’t buy Azenta.

I’ll be reading here and already like the eds - zip-file info. Thanks for that.

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I just read a blog about a new flexible qPCR system.

Seems interesting and something to keep an eye out on.

Although it seems like a lot of qPCR work is starting to make it’s way to dPCR’s.

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@Florian Would you recommend using a different software for Ct calling, or just have manual confirmation of the software’s analysis?

Our lab currently likes using the BioRad hard shell plates, but I have successfully used TwinTec plates with automation before. You’re right that single component plates tend to be too flexible for reliable automation.

I plan to use PlateLoc sealers. I have never used the a4s, but I have never hear anything bad about it.

@luisvillaautomata If N6 can deliver on their promises, then that system will be incredible. Thanks for linking to it.

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In my experience BioRad sometimes has issues correctly setting the baseline, which in turn may lead to false results. Thus here in the lab we rely on human expertise.

Well, if you are using the PlateLoc, you definitely don’t want to try the Azenta alternative, even if the price point is much lower. We have adopted the PlateLoc a while back and it has been night and day.