I have come across this problem too. A program called Minianalysis is very good for analyzing this type of data and I found it very useful. It is freely downloadable here.

http://www.synaptosoft.com/

good luck! =)

My experience with minis is that it is not only going to depend on the age/treatment, it is also going to vary significantly from cell to cell. I used MiniAnalysis (synaptosoft), pretty cheap and works fine. Automatic detection, unless you have really low noise is not great. I usually set my treshold at 5pA. If most of your minis are from 10 to 30 those should be easy to see. Play around with the block size to see better the minis.

Regarding numbers, I would recommend counting 5 minutes or 500 events, whatever comes first.

Hi Cherry

Mini Analysis Program from Synaptosoft Inc. is good!

http://www.synaptosoft.com/

It allows one to play around with the detection parameters (rise time, decay time, amplitude threshold and so on). It also has a noise analysis function which helps one set the thresh hold for signal detection). If noise is low, the auto detect option works fine. However, in cases where noise is high I would depend on manual selection! Its tedious that way, but preferred, I don't have to deal with false hits later. I find that manual selection in Mini Analysis is far easier than in Clampfit . However, only the demo version is free but it allows you to do most of things the paid version does (250 $ or so I guess for the paid version). I feel you can check it out. I hope you find this information useful :)

You should try minianalysis. I have had good luck with it. http://www.synaptosoft.com/MiniAnalysis/

There is a free trial version.

HI, Put on gap-free mode when you edit the protocol....

I wonder whether doing a moving average might help get rid of some of the noise - haven't thought this through to see what other issues you might run into - but worth a go to see what happens. happy to give it a go if you want.

Hi Cherry, I used a similar strategy for detecting miniature currents in mouse brain cells. From memory I would make a template from a typical miniature for each cell, and then have a program to slide it along and detect miniatures by scaling it in amplitude. It worked pretty well and like yours you could vary the allowable error threshold up or down so that it would match what you detected by eye. It was done in Igor Pro which is an analysis program with some programming tools in it, so the program was easy to mess with. I think very noisy recordings will always be a problem. If you like I could dig out the detail of the methods we used, or I could contact my (then) supervisors and ask them for the program, they would probably be happy to supply it. Full copies of Igor are a bit expensive but you can get a student license for I think $80 or so if you are a student (or know someone who is :). Cheers, Andy

Might be worth trying a moving average and see what happens? havent thought it through but might be worth giving it a go....

It would be good go back to the classic electrophysiology and try to minimize the RMS. There is no magic with a randomic event... How can you use a template?

We've been able to use Clampfit for such analysis. In fact, we are doing almost exactly the same but use a higher event detection threshold (4 to 5) and always (!) do it manually. For most types of analysis it is enough to accumulate 200-400 events. Therefore, the selection can be done within 10-20 min. Alternatively, you can run it automatically and then reject the ''bad'' ones but it is more time consuming.

I used minianalysis, igo pro, and clampfit to analyze minis. I found that clampfit is the best. It is fast and easy. According to you problems, I suggest that you may need to make several templates. I make a template by averaging more than 20 events with the similar waveform. After you set templates from a trace, you can detect minis with multiple templates and exclude overlapping.

By the way, there are glutamatergic, gabaergic and glycinergic postsynaptic currents in the brain stem, and they have different kinetics. You may need to add antagonists for others if you are interested in one of them. If you use low chloride intrapipette solution, such as, K gluconate (135 mM in J Neurosci. 29:12428-39), all of inward currents are glutamatergic, outward currents are either GABAergic or Glycinergic.

Any analytical tools cannot overcome the difficulties that noise brings to the signals. In my recordings, after 1 kHz filter, the P-P levels of noise is less than 2 pA, I can detect almost 100% events with a single template (at the threshold level of 4) in my recordings from the basal ganglia and cortical neurons. Therefore, you may need to improve the noise level in your rig.

Hi Mao

can you let me know how you have the recorded in the gap free mode. I mean I wants to record in these miniature curents from DRG neurons in p-clamp 9. how to make the protocol in pulse and other parameter. if possible let me know at my e-mail [email protected]

Thanks

Hi Jitendra,

It is really simple indeed. I just opened up a gap free protocol, set the time you want to record for and set the holding potential. If there is spontaneous currents at that holding potential it will be recorded.

Thanks mao, I will try

Hi everybody!!

I know long time is passed from the last answer, but.. Is someone still present?

I have the same problem as Mao (by the way: did you solve it in the meantime?), but for mEPSCs I found a single template that I have used for the entire data analysis, I hope it's not too bad... Empirically it seems to work fine.

For mIPSCs instead I have more troubles, 'cause my amplitudes are lower and the noise is around 10 pA, and detect the event is very difficult. Moreover, when detecting events, Clampfit founds often many events within some of them, thus calculating for example even 5-6 events instead of one! Basically it detects more events within the same event, overlapping them. Shall I reject the undesired events? I think it is an incredible work, since I have thousands of events too...

About minianalysis: I tried it, but I cannot find a way to use this software, there are no instructions and it seems quite difficult!!

Sorry for hijacking this old question, but I have an current issue that fits well to this discussion:

It is interesting, that after all this time, MiniAnalysis by Synaptosoft is still popular, although, according to my experience, support has ceased long time ago. I also used this software until recently, but now I guess I have to switch, because the new ABF2.x files are not at all supported any more, and Molecular Devices is not offering a batch conversion tool to "downgrade" the files (reply from their support). A question to the Synaptosoft technical support concerning this did not produce any answer yet. And I have my doubts that there will be neither an answer nor an update, because there where no updates since a long time and content is not updated either. I also would hesitate spending money onto this company, because I am not sure whether it is actually still running?

So I am considering doing the analysis in Clampfit, too, using the hints provided here, thanks. Although I know that high troughput and (semi) automated analysis is bit cumbersome there (even the guys from Molecular Devices admitted during a conference, that they can't beat Synaptosofts software).

But my question is, why is there no competitive, fresh new software like MiniAnalysis? Is everyone using mostly ClampFit or are people using custom scripts (MatLab, for example)?

Hi Simon

Did you find a solution? I have the same problem as you, I think MiniAnalysis is obsolete now and I was thinking about a new program to analyse my abf2 files... Clampfit is not bad but is there really no other program to do that at this time????

Hi Rachel,

I tried Axograph but it does not seem to do much better than Clampfit, so I am still stuck with the latter one. Anyhow I do not have Minis to analyze right now.

We had a script to do this written in Igor a few years ago, I could see if it is still around if you want :)

Dear all, 

I have been using Clampfit template search to detect spontaneous EPSCs of NMDA (clamped at 40mV) and AMPA (-70mV). 

For NMDA upward spikes, occasionally there are bursts of connected spikes and the Clampfit doesn't pick up each spike. The template was selected from the ones that are commonly appear and the threshold was set to 4. Is there a method to pick up all the spikes within the burst?

I also faced the problem when the detection was in autorun, it detected a few events within one event. The EVENT STATISTICS - the COUNT column should always have the same number of counts for all event properties?

Thank you.

I've also run into problems with analysis using Clampfit.

You can also use a program called Axograph. 

i'm pretty sure you can get a free version and it's one of the best electrophysiology analysis programs. You can detect events based upon threshold/kinetics/templates etc. You should have no problem detecting your events with it. (even ones within a burst)

Can I ask also about the quality of your data? You mentioned that your data contains some noise/interference (which is about 50% of your lowest signal). Can you try to optimize the setup to decrease this which will make your detection much smoother.

Simon 

you can also try wdetecta, on my web site.

https://hlab.stanford.edu/public/wdetecta.exe,

and if you don't have clampex 10.6 installed, you will also need the relevant library file:

https://hlab.stanford.edu/public/ABFFIO.dll

John

Having a stable baseline is critical. Do keep exclusion criteria for unstable traces as constant baseline fluctuation either because of flow issues or 60 Hz noise may lead to burying of events in the baseline leading to underestimation of frequency.  

Which 1KHz low pass filter are you using? I prefer 8 pole filter over Gaussian filter and recommend using Bessel.1000 events in how much duration? I recommend keeping the threshold same otherwise your comparison will not be fully reliable. 

Also, 10pA events are tough to pick with the noise level that you have mentioned. You can rely on the sharpness of rise phase to distinguish from baseline fluctuation. 

I realized that the links I provided above do not offer much in terms of usage information.  Here is a page that provides a bit more information. I hope you find it useful.

https://hlab.stanford.edu/wdetecta.php

John

Try this soft from Maria Bykhovskaia https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2290970/

this soft good detect mini

Maybe also check out Christoph Schmidt-Hieber's Python-based (i.e. free) Stimfit:

http://www.stimfit.org/doc/sphinx/manual/event_extraction.html

http://www.stimfit.org/doc/sphinx/manual/event_extraction.html

miniAna: an Igor Pro GUI might be a solution.

https://github.com/yuichi-takeuchi/miniAna

Hi guys,

Was searching for a solution to the now unsupported MiniAnalysis software and came across this trend. I tried contacting Synaptosoft but was unsuccessful.

Thanks for the very current input.. from John, Tobias, Yuichi and Dmitri.

I had issues using Clampfit. Will give suggestions a go ...

Sindy

Quantal analysis, Mini analysis are both quite useful to analyze minis.

Thanks Zhitao,

Unable to obtain Minianalysis nowadays.. But will check out Quantal analysis.... We are currently using clampfit but would prefer minianalysis