Re: [DIYbio] Re: Cheap capillary electrophoresis

Did I understood correctly? You want to use florescence as a detection instead of typical absorption and you can do that with labeled DNA. It might work in that case. Like I said UV is a problem since you need special optics components. If you just need UV excitation you just need to make sure that the microscope slide that the capillary is fixed on is UV transparent (and maybe remove the capillary coating). I like the idea of using microscope optics but florescence will be weak (small sample amount and florescence is weak in general) but adding all the pixels will help. One problem that could arise is that DNA ladder needs to have polymer passed with them. These would also absorb excitation wavelength and considering higher mass/molarity they would probably reduce your signal a lot. Another thing is heating, florescence would increase heat and that is not good for capillary stuff. I am curious how it will turn out.

On Friday, May 5, 2017 at 10:34:19 AM UTC-5, Harristotle wrote:

Hi Gordana,
what I had in mind for optical detection was a simple USB microscope focused onto the capillary. You could then take a video of it (amcap), and then pick the video up with ImageJ and plot color vs time/frame number.

My ImageJ skills are weak - I knew it as NIHImage, and I never had my own Mac to run it on, so I have not yet figured how to do this. I know from the examples that it is possible to step through the video (avi) file and treat the images sequentially. I am short of time, so only playing with this a little.

Possible cheap detectors for this:

1) a uv source shining through the capillary onto yellow highlighter drawn on paper behind - monitor diminishing of the fluorescence when an aromatic floats past.

2) some variation of sybase green/ DNA ladder


I am quite impressed with the magnification/focusing possibilities of those usb cameras. It is not near spectrophotometer level, but it is very cheap, and looks quite robust. When I get it all going, I'll look at linearity.

My current update is that capillaries with silicone oil run well with pillbox red and 50mM NaHCO3 as running buffer (pH 7.8, supposedly, but not measured).

In answer to your other question, both ends of the capillary were in fluid.

Cheers,
H.



On Wednesday, May 3, 2017 at 10:00:38 PM UTC+8, John Griessen wrote:

On 05/03/2017 08:52 AM, Gordana Ostojic wrote:
> Here is one

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Hello Gordana,
You said about harristotle1's video:  https://www.youtube.com/watch?v=jSwYqz_R9n8&feature=youtu.be

 > I can't see from the video the other end of the tube and if that is in liquid. Anyway I did use similar thing (it's somewhere on
 > this board). It turned out that optical detection is hard (material issues with UV, lamps are expensive, power is demanding...) so
 > I found out most people work with dielectric detection.

Are you sure optical detection is still "hard" today?  How did you do the dielectric detection?  Measure capacitance of the
capillary circuit?