$24,000 DNA Microarray Scanner Teardown

[dexters_lab]

taking a look at a $24,000 DNA Microarray Scanner

lasers, photomultipliers and expensive engineering 

[orion242]

So how much did this guy set you back?

Looks like some nice bits and bobs and a crap load of AL for the scrappers.

[dexters_lab]

So how much did this guy set you back?

Looks like some nice bits and bobs and a crap load of AL for the scrappers.

it cost me £30 which is quite a large depreciation!

yes there are a few nice bits in there, quite happy with the haul!

[PA0PBZ]

Thank you for (another) very interesting teardown! I appreciate that you took the time to take it apart and explain all the bits and pieces. Another great dexters lab production 

[dexters_lab]

Thank you for (another) very interesting teardown! I appreciate that you took the time to take it apart and explain all the bits and pieces. Another great dexters lab production 

thanks!

i do struggle to explain things sometimes... i know what i want to explain but sometimes i just end up talking jibberish, problem is i dont always realise until i have a pile of bits and i'm editing! No chance for a second take!

[orion242]

it cost me £30 which is quite a large depreciation!

Nice score and made a great teardown.

thanks!

[Synthetase]

I used to use these. So outclassed by modern high-throughput sequencing, but very good at the time as long as you had a well-designed microarray.

Also, fricken lasers! What's not to like?

[Synthetase]

Okay, just finished watching the video. Nice job.

I suspect those filters before the photomultipliers are not just to reduce cross-talk between channels. The fluorescence of the dyes used (red example: https://www.thermofisher.com/au/en/home/life-science/cell-analysis/fluorophores/cy5-dye.html ) are only a few nm different to those of the incident light, so those filters may also be cutting down on reflected incident light as well.

For me, the most interesting part was the disparity in the power of the red and green lasers. When I used to do experiments with DNA microarrays, we used to do multiple repeats to average out the data, but we'd also do a 'dye swap' - we'd swap which sample was attached to which fluorescent dye. This helped us to compensate for the less than equal intensity of the two different lasers and dyes. Of course the arrays themselves had lots of control spots, too. It was quite an involved process.

This technology is actually still in use. It's a waste of time for DNA samples because now we can just brute-force sequence everything, but it's still used for protein work.

[dexters_lab]

Okay, just finished watching the video. Nice job.

I suspect those filters before the photomultipliers are not just to reduce cross-talk between channels. The fluorescence of the dyes used (red example: https://www.thermofisher.com/au/en/home/life-science/cell-analysis/fluorophores/cy5-dye.html ) are only a few nm different to those of the incident light, so those filters may also be cutting down on reflected incident light as well.

For me, the most interesting part was the disparity in the power of the red and green lasers. When I used to do experiments with DNA microarrays, we used to do multiple repeats to average out the data, but we'd also do a 'dye swap' - we'd swap which sample was attached to which fluorescent dye. This helped us to compensate for the less than equal intensity of the two different lasers and dyes. Of course the arrays themselves had lots of control spots, too. It was quite an involved process.

This technology is actually still in use. It's a waste of time for DNA samples because now we can just brute-force sequence everything, but it's still used for protein work.

yes, i think mikeselectricstuff mentioned the point about the slight difference in laser & florescence frequency in the youtube comments

it's interesting that as a user you were doing a manual step to effectively balance the laser power between the red and green, wonder why they just didn't match them? - I guess they had their reasons!

[Synthetase]

Like I said it's not just for the laser, it compensates for the dyes as well. To be completely honest, I'm not sure whether it was entirely necessary - but if you're doing repeats anyway, just switching which dye is attached to which sample is simple so it's good practice.

[PedroDaGr8]

Synthetase is bringing up some good points. I have not been able to watch the video yet, but there are a variety of reasons you use these filters.

Time for me to start rambling on about stuff nobody cares about  This is VERY much my realm of expertise, being a bionanomaterials chemist with my recent focus having been on bioconjugation of fluorescent materials.

The first is that the stokes shift between the excitation and emission is usually around 10nm difference. (So a dye that emits at 545, would have a peak absorbance around 535nm). This means that you need a solid filter set to prevent the laser light from actually entering the detector and obscuring any of the signal you are trying to detect. For microscopy, we often use filter cubes which have a variety of filters to handle the various wavelengths needed. Here is an example of a common filter cube for FITC The blue is the filter for the excitation wavelength, the green is the beam splitting filter (it reflects the excitation up into the sample, while letting the return fluorescence through) and the red is the emission filter.

Also, organic dyes have a phenomenon which is called a red tail. What this means is that their emisison profile is not symmetric. If you see the following image:


The solid black line is the emission spectrum, you can see how instead of returning to baseline it has this tailing off effect in the longer wavelenghts. This bleeds into any other channels that are to the red side of the dye. This is a VERY big issue in flow cytometry and microscopy, as well as many other polychromatic analysis methods. The following image shows the spill through:

You can see in particular how the green dye overlaps with the yellow dye, giving a significant signal represented in the yellow channel. This signal is represented by the green hatched triangle.

Compensation is damn important in flow cytometry, so much so, that my old place of work made LARGE amount of money selling compensation beads. Sub-um scale beads that helped aid in the compensation of the instrument. This shows before and after compensation of the instrument to show you how important compensation is. For these signals you want them entirely parallel with their axis, because any deviation from this is considered a positive signal.



As you can see, compensation can make a pretty big difference. Hopefully, my rambling helped explain the use of the filters in the instrument.