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[Jebus Jones]

OK be gentle with me. I'm brand new. I live in I am a mature student (aged 44) studying for a degree in cellular and molecular biology in Newcastle upon Tyne in the UK. I have been interested in science and technology since I was a very young boy, but for various reasons (family/work) etc. never really got around to applying myself to a formal pursuit of science until quite late in my life.

Anyway it seems to me if/when I complete my degree that the opportunities for middle aged newly qualified biologists is likely to be rather limited, particularly if I am interested in any sense in being at the bleeding edge of research. I have long read about the Open Biology movement, and to my eyes in a great many ways this is the bleeding edge. An attempt to bring biology and make it accessible to the masses is a deeply noble aspiration I feel. College professors spend too long isolated in their comfortable crystal towers. However due to having been intensely busy for this last 2 years I have not been able to delve into the topic in significant depth.

My initial response in finding this forum (and any other resources that users here might care to recommend?) is one of both astonishment and enthusiasm. I feel it is astonishing how far this community and this movement have developed and can't imagine why I have waited so long to explore it.

However, I have hit something of a small snag. I enjoyed hugely reading about the DIY projects available on the DIYbio web site. I initially set out and found this list because I had been vaguely contemplating for some time building my own PCR thermal cycler. However lacking sufficient knowledge and experience to do so, I concluded that it was likely someone had probably already beaten me to it - and that they might have plans. Unfortunately while this has proved to be the case, all of the DIY equipment projects I have come across so far seem to require access to a laser cutter and/or a 3D printer.

This seems to rather go against the spirit of 'democratizing biology', as very few ordinary people are likely to have access to these resources. Indeed as I live in a small city centre apartment close to my University, these resources are likely to be forever beyond my grasp. We do have a Maker Space in Newcastle and they do have a laser cutter and a number of 3D printers. But the laser cutter is very jealously guarded by those few members who paid for and bought it - and the 3D printers are in general all the property of individual members. The attitude appears to be 'if you want to use a 3D printer, you should probably make one.' There may be some merit in this argument, but really what I want to do is learn about biology, so this seems like a bit of a side issue for now.

I am also almost certain I must be the only person in this entire city who is interested in, or even remotely aware of these topics. I would love to be proved wrong however. But my question is, how do I get my hands on the relevant parts for any of these DIYbio projects without needing a laser cutter and a 3D printer?

I know some folks here might ask why don't I just use the lab equipment at my university? Unfortunately as an undergraduate I am not presently granted the luxury of just messing around in a lab and doing what I want - and in any case, the point in building stuff from the ground up is to better understand how they work, so I can build on that experience in the future.

Anyway sorry for the slightly long intro. As a side note I am also desperate for any spare lab equipment I can get my hands on, as I am archetypal penniless starving student and the prospect of being able to find the money to set up my own small domestic lab seems daunting. So if anyone has anything going spare they can share or give me please feel free to get in touch privately. In any case I hope I can play a very small part in this vast and exciting revolution.

[Brian Degger]

Come down to makerspace.org.uk
Your friendly local newcastle hackerspace.

I am interested in starting a local diybio/bioart group, and already do quite a bit of the stuff you are interested in. Plus we have laser cutter and 3d printers ;).

Be great to meet.

Cheers
Brian

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[Jebus Jones]

We already have met Brian. I'm the slightly pushy Scots dude you met at Maker Space a couple of months ago.

Nothing really appealed much at the time, but I would love to go 'in with you' as it were on a PCR build if you're interested? http://openpcr.org/

I am already on the trail of a Sanyo Centrifuge on eBay, but if you know any way we could start to put a small local lab together, or are interested in getting a local DiyBio Hacker group off the ground in Newcastle, I would be well up for helping out.

I really think Newcastle needs to get in on this act. We are too far behind the rest of the world in this respect I feel.

[Josiah Zayner]

I just started up a company that sells chemicals and equipment(soon) to DIYBio folks for fairly cheap (http://www.the-odin.com/)
I don't know how cheap when including shipping costs from US to UK but I don't think they are too bad?

As for PCR machines, I suggest you troll eBay. I have found many PCR machines that are functional in the $100-200 USD price range which even with shipping to the UK would be drastically less than the $600 USD OpenPCR. In fact if you are looking for a PCR machine in the $100 USD range (maybe closer to $200 including shipping) I can work with you to find something. Or even a centrifuge. Shoot me an email and I can give you an estimate on what it would cost to get the equipment and run the experiments you want.

Though PCR is not very usable unless you plan on running gel electrophoresis or direct sequencing afterwards so make sure you take these things into account.

I hate to see people use up their hard earned money on Bio equipment that can be found cheaper.


Thanks,
     Josiah
     josiah...@gmail.com

[Jebus Jones]

I was kind of up for building a PCR machine (AKA the maker mentality) just so I could learn some of the underlying technology and principles. But you are right, the cost is rather prohibitive, which is why I was looking to maybe share the cost with Dr Brian, or anyone else that might be interested?

Having said that, beyond building my own kit, I would be interested in any deals on cheap lab equipment (including PCR, centrifuge and anything else for that matter). As far as I understand it, the main hurdle will be import duty/VAT at 20% in the UK. For reasons I don't understand, depending on the shipping company, they might charge another £15 dollars per shipment for handling too for all items shipped from the USA. If things could be shipped from Europe, this would not be an issue however.

[John Griessen]

On 06/02/2014 01:43 PM, Jebus Jones wrote:
> I was kind of up for building a PCR machine (AKA the maker mentality) just so I could learn some of the underlying technology and
> principles. But you are right, the cost is rather prohibitive, which is why I was looking to maybe share the cost with Dr Brian,
> or anyone else that might be interested?

Would you be interested in using a micro python controller for the PCR?
If so, we could maybe collaborate depending on timing.

[Josiah Zayner]

I would suggest looking on eBay in the EU or UK you might find stuff. Feel free to email me for opinions on what you are looking at. Though I looked up the the VAT and duty taxes and they are only about 30-40 pound sterling.

The Maker Mentality is awesome! But in my opinion it would be super cool if you built something using DNA rather than overly expensive PCR machine.
If you still want to build a PCR machine I would suggest (http://www.instructables.com/id/Arduino-PCR-thermal-cycler-for-under-85/)
I have built that one and it is really easy to do!

[Jebus Jones]

That depends where you're based? I would still welcome Dr Brian's input? As I said I met him and he's a really cool guy. We spoke briefly about Open Biology, but never got as far as trying to set up a group.

[Jebus Jones]

Yeah I looked at that a couple of days ago. But (as I recall) it was milling out that heatsink from a solid block of aluminium (or aluminum since you're American) that proved the stumbling block. I just don't have the equipment lying around to do this.

I had a look at the price of some Micro Pipets and I nearly swallowed my tongue lol. We throw these things around in our college lab every day and every day one or two get broken by careless use. I had no idea they were so expensive.

I guess the reason why science is exclusive is because its so expensive. This is why I think it's vital that folks club together and try to share the cost.

[John Griessen]

On 06/02/2014 04:38 PM, Jebus Jones wrote:
> But (as I recall) it was milling out that heatsink from a solid block of aluminium (or aluminum since you're American) that proved
> the stumbling block. I just don't have the equipment lying around to do this.

Heat flow can be equilibrated quickly bystirring vigorously. Big hunks of metal are not needed.
vigorous stirring of fluid, (which can be air), around your test vials is just as good
as fancy machined metal for transferring heat. Controlling heat is easy for microcontrollers,
just throwing switches, (and needing only cascades of heavier and heavier switches to get up
to high powers of some heaters).

[Jebus Jones]

Dammit I already replied to this and my post got deleted? Sigh, lol. I would really like to build the $80 PCR, but I would also like a decent heatsink for it too. I think a heatsink would be useful, as it would help get things to temperature efficiently, and I don't have to be there to baby sit the reaction. I don't want to play games. I want (preferably) a viable PCR option. $80 sounds good. This sounds like democracy in action to me. Although of course the practical limitation is that the vast majority of people are unlikely to have CNC machines lying around to carve out the kind of heatsink this design requires.

I don't suppose anyone here would be willing to do this for me and I would pay them for the cost of their materials and time etc?

[Jebus Jones]

On a side note Yay! I am now the proud owner of a 13,000 rpm Sanyo MSE Centaur Micro centrifuge. I wonder if this is likely to be sufficient for most small lab applications however? I have read about guys here building centrifuges from Dreamels that can hit 50,000 rpm?

[Jebus Jones]

"I don't think [heatsinks] are useful for heat transfer through plastic vials. Oh well .."

That's an interesting comment. However could not the same be said for the liquid medium in which the vials are contained? How useful are plastic vials in this context, when you want to get things to temperature quickly in as few intermediate steps as possible? Why isn't glass used instead?

Also if this is the case I presume it is the volume of the liquid medium that is likely to have the greatest cooling and heating effect on the sample and that a large volume of liquid therefore would have the greatest effect in this heating and cooling process? Are rapid changes in temperature beneficial, or are more gradual changes preferable?

Also if getting to each temperature quickly and efficiently is the main goal, then presumably the purpose of a heatsink should be to have the capacity to cool the liquid medium as efficiently as possible? There are many ways this could be achieved, as those familiar with working with CPU cooling methodologies might be aware. (Like a water cooling via a radiator and a pump etc.)

Lastly in PCR (in my limited experience) I have tended to find that water is used as the main thermal medium. However I am aware that there are other potentially more efficient thermal mediums that can endure much more rapid heating and cooling cycles, should this be preferable?

On Monday, 2 June 2014 23:06:51 UTC+1, John Griessen wrote:

[John Griessen]

On 06/03/2014 06:15 AM, Jebus Jones wrote:
> "I don't think [heatsinks] are useful for heat transfer through plastic vials. Oh well .."
>
> That's an interesting comment.

The context is: "When you want to cool down, what?" -- You have a big heat sink heated up then.

However could not the same be said for the liquid medium in which the vials are contained?

If the fluid, (usually air is most convenient), is vigorously stirred, you get rapid heat flow.

How
> useful are plastic vials in this context, when you want to get things to temperature quickly
in as few intermediate steps as
> possible? Why isn't glass used instead?

Plastic is inexpensive, disposable, reusable, some kinds don't absorb water, inert for water chemistry...
Glass could be good also, if you can get it. Is the closure going to be plastic? Why not all plastic then?

Vials of plastic are what I am going to be experimenting with for heated/ambient
air thermal cycling.

> Also if this is the case I presume it is the volume of the liquid medium that is likely to have
the greatest cooling and heating
> effect on the sample and that a large volume of liquid therefore would have the greatest effect in this heating and cooling
> process?

I don't see that. Rapid flow of controlled temperature air is going to transfer heat quickly.
Having no thermal mass besides the vials and a low heat capacity holder for them lets you change to cool
down very quickly. Just open a door for ambient air to be fan forced around the vials,
close a door to where the heater element is, and your temperature differential will be greatest
and about equal in both decreasing and increasing directions. Controlling hot air temp with an
electric heater is easy. Controlling slightly above ambient air temp with an
electric heater is easy.

Are rapid changes in temperature beneficial, or are more gradual changes preferable?

Rapid allows more cycles quickly, benefiting a busy user, and I think there is a benefit in
basic PCR reaction also.

[Jebus Jones]

In that case, if air can be used as a medium for rapid heating and cooling cycling, then would not some form of heatblock arrangement (like a CPU heatsink) on a CPU be used (with a suitable gap to contain a small volume of air) with a heat/radiator source (again like a CPU) with a fan arrangement blowing over the heatsink in preprogrammed cycles to produce this rapid cycling effect?

To make this clearer, think of something analogous to (again) a CPU (the heat source) on a motherboard, upon which the plastic vials could sit, which could then heat the contents of the vials rapidly to a specific temperature. Then when the vials had reached that temperature for a given set period of time and a new lower temperature had to be attained, the fans on the associated heatsink arrangement could kick in to rapidly bring the vials down to the desired temperature, while simultaneously the 'chip' could be switched off until it achieved the new desired temperature. When the chip and the contents of the vials had equilibrated at the new desired temperature, the chip (or heat source) could then be switched back on, so that this new temperature could be maintained.

Of course a CPU type chip might not be the ideal heat source as it would give a very uneven heat distribution (heating from the bottom of the vials up first), so perhaps having a more evenly distributed heating source, (like a small coiled heating element) in the centre of the heating block (or heatsink) inside which the vials might sit would be a better arrangement?

If the idea is to encourage rapid temperature cycling, would this not be the most effective way to achieve this goal?

The issue with the current methodology, is as you say while its effective, it is terribly slow.

[John Griessen]

On 06/03/2014 05:41 PM, Jebus Jones wrote:
> would not some form of heatblock arrangement (like a CPU heatsink) on a CPU be used (with a suitable gap to contain a small volume
> of air) with a heat/radiator source (again like a CPU) with a fan arrangement blowing over the heatsink in preprogrammed cycles to
> produce this rapid cycling effect?

The air gap you describe would not be moving, thus it would be a high barrier to heat flow -- an insulator.

[Jebus Jones]

It depends how big the air gap is. There would of course be an optimum. I haven't seen how this $80 PCR works yet. But it strikes me that if it's just the plastic tubes in direct contact with the heatsink then that that really is a dumb idea.

Heatsinks would be pretty bad at directly removing heat from plastic tubes in this way. However they are very good at removing heat from air, or water or other fluids.

What I described is essentially a CPU heatsink - which of course do work by allowing sufficient airflow around the heatsink. In general these work very well and offer quite accurate levels of temperature control. So why not sit a small water bath (or air bath - whichever you prefer and whatever tests prove will work more efficiently) in the middle of a heatsink carved from a similar piece of aluminium to the above $80 design and have fans blowing down (or across) the heatsink to remove the heat? I don't think it matters so much if the water (or air) inside the bath itself is moving or not, just so long as there is enough of it (although still a relatively small volume) to allow rapid heat transfer to the heatsink?

Inside the water bath sitting in the middle of the heatsink, you would of course need a small (possibly coiled) heating element to heat up the water (or air) when required. You would also need a temperature sensor and a means of controlling the temperature at precise intervals. The only point of having a relatively small volume of water (or air) is to be able to cool and heat this as rapidly as possible.

I am not describing anything revolutionary. This is as I said pretty much how CPU heatsinks and fans work, as I'm sure you know. I assumed this was how the $80 PCR worked anyway?

[Nathan McCorkle]

I've thought for a while it would be a great experiment to try flushing hot or cold water from separate reservoirs past tubes, that way you just get rid of the heatsink rather than cycle it.

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[John Griessen]

On 06/04/2014 01:20 PM, Jebus Jones wrote:
> It depends how big the air gap is. There would of course be an optimum.

Just remove the air gap.

Removing the air gap by having air be the cooling fluid is the optimum
for low costs and high performance. Thin walled vials would be compatible with
air cooling/heating. Any kind of thick vials, or vials dropped in holes
in blocks of metal, (as was done in PCR machines of 1995), is going to cost more
and probably be lower performance.

[John Griessen]

On 06/04/2014 01:58 PM, Nathan McCorkle wrote:
> I've thought for a while it would be a great experiment to try flushing hot or cold water from separate reservoirs past tubes,
> that way you just get rid of the heatsink rather than cycle it.

Yep. Open and close doors to get rapid cool and heat up rates that are about equal
in dT/dt. I like air as the coolant.

[Winand Slingenbergh]

I met someone recently who built one of these out of an old drip coffee maker heating element. This yielded some pretty great ramping times. Unfortunately this was many 20+ years ago so the device no longer exists. I do believe it would be a great way to do it cheaply. Peltiers can only take so many cycles and I am not too sure about the reliability (read temperature stability) of the lightbulb approach. Something with a heated lid would of course be preferred.

I am also trying to collate all the homebrew PCRs out there on this page:

http://www.diybiogroningen.org/mediawiki-1.22.0/index.php?title=OpenerPCR

If you know of any that are not on that list but should be please let me know.

To view this discussion on the web visit https://groups.google.com/d/msgid/diybio/CA%2B82U9LAPmme-PXEz_o1O_tZ9C-M%3DGdE4AOhbFZ_G32Vr_MGRQ%40mail.gmail.com.

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Regards,

Winand Slingenbergh

[Jebus Jones]

Bare in mind I am a total novice, but I am interested in how one might maintain an even temperature gradient through (say for example) a capillary type system?

I thought initially of a 'turntable' (or 'stage') type arrangement (like an old turntable from a record player)  around which a capillary type tube could be wound. This tube could be filled with the PCR target under consideration. Three (or perhaps more?) distinct temperature zones could be maintained by adding three heating elements and/or heatsinks (perhaps coiled around the capillary?) set at different temperatures at different zones around the outer edge of the disk. I thought to make the disk 'wobble' about it's axis slightly to both facilitate and enhance fluid flow through the tube?  A small air gap could be left in the tube to allow the fluid to flow freely. One could in effect set up a 'production line' type arrangement where each segment in this apparatus would be at a different stage in the PCR process.

This of course is just a crude thought experiment. There has already been some extremely advanced work done in microfluidic and other systems that employ a temperature gradient type arrangement to a similar effect.

http://www.abebooks.com/Continuous-flow-temperature-gradient-microfluidics-Spatial-polymerase/10797610401/bd (I wish I could read this full article!)

http://www.newscientist.com/article/dn11763.

The problem however (although it's likely significant progress has been made in recent years) is how to prevent 'mixing' between the different temperature zones?

Nonetheless one of the above articles appears to indicate that mixing between different temperature zones might be less problematical than one might think? I would certainly like to think more about taking a pop at building something along these lines, providing a stable temperature gradient cold be maintained.

On Sunday, 1 June 2014 09:16:06 UTC+1, Jebus Jones wrote:

[Nathan McCorkle]

Water is not the most common heat sink for PCR these days, I think...
metal probably is, as in heat sink blocks. You don't need a small air
volume near the heat exchanger, what John is describing is basically a
box you've pointed a hair blow drier into... you get even temperatures
throughout the samples in the box by stirring or swirling the air
(actively or through some vortex/twisting design of the chamber) or
rotating the samples in the air (temperature controlled centrifuge,
then, basically).

You could do this many ways... you could have a carousel (like at an
amusement park) where some rotating arm with a wheel at the end rests
on a circumferential, which would be shaped like a wave with the
height of the track going up and down. When the arm rotates, it
'drives' on the outside track, since the 'arm' has a hinge, when the
track goes up or down, the arm 'drives' up or down to follow. Now
imagine just past this circumference are pools of hot or cold water,
every hill on the track would lift a basket at the end of the arm out
of the water, then when the arm is coming down the next hill on the
track, it would be lowering into the next water bath of a different
temperature.

See this reference to a childhood amusement park I frequented:
http://nicolecoomer.com/jour232/wp-content/gallery/kennywood/kangaroo.jpg

Note that the hinge in this picture is associated with a
pneumatic/hydraulic arm... that isn't to lift up the basket (full of
people here) but rather to slowly lower them down after the car has
been driven up a hill (simulating a gliding sensation).
http://www.themeparkcritic.com/Uploads/2108/Kennywood%20074%20copy.jpg



for capillaries, some have found just constant cooling from below is
fine, then heating with laser pulses.... others have had different hot
and cold zones, then snaked a capillary back and forth between the
zones. Pumping the fluid at a constant rate allows the cycle time to
remain constant (as long as the zig-zag of the capillary is the same
over the entire stretch).

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-Nathan