Re: [DIYbio] Re: Making my own incubator inexpensively

But if I need a reference, and I only have one incubator, I'll still need to buy at least one good temp sensor. So I guess we should be talking about two designs, a reference or golden standard, and "slave" devices which get calibrated against that, as our lab needs grow

On Apr 24, 2012 4:20 AM, "Cathal Garvey" <cathalgarvey@gmail.com> wrote:

It's not the error in my own samples that matters, so much as the
variance between incubators. Also, it's not about the cells' survival;
it's about maintaining them at a known growth state.

We grow mesophiles like E.coli at 37C because, for most of them, that's
the ceiling for comfortable growth. Higher temperatures mean more
reactions per second, meaning faster growth. However, any higher than
this ceiling, and heat-shock starts to get induced. While the cells will
survive, their gene expression profiles will change markedly.

So, if you and I set our incubators to 37C, and mine hits 36.5 while
yours hits 37.5, that could have a significant effect on our experiments
with the same DNA constructs. If any protein folding effects are
interrupted by heat-shock, you'll have reduced apparent expression. If
we're using heat or chemically induced promoters, you'll seem to have
higher expression at the same temperature.

At the very least, you'll have faster growth than I will at the same
apparent temperature, even controlling for all other factors.

If this were unavoidable I wouldn't complain, but for the difference of
€0.5, it's not worth skimping on accuracy. Science is about good
experimental control and reproducible results. If our equipment has such
significant error, you and I can't really compare or reproduce one
anothers' work; that's bad science.

On 24/04/12 00:23, Simon Quellen Field wrote:
> This is an incubator, guys.
> What organism do you know that maintains its temperature to 0.1C?
> Why would you think that was important for anything you are going to
> incubate?
>
> Let's suppose that you need to incubate something for 10 hours (600
> minutes).
> Suppose you set one incubator for 310 kelvin, and another for 310.1.
> Thus one is to get 186,000 degree minutes, and the other gets 186,060 degree
> minutes.
>
> You open them both, and immediately the air temperature changes by 11
> degrees
> to 26, the temperature of the Petri dishes full of agar.
>
> It takes both incubators 20 minutes to settle down to their set temperature.
> You have an average of 5 degrees below the set point during those 20
> minutes.
> That is 100 degree minutes off already, which is more than the 60 degree
> minutes
> by which the two incubators have been set.
>
> But more to the point, suppose we are incubating E. coli.
> Let's look at some of the protocols in
> use<http://www.daff.gov.au/agriculture-food/food/regulation-safety/antimicrobial-resistance/antimicrobial_resistance_in_bacteria_of_animal_origin/appendix_2_bacterial_culturing_protocol_for_e._coli,_enteroccus_spp._and_cambylobacter_spp.>.
> One says to incubate at 37 Celsius for
> 18 to 24 hours.  Another of
> them<http://www.neb.com/nebecomm/products_intl/productC2988.asp>says
> to expect a 2-fold loss
> of transformation for
> every 15 minutes that a 1 hour incubation at 37 Celsius. A
> third<http://openwetware.org/wiki/Maxiprep_of_plasmid_DNA_from_E.coli_protocol>calls
> for incubating
> at 37 Celsius for 30 minutes, then incubating 'at room temperature' for 10
> minutes.
>
> It does not look to me like anyone is calling for one tenth of a degree of
> accuracy in
> temperature. But more to the point, given that how long the cultures are
> incubating
> is so variable, being off by a couple degrees does not seem like it is
> going to make
> any difference.
>
> If it *does* make a difference, is temperature the right thing to be
> measuring?
> If the reason we incubate at 37 for an hour is to get a certain density of
> critters
> per milliliter, why not incubate until the optical density is some
> particular value?
>
> Maybe we shouldn't bother with electronics at all.
> Run some hot water from the tap and adjust the temperature to 37.
> Get a 5 gallon insulated beer cooler and fill it with water from that tap.
> Put your Petri dish in a zip-lock bag, set it in the water, and close the
> cooler lid.
> Take it out an hour later. The water temp is still 37.
>
> But that takes all the fun out of building the device. So let's use the
> device to
> do *sous vide* cooking, or to incubate chicken eggs (which will hatch in 21
> days
> even if you are off by 10 degrees). Your *sous vide* pork chop will come
> out great
> even if you are off by 5 degrees.
>
> In other words, the temperature sensor in the microcontroller is already
> more
> sensitive and accurate than you need.
>
> -----
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>
>
>
>
> On Mon, Apr 23, 2012 at 2:20 PM, John Griessen <john@industromatic.com>wrote:
>
>> On 04/23/2012 03:28 PM, Cathal Garvey wrote:
>>
>>> Most other forms of temperature readout
>>> that I've encountered are a bit batch-variable, and I wouldn't be
>>> surprised if the same is true of something cool but hack-ey like your
>>> onboard diode idea
>>>
>>
>> It's the physics behind all the temp sensors, the differences are in the
>> implementation details.
>> Why would you pay more for a method or mechanism if you can't get any
>> better results
>> than with the low cost method?  You don't just always pay more, you shop
>> for a low accuracy need for low dollars
>> and hi accuracy for hi dollars.  Precision, or fine grained resolution, is
>> inexpensive,
>> and you can transfer the accuracy from another temp probe to your machine
>> and get repeatable results
>> without a temp standard built in.
>>
>> This particular MSP430, for example, MSP430G2230IDR, has a nicely
>> engineered diode temp sensor
>> inside, that can be switched in to one of its ADC channels to read the
>> temperature of the chip.
>> The diode will have a small batch to batch random variation, but will
>> always repeat very closely
>> as temperature differences happen.  There is a temperature compensated
>> volt reference for the
>> ADC so volts measured are truly accurate.  Temperature repeatability of
>> .01 deg C is probably
>> possible, although that's not accuracy, and would only agree that closely
>> on long equilibration
>> soak times after a temperature change.
>>
>> All you need is to put the chip in the stirred air you want to heat and
>> you're accurate
>> to a very reasonable precision, and you could get better accuracy by
>> calibration.
>> Repeatability and precision are easy to get with silicon microcontrollers.
>> That one I mention costs $46/qty 100, or 46 cents each and has four
>> channels of 10 bit analog converter.
>>
>> Is 0.1 degree C inherent reference standard accuracy necessary for
>> incubation?  No...
>> repeatability is nice, an easy cal feature is nice...  Your code can have
>> a mode where
>> when you press a calibrate button, it changes its look up tables to use
>> what your external
>> fancy temp sensor reference standard says.  The code could take input as
>> up/down buttons
>> to adjust the temperature lookup table while controlling temp at 35 deg C.
>>  You would put the tip
>> of your hi res. platinum temp probe in the incubator for the calibration
>> for a few minutes for each
>> up/down command until stable at 35.0 deg C.  Then it will repeat, and be a
>> transfer standard
>> for the cal thermometer for many months or years with as good accuracy and
>> precision
>> as the cal thermometer even though it does not have an internal temp
>> standard that
>> accurate.  It does need a very stable, repeatable volt standard inside or
>> on board, and
>> to make no changes to the chips used to keep its cal.
>>
>> John
>>
>>
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>>
>


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