DIY spectrometry for DIY bio ?!?

[Liz Barry]

Dear all, 

I wanted to send a quick email to introduce folks i've been chatting with about DIY spectroscopy and copy the main Public Lab spectroscopy list. On this very email, there's some amazing people representing DIY Bio communities:

DIYBio PDX in Portland, Oregon: Justin Dormandy

LA BioHackers in LA, California: Cory Tobin

Genspace in Brooklyn, New York: Ellen Jorgensen

Hackteria in many places: Marc Dusseiller, Rüdiger Trojok, Urs Gaudenz

(anyone else?)

and Sean of AlphaOneLabs and myself in NYC. 

So, to get started, I'd love to hear what the current and hoped for uses of spectroscopy are in your DIY Bio communities.

Just reply all and we'll gather them in this thread and see if any opportunities emerge.

We have a #SpectralChallenge open right now, and framing up some issues / events would be awesome.  

Thanks!

Liz 

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Liz Barry
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[Liz Barry]

Including Cory's correct address

publiclaboratory.org

+1-336-269-1539

[Brian Degger]

Am one of the diybio hackteria people. Love the spectrometry project by plots and pointed marc to the kombuchia post in march.
Cheers
B

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[Don Blair]

Hi Liz, All,

Liz, thanks for this!  Some quick notes, with a question for the list at the end ...

GFP. I've been playing around in a fruit fly biology lab at UMass Amherst, and one of the main techniques in the lab (and, as I understand it, in genetics these days) is to correlate the expression of certain genes with the production of a florescent protein, "Green Florescent Protein (GFP)" -- the neat result is that if a fruit fly is expressing a certain gene, it will glow green when UV light is shone on it.  From what I understand, techniques like this are widespread in biology, and there are an array of different proteins being used this way, each of which requires a different 'stimulation' wavelength (not all of them require UV), resulting in a different 'emission' wavelength (not all of them glow green). 

Currently, most labs use some fairly expensive hardware and software to accomplish this technique, but it seems as though the requisite hardware is really pretty cheap:  the relevant LEDs for stimulating these proteins can be purchased for a few dollars online, and the sort of techniques used in the Public Lab spectrometer seem appropriate to analyzing the results. There are also some filters, and protective eyewear to consider, but these generally seem low-cost, too. (In fact, the most basic case simply involves having a "flashlight" with the appropriate LEDs, and some safety glasses, and simply shining the flashlight on the organism to see if it glows. But when multiple florescent proteins are used, a spectrometer might be a really nice way of distinguishing the emissions.)

Bringing DIY spectroscopy techniques into the biology lab would likely save researchers a bunch of $$ -- but it also might lead to the prospect of novel 'biomarkers' in environmental applications.  E.g., there may already be research genetic variants of insects that could easily be tweaked to express GFP when an environmental toxin is present.  You could raise a crop of these insects locally, and if they're glowing, you know your back yard is contaminated with toxin X.  And if they're crickets, and they don't glow, you could fry them up and eat them.  For example.

But more generally, it seems that GFP and related techniques represent a safe, visually compelling, potentially very inexpensive way of getting folks engaged in DIY BIO research, and the Public Lab spectroscopy community could really help out here.

Colorimetry.  There are also a bunch of other applications in biochemistry and chemistry in which the color of a material indicates something important about the processes being studied.  There are lots of chemical assays in which the concentration of a chemical correlates with the color of the resultant solution, and this is widely used in analyzing specimens for environmental toxins.  E.g., there are a lot of "paper strip" tests that are very inexpensive, but which are difficult to calibrate, as they rely on ambient light conditions and the user's subjective assessment of what shade of color on the strip best matches a printed rubric.  Using a DIY spectrometer to assess these strips -- e.g. in a light-tight box, with a standard LED for lighting -- could be a nice, cheap way of developing a highly-calibrated chemical test.

Question to the list ... I'm sure folks on this list would have a lot more examples of ways in which cheaply & accessibly determining the precise color of a thing in a biology lab would make life a lot easier, and might lead to novel assays and experiments.  If you could set up a device that cost around $100 (cheap webcam, filter, Rasbperry Pi, power cable, ethernet cable, SD card) that would capture visible and near-infrared wavelengths (and perhaps some UV, too), and it could visually monitor e.g. a petri dish in the lab, capturing the spectrum every X minutes (storing it on the SD card, or putting it online)  ... what might that be used for?  Are there stains or reactions that evolve over time such that this information would be useful?  Note: if you're looking for a particular signature in a certain part of the spectrum, we could set up the software to notify the user when that part of the spectrum shows higher intensity.  E.g. perhaps growing cultures could be made much easier if folks had some automatic way of telling when the petri dish had filled up with the organism of choice ...

Very exciting stuff!  So far, when I've wandered into biology labs (quite ignorant of biology) and shown off some of these DIY tools, it has prompted the researchers into some immediate brainstorming around various applications in the lab ... can't wait to hear what you folks come up with!

Cheers,

Don

 

On Thu, May 2, 2013 at 1:43 PM, Liz Barry <l...@publiclaboratory.org> wrote:

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[Don Blair]

Hi Ellen,

Thanks! This just sent me on a very useful Wikipedia journey

http://en.wikipedia.org/wiki/Fluorometer

http://en.wikipedia.org/wiki/Spectrophotometry

I'm going to dig into these designs soon and try to see what aspects / applications might be amenable to inexpensive webcam sensors and lenses, filters, and LED light sources ... 

Quick question: is there a nice example / online reference re: GAP emissions?  GAP seems to be a difficult search term for Google to resolve to biology (without my knowing other info), instead of hip clothing :)

Cheers!

On Fri, May 3, 2013 at 9:16 PM, Ellen Jorgensen <ejorg...@genspace.org> wrote:

I think er have to distinguish between fluorimeters, which measure fluorescence like GAP emissions, reflected color like a colored bacterial colony on a dish, and absorbance of specific wavelengths which is what a spectrophotometer measures.

[Justin Dormandy]

Hrmm... lets try this again:

Hi everyone,

Sorry for the delay in getting back to this discussion. I feel like I've been working two full time jobs lately with school and trying to organize our little emerging DIYBio community here in Portland. I've only ever used spectroscopy to quantify DNA and determine protein contamination. I've also used it for colorimetric assays. However, I'm sure there are a very wide range of uses for spectroscopy that I'm not aware of or haven't thought about. One thing that might be useful for me to generate some more interest in my group is if I could report back to people, in laymens terms, why spectroscopy is useful and important. Generally when I've used spectroscopy to identify the quantity of some compound I have used samples that have been purified and only contains that single compound as an extra component over my blank. In the case of investigating DNA vs Protein in my sample it has gone through a DNA purification process where "ideally" the sample should contain DNA with maybe trace amounts of proteins.

However, I'm curious if spectroscopy can be used to identify compounds in a sample (tap water, lake, river, resevoir water, etc) when you don't know what compounds it might contain. Though I feel like, just thinking about it off the top of my head, it would be really difficult to identify whats in a sample since the optical densities/transmittance of each compound would interfere with the transmission and absorbance of others.

Thoughts?

-Justin (DIYBio PDX)

[Justin Dormandy]

This seems really interesting. The one application I can think of off the top of my head when it comes to excitation-emission in biology is used in flow cytometry. Basically you have a number of antibodies that bind to specific proteins. These antibodies then have a fluorophore conjugated to them that have specific exititation/emission spectra. Stains and the excitation-emission spectrum of the molecules themselves can also be used. The flow cytometer is designed so each drop should (but not always the case) each contain one cell. You analyze the resulting data to see how many cells contain X compound, Y compound, Z compound, what cells are in your sample, how many cells etc.

Your suggestion is interesting. If you had a heterogeneous mixture of cells you wouldn't be able to tell how many cells contain protein X over Y, etc, count them, nor see what different cell types are in your sample. However, if you had a homogeneous mixture of cells that started from a single colony (lets say we are using bacteria for the moment) you would be able to tell if they contain certain proteins or compounds just by going into a dark room and using a flashlight and LEDs calibrated to emit your excitation frequency, as you mentioned. The more I think about it you could actually roughly determine how many cells contained a certain compound. If you used a microscope with a hemacytometer (and your light emitted the frequency you needed for excitation) then you could count your cells of interest, establish a ratio of cells containing the compound your interested in, and statistically determine how many cells in your sample contain protein x,y,z, etc. Though, I just now realized I've just described fluorescence microscopy. But maybe there is potential here for creating a really cheap fluorescence microscope - they're not exactly cheap https://www.google.com/search?q=fluoresence+microscope&aq=f&oq=fluoresence+microscope&aqs=chrome.0.57j0l3j60.5484j0&sourceid=chrome&ie=UTF-8#q=fluorescence+microscope&source=lnms&tbm=shop&sa=X&ei=WX-EUYHxMqOWiAKixYAY&ved=0CAwQ_AUoAw&bav=on.2,or.r_cp.r_qf.&bvm=bv.45960087,d.cGE&fp=8bcec0c6f29b8c8d&biw=1317&bih=656

"If you could set up a device that cost around $100 (cheap webcam, filter, Rasbperry Pi, power cable, ethernet cable, SD card) that would capture visible and near-infrared wavelengths (and perhaps some UV, too), and it could visually monitor e.g. a petri dish in the lab, capturing the spectrum every X minutes (storing it on the SD card, or putting it online)  ... what might that be used for?"

Few things off the top of my head:

1. Growth of a protein product over time

2. If you were culturing cells in a flask and they had a unique absorbance spectra and if the intensity of an OD could correlate to confluency then you could set up a monitor to alert you when your cells are ready to be passaged to another flask

3. Similar to number 2, except use it to establish a growth curve for your cell/tissue cultures

4. An IGEM team made some bacteria a while back which that smelled like bananas in the presence of arsenic. How about instead of producing a smell, produce a product that has a very specific emission under white light. Spectrometer picks up this emission then sound alarm bells, start recording data, etc etc.

5. Find a protein (or another indicator) that has affinity for heavy metals and wont denature in the medium your trying to measure (ie blood urine, etc). Take a baseline reading, of urine for example, that will be your blank. Then add the protein (or other indicator). Subtract your blank and use the OD spectra of that protein (or exitation/emission) to detect metals ---or other compounds (drugs, toxins, etc).

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[Don Blair]

Hi All,

(Brian, Ellen, Justin -- thanks so much for the input and great ideas!)

Because Justin generated a ton of useful ideas and material, I didn't want it to get buried in the email thread, so I tried to distill a bunch of ideas that you folks generated into a wiki page on publiclab.org:

http://publiclab.org/wiki/diybio-ideas-and-applications

(I actually hadn't created a wiki page on the new site -- it's super fast and easy to edit!  If you haven't yet used this sort of wiki, putting in a hyperlink is really easy:  if I wanted you to go to CNN, I'd just write "Hey, you should check out [CNN](http://cnn.com)" ... and adding images is drag-drop!. Not your grandpa's wiki system.  Slick!)

I pulled out the following topics related to DIY BIO:

- General imaging

- Flow Cytometry

- Spectroscopy

- Research Questions

- Groups / Resources

And started to flesh them out (mostly using Justin's awesome text -- hope that's okay, Justin!)

Perhaps we can use that page as a place to collect ideas and links related to DIYBIO?  Also, in the Groups / Resources section, maybe you could all put links to your own organizations, and to any related groups you think might be useful ...

Looking forward to continuing this conversation / idea exchange!

Cheers,

Don

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[Liz Barry]

Hey all!

Getting back on this thread with a quick event description that Don and I just wrote up: 

http://pad.publiclab.org/p/diybio-ideas-and-applications

We'd love to hear your thoughts! feel free to edit that etherpad (which is linked from the page Don created http://publiclab.org/wiki/diybio-ideas-and-applications) and write back on this thread. If anyone is still have trouble posting to the spectrometry list, please let me know in a direct email. 

Thank you!

Liz

publiclaboratory.org

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[Liz Barry]

once again, this time with Cory's correct email! 

[gbathree]

Hey guys, I thought I'd jump in as well.  I'm working on a project called Photosynq (photosyqn.org) where we are measuring fluorescence (that is, IR light) from photosynthetic organisms (including plants and algae).  To read more about the basic idea see here:

http://en.wikipedia.org/wiki/Chlorophyll_fluorescence
and
http://www.hansatech-instruments.com/modulated_overview.htm

Pulse modulated fluorescence is more interesting than just measuring how much plants fluoresce (which can be seen from a properly filtered camera like this - or from satellites in space for that matter), because it can tell you exact information about how efficient the plant is at absorbing light, how much linear electron flow is happening, and other physiological data which relates to drought stress and plant health.  However, it's more complex than just reading fluorescence because, to take proper measurements, it requires that you also flash the sample with some light, which requires a microcontroller and fairly accurate timing on the LEDs.

I'm not sure if this is possible to include this kind of measurement, but it is interesting and very useful in plant science.  One example I can image being interesting is using this to measure algal concentration in water sources, as well as algal activity (so not only how much is there, but how healthy and photosynthetically active is it).  I'll ask some other members of our team to see if they have any other ideas.  I've done a lot of searching for different types of detectors and LEDs appropriate for this and would be happy to share them if that's helpful

Greg

[Tiberius Brastaviceanu]

Greg, what are the time parameters? 

We're starting to work on a pulsed system, containing a fast pulsing LD and a PD connected to a  luck-in amplifier, all in the ns time scale. The timing will be flexible on these circuits. But in this case, you're not using a camera, but only a photodiode. So you'll not have spatial information. We can share the electronics. In order to adapt this system to spectroscopy you'll need to add a variable filter. 

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[gbathree]

Wow, I've never seen Sensorica - fantastically interesting!  I just finished a talk at Ann Arbor Nerd Nite about many of the things you discuss in the "about" section of your site - it seems that you're very far ahead of the curve!  Very very cool, I'd love to learn more.

The time parameters are ~5us for a single pulse, and of course you need to be able to take at least 1 data point from the detector during that time.  The timing between turning on the measuring LED and saturating LED on or off at the same time should be as close as possible (right now it's ~500ns) and consistent throughout the run and between different units.

Our benchtop units have ns accuracy but they are using FPGAs which are hard to program and adjust.  We feel that for something to be truly accessible, we're trying to keep it as close to an arduino device as possible.  Then, typical arduino users will feel comfortable trying to fool with it.  If that's not possible, then I think we'll be significantly limiting our impact. Also, we chose the Teensy in part because there's a fantastic Teensy community continually making the unit better.  That being said, if we can get ns time scale resolution in a user accessible / low cost package that would be awesome!

Within the next month we'll have the schematic and board drawings for our device, which we'll share on our project page, but I'd love to discuss it further to see if there's ways we can make it better, or make it appropriate for other applications.  I'd also want Robert, our electrical engineer, involved since he can respond a lot more accurately about the technical details.

Greg

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[Sara C]

Sorry if this is a repost, but I'm interested in spectrophotometry for enzyme kinetics In the classroom. In particular, p-nitrophenol, which absorbs around 400 and nanometers is a nice colorimetric product, Since it is provided with many different kits, such as: http://www.bio-rad.com/prd/en/US/LSE/PDP/KVHOJU15/Biofuel-Enzyme-Kit

Might this be something the Desktop Spectrometry Kit could handle, with the correct calibration?

[Jeffrey Warren]

sure, 400nm is in the right range... are you looking to quantify its presence, or just detect any absorbtion?

On Thu, May 30, 2013 at 3:20 PM, Sara C <skyti...@gmail.com> wrote:

Sorry if this is a repost, but I'm interested in spectrophotometry for enzyme kinetics In the classroom. In particular, p-nitrophenol, which absorbs around 400 and nanometers is a nice colorimetric product, Since it is provided with many different kits, such as: http://www.bio-rad.com/prd/en/US/LSE/PDP/KVHOJU15/Biofuel-Enzyme-Kit

Might this be something the Desktop Spectrometry Kit could handle, with the correct calibration?

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[Sara C]

Ideally, it would be quantification over a time course. However, it could simply be end point.