Re: [DIYbio] GFP visibility vs concentration?

Or a Bradford or Lowry type assay:
http://en.wikipedia.org/wiki/Bradford_protein_assay
http://en.wikipedia.org/wiki/Lowry_protein_assay

On Fri, Jan 17, 2014 at 4:01 AM, Cathal Garvey
<cathalgarvey@cathalgarvey.me> wrote:
> IIRC, protein absorption is measured at 280nm? Make sure that's not in a
> significant absorption band for your GFP fluorophore, or the fluorophore
> might throw off your readings.
>
> That is, I don't think the 280 readings generally assume that the
> protein content of the solution has an unusually high absorbance at that
> wavelength, but a GFP variant might! :)
>
> On 17/01/14 03:35, kenny kostenbader wrote:
>> Nathan, remind me to come back to you when I have a scientific
>> heavy-lifting problem on my hands :) that's quite the solution!
>>
>> Avery, I'd be able to get a value for mg/ml of protein from an absorbance
>> reading.
>>
>> I was hoping to find a set of pictures that compare jars of GFP with
>> differing concentrations, something like that. When I get to my own
>> samples, I'd be happy to perform some dilutions and post pictures
>> somewhere. Someone out there will find that interesting :)
>>
>>
>> Kenny
>>
>>
>> On Thursday, January 16, 2014 7:50:08 PM UTC-5, Avery wrote:
>>>
>>> or you could get some GFP and serially dilute it until you cant see it.
>>>
>>> Filters help block the light that causes the excitement, but also make the
>>> overall image dimmer.
>>>
>>> How do you measure GFPs/volume?
>>>
>>> --A
>>>
>>>
>>> On Thu, Jan 16, 2014 at 6:17 PM, Nathan McCorkle <nmz...@gmail.com<javascript:>
>>>> wrote:
>>>
>>>> you might try finding the dosage (watts of light per area... i.e.
>>>> w/cm^2) that photobleaching of GFP occurs at, then from there
>>>> determine the maximum fluorescence (watts of light per area, or # of
>>>> photons), then find the minimum detection limit for the eye for that
>>>> wavelength (watts/area or photons).
>>>>
>>>> With that you should be able to determine how much light a single GFP
>>>> gives off at full brightness, then you can divide the minimum
>>>> sensitivity of the eye by that number to get # of GFPs needed. You
>>>> could then modify the equations with lower input power, in case you
>>>> don't have a super-bright UV light that is capable of photo-bleaching.
>>>>
>>>> On Thu, Jan 16, 2014 at 2:26 PM, kenny kostenbader
>>>> <kenst...@gmail.com <javascript:>> wrote:
>>>>> Hi All,
>>>>>
>>>>> I was wondering how much GFP (in aqueous solution) would be required to
>>>> glow
>>>>> bright enough to see with the naked eye? Or does it depend more on the
>>>>> intensity/color of the excitation lighting?
>>>>>
>>>>>
>>>>> Thanks!
>>>>> Kenny
>>>>>
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>>>
>>>
>>



--
-Nathan

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