Measuring GPF's light absorbance

[deleted-112378]

Can I find the structure of GFP according to its absorbance of light through the spectrophotometer?

[sunmoonstars]

Hi Han,

If interested, you can read this paper deiscussing GFP - there is alot of good background in there, as well as the information about how the structure was originally solved. http://www-bioc.rice.edu/Bioch/Phillips ... fpbio.html
The spectrophotometer wouldn't tell you much about the structure itself. The spectrophotometer is good for measuring the light that passes through a solution, which correlates to the concentration of the solute it contains. This gives some good background about that: http://biology.clc.uc.edu/fankhauser/La ... ometer.htm
Please let me know if you have further questions.

Tonya

[deleted-112378]

Thank you for responding Tonya!
I looked at the first website and I found that GFP can be used as a protein tag, does this mean that it serves as a tracker?
How would I physically find the structure of GPF if I attempt to denature it at high and low temperatures and can I use the glow to measure the light absorbency in the spectrophotometer?

[sunmoonstars]

Hi Han,

Yes, GFP is a popular tracking protein. Once folded, it glows.

I don't think you will actually find the physical structure of the protein based on the use of the spectrophotometer, but you will be able to tell if the protein is folded (glowing) or not (not glowing).

Did you find any other sites to use for background, or to help decide how you are going to measure the glow?

Tonya

[deleted-112378]

I have not been able to find how I can measure that glow but would there be a blacklight setting on a spectrophotometer? How can I measure the glow of GPFs with a spectrophotometer? Is there a way for me to use the same amount of GFPs in my research that's easy to separate to use in temperature testing (should I have a constant weight or another constant variable?)?

I found this project to measure the glow but would this be a realistic project to do on top of the temperature and protein research or should I continue to search for methods?
https://www.sciencebuddies.org/science- ... ml#summary

[sunmoonstars]

I have not been able to find how I can measure that glow but would there be a blacklight setting on a spectrophotometer? How can I measure the glow of GPFs with a spectrophotometer? Is there a way for me to use the same amount of GFPs in my research that's easy to separate to use in temperature testing (should I have a constant weight or another constant variable?)?

I found this project to measure the glow but would this be a realistic project to do on top of the temperature and protein research or should I continue to search for methods?
https://www.sciencebuddies.org/science- ... ml#summary

Hi again,

That's a nice project! Based on the difficulty of the questions you are asking me, I think you can do something more advanced than that one.

There are spectros that have UV setting. You should first find the spectro you may use and see if it has the capability you will need. If not, then I do suggest finding an equipment to use before you are locked into a project. Since the spectros operating instructions vary, someone there would be familiar with its use and can help you.

You can buy the protein to use as your starting sample: (one of many places to buy it from: http://www.abcam.com/GFP-protein-ab84191.html ).

[deleted-112378]

My instructor has spectrophotometers for us to be using in our classroom so I hope that it has the setting you are talking about, I will be checking it today. Just to be sure, would this be a good way to check the lumiosity of the GFP?

Since I'm still getting started I am still readjusting the project to my tastes so I was thinking of posing this question: How does temperature affect the stability of GFP and will this affect the effectiveness of it as a biomarker?
I was planning to use fungi as the marked specimen because I still need to find out how to use animals or other plants to help find cancerous cells. If you have any suggestions please help! But I would much rather prefer looking into how this would help locate cancerous cells.

Also, I was planning to culture my own E. Coli colony and produce pGlo to extract the GFP because it is more cost friendly.

Did you mention that you have a more complex project in mind? I am open to suggestions and would be grateful if you shared your ideas!

Thank you so much for your constant help!

I was also wondering if you work for Biorad? If you do then are there any other professionals that you are aquainted with that I can email and gather more information from?

[deleted-112378]

I was also wondering how we would be able to make our own biomarkers.
Thanks!

[sunmoonstars]

Did you mention that you have a more complex project in mind? I am open to suggestions and would be grateful if you shared your ideas!

Thank you so much for your constant help!

I was also wondering if you work for Biorad? If you do then are there any other professionals that you are aquainted with that I can email and gather more information from?

Hi again,

I do work for a major life science company, but we have to keep all of our correspondance here on the forum, ok? I will ask the others if anyone has more input for you here, ok?

I don't have a more complex project in mind yet. Do you? I think you started this conversation with a question to be answered.

Have a great day!
Tonya