I am unable to understand how the data from ABI 7500 Fast can be analysed after the experiment is run. I tried to measure the melting temperature of a protein. I performed the experiment using 5X concentration of Sypro Orange and my protein at 4.5ug overall in a reaction of 20ul. I have run the experiment with initial temperature at 25oC and a final temperature to reach 99oC with 1% rise in temperature as a ramp speed. The experiment is run in triplicates. So, now I would like to know how can I make the average of three and get the melt curve, how can I draw the negative derivative curve and normalized curve, and finally how can I draw the melting temperature values for the analysed protein. The experiment is performed in Applied Biosystems 7500 Fast instrument. So, can someone share a detailed protocol of post experimental analysis of DSF stability assay to draw the melting point and for proper representation of the same by a melt curve.
Hi Vishweshwar, for analyzing your data after the experiment you need to have protein thermal shift software. This software will need the result file generated after your experiment, once you upload that file in this software, it will automatically show you the melt curves. It is a user friendly software, easy to understand too.
Thank you Deekshi for responding, but the software is expensive to procure. So, can you suggest me some manual way to do?
https://www.researchgate.net/post/Is_there_any_free_software_to_analyze_Thermal_shift_melting_curves
Shahid sir, I have gone through that, but the protocol is not clear, also they used some high end instrument for experiment, I am not getting the protocol they are describing
look for someone's article who has worked on the protocol you have used and then mail the corresponding article regarding the same. Try number of people.
- Badges
- Science topic
- Similar topics
- Biological Science
- Kinesiology
- Running