Dear Anita Dhara

can you clarify better yuor question?

Is the a purified lyophilized powder is purchased from a company, a protein?

Would you like to dissolve it and test is interaction with a different protein?

there are a many tecniques to detect protien protein interactions

e.g:

- SEC

- BLI

- SPR

- ELISA

- NMR (2D NMR 1H-15N- HSQC)

- Xray

- Cryo-EM

- Calorimetry (ITC)

- mass spectrometry:

- Cross linking mass spectrometry

- Non denaturing mass spectrometry (eg. ESI-MS)

- Deuterium exchange;

- Non reducing SDS page (in case of very strong interactions eg. streptavidin with biotinilated proteins)

there each of this technique as several pros/cons. Some techniques are able only to detect if the interaction happen (eg SEC) while other technique can provide you an idea of the affinity of the interaction (eg. BLI, SPR, ITC, ELISA) or the region involved in the interaction (eg NMR, mass spectrometry deuterium exchange)

There is not a perfect approach and you need to evaluate if your protein has the properties to be detectable through the specific approaches;

eg:

- BLI and SPR require that at least of of the 2 proteins contain a tag (e.g His) or modification (e.g biotinylation) that allow the immobilization of 1 of the 2 proteins in the sensors.

- NMR require the production of one of the 2 proteins uniformly labelled in 15N form

you can found more information about production of uniformly labelled proteins on the following link

https://www.blogger.com/video.g?token=AD6v5dwMieb--rE6-qY-WfM4fv8o-2PbxqaWuGxZ7_U3QB5tB113BQ1lEjHxpZhRMLWIWkeHmZV_LcuaESWuqDtZe1L3JI62cDvZaMQyL6BLVQxIRJtuE7IPw8SYCQHKWSV4_CCL0cs

good luck

Manuele

I am not an expert in this field, but I am very interested and have researched to find an answer. I received some assistance from tlooto.com for this response. Could you please review the response below to see if it is correct?

To study protein-protein interactions in vitro using purified lyophilized powders, a variety of techniques can be employed. One primary method is surface plasmon resonance (SPR), which allows real-time monitoring of interactions without labeling [1]. Isothermal titration calorimetry (ITC) can be used to measure the thermodynamics of binding events [2]. Additionally, pull-down assays and co-immunoprecipitation are useful for confirming physical interactions between proteins [3]. Other techniques include yeast two-hybrid assays for detecting interactions within a cellular context, and Förster resonance energy transfer (FRET) for studying interactions at a molecular level in solution [4].

Reference

[1]Nejati-Yazdinejad, M. (2007). Indirect determination of ascorbic acid (vitamin C) by spectrophotometric method. International Journal of Food Science and Technology, 42, 1402-1407.

[2]Linster, C. L., & Schaftingen, E. V. (2007). Vitamin C. The FEBS Journal, 274.

[3]Arya, S., Mahajan, M., & Jain, P. (1998). Photometric Methods for the Determination of Vitamin C. Analytical Sciences, 14, 889-895.

[4]Li, H., Tu, H., Wang, Y., & Levine, M. (2012). Assay of Vitamin C in Red Blood Cells. The FASEB Journal, 26.

Dear Manuele Martinelli ,

I want to dissolve it in a buffer and then test for protein-protein interaction. To check if those two proteins are interacting with each other or not. Also, they do have tags.

Thank you for your suggestions. It is helpful.

Best,

Anita