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Traditional Bead and Biotinylation Technology (TBB)

TBB is a well established, albeit dated, method for target identification, and validation.
[Optimum experiment design and higher probability of success.]


Working Principle

This method relies on enriching target proteins of a ‘bait-molecule’ by preparing ’bait-molecule’ specific affinity matrix. Affinity isolation can be achieved by either immobilizing bait-molecule on solid-phase beads or by coupling the bait molecule to universal isolation agents such as biotin.



Workflow

In TBB, the ‘bait-molecule’ is appropriately derivatized and coupled to either solid-phase or biotin. Target proteins due to inherent affinity interact with the bait-molecule and can be isolated by pre-immobilizing bait-molecule with solid-phase or biotin. These proteins are then identified and quantified using mass-spectrometry. Specific target proteins are de-convoluted by comparing the identity and quantity of identified protein with the proteins that were identified from control experiments.


Key Advantages


Points of Consideration


References
Technical Note 1
(PDF, 484KB)
This note explains various steps involved in de-convoluting the right target(s) of small molecule by immobilizing the bait-molecule on solid support.

Application Note 1
(PDF, 172KB)
This note describes the application of the technology in identifying/de-convoluting true positive targets of a test-molecule BIS-III.


FAQs

1. Can Shantani help analyze the SAR for identifying a site on a molecule that can be derivatized without activity loss? Yes, Shantani can synthesize the compound on case to case basis.


2. Can Shantani synthesize the primary amine derivatives? Yes, Shantani can synthesize the compound on case to case basis.


3. We can provide primary amine derivatives but how much molecule typically is needed for a typical target deconvolution experiment? Typically molecule provided in the range of 5-10 mg is sufficient enough for the purpose.


4. How much cell-lysate is needed? Can Shantani culture the cells? We prefer that ~15 mg of protein(cell-lysate) is provided for the purpose. In case not, we can culture the cells and generate the required amount of cell-lysate as long as either seed-culture of the cell and culture protocol is provided OR commercial source for the same is located.