I came across a new fluorescent molecule (like fluorescein , BODIPY, etc.), which has a strong reactivity towards nucleophiles (through SN2) and basically it has a binding affinity towards different divalent and monovalent ions too. This dye is relatively very fast in changing its emission spectral properties immediately after binding with metals or reacting with its nucleophiles which brings an immediate shift in its spectral properties to identify different aspects.
How can this dye be shown or used to prove its applications in the aspects of biology (tagging, staining, etc.)?
I am interested in your views and suggestions to design some experiments for showing its biological applications.
if the molecule selectively interacts with certain metal ions then it can be used as a metal ion sensor e.g to detect the degree of certain metal contamination. This is one useful application which can have industrial significance too. Good luck !
The first experiment that I would do is to determine whether your molecule chelates/binds to biologically relevant metals such as Zn, Fe etc. If it does, then you have a lead.
In parallel and different experiment, you can do an affinity test of this tag with a protein (any protein for that matter) and check its spectral properties. If this happens, expand array of proteins(minimum 3 different type of proteins for first experiment). If it binds to everything, you basically have a non-specific tag, which is less useful.
If you have selective binding, you have a lead.
Dear Upendar,
for a concrete help and according to the message of Martin, you must give some infos about your fluorochrome.:
- Have you bought or synthesized this fluorochrome?
If bought, try to make a structure determination like elemental analysis, Fp, pH of the solution of your new fluorochrome in water UV, IR, NMR, solubility in aqueous biological media,...
If synthesized, you must be able to determinate the part of your fluorochrome reacting in the SN2 manner and the part acting as chelat (p.e. nitrogen or another group with oxygen....)
- When you write : " .. strong reactivity towards nucleophiles (Through SN2)..." the SN2 reaction gives a leaving group and it can affect the rate of the reaction. This is important concerning the biological application p.e. in vivo.
- 3 decades ago, I synthesized various news fluorochromes with a cyanine part for the conjugation trough the molecule and a phenanthroline part for chelating metal in the cells. Mitonchondria were stained in vivo but all the fluorochrome were toxic because the cells need metals for a " good life".
As you can see, the biological application will be certainly interessant IN VITRO for your fluorochrome and not IN VIVO. I agree with the message of Rupashree for industrial application.
More if you wish in due course!
JRG
Dear Grezes,
This fluorochrome was synthesized in the lab. All the elemental analysis was done and its solubility (Mostly in DCM, Chloroform, methanol.).
Just now I started to check for its toxicity in vivo.
All the above suggestions were good to start something with this ligand.
Thank you all..!!
Thank you for your fast answer. Concerning the toxicity in vivo during your next investigation:
- Certainly a selective toxicity will appear.
- Selectivity occurs through differences in distribution.
- The basis of selectivity can be found in cell wall, cytoplasmic membrane, organelles, EPRet. , microsomes a.s.o.. or in biochemistry (Human or animals.
When you work in water slutions, it is useful to know the ionization constant of your fluorochrome, the PKa and the stability constant.
- The oil/water coefficient of your fluorochrome must be found prior to an uptake by cells.
Good luck
JRG
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