|Register||Search||Today's Posts||Mark Forums Read|
|Protocols and Methods Forum Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.|
| ||LinkBack||Thread Tools||Display Modes|
I need to stain polylisine - free and conjugated with peptide. Coomasie
Brillant Blue fails with free polylysine and - for my suprise - so does
I'm using silver staining method from Amersham Technical Manual (as it say -
based on Heukeshoven and Dernick 1985) but they do not describe which
aminoacids are essential for detection.
Maybe, the using of this method is limitted by F.W. of your
polylisine. It cannot apply to a small peptide. You can change another
polylisine with greater F.W.
Chris B. schrieb:
I used already Ag staining method described by Blum et al.
Electrophoresis 8 (1987) 93-99 for protein staining.
Also, this autor didn't describe which residues are necessary.
But there are just two possibilities how the method works:
(1) Ag+ from the fixing solution is reduced to Ag (colloid)
(2) Ag+ reacts with free -SH groups and forms an Ag-S-R complex
In the first case there must be amino acid residues present
that can be oxidized by Ag+ (cysteines and others).
In the second case there must be amino acid residues present
that can form sulfides with Ag+-salts (cysteines are the only
Chris B. wrote:
What is the molecular weight of your peptides? If it is too small, they
will be eluted from the gel during the staining procedure. Cross-linking
with glutardialdehyde may help in this case.
Also, polylysine will have a considerable positive charge, which would
interfere with Ag+ binding. Staining with an amine reactive fluorescent
probe may be the preferred option, either before or after
electrophoresis. Check the Molecular Probes catalog ([Only registered users see links. ]) for
|silver , staining|
|Thread||Thread Starter||Forum||Replies||Last Post|
|Immidiate dissappearing of bands on DNA sequencing gels after silver staining||himanshu||SDS-PAGE Gel Electrophoresis Forum||2||07-24-2008 06:17 PM|
|Problems with silver staining||terence diane fabella||Protein Forum||2||01-16-2008 05:04 PM|
|USP silver staining method help||leelavati shetty||Protein Forum||0||01-16-2008 04:17 AM|
|Silver staining||Chris B.||Protein Forum||3||08-30-2005 01:24 PM|
|Silver staining Western membranes||EK||Protocols and Methods Forum||14||09-27-2003 02:37 PM|