Western-blot is not the easiest way to detect short peptides but if you have the right antibody or probe, it could be possible.
1/ You have to load an high amount of your peptide compared to a regular mid or high MW protein. I guess you have to load µg of your peptide if you want to detect it with a classical ECL system.
2/ For electrophoresis, you can use highly concentrated gradient gels (eg 4-20%) or Tricine PAGE type gels (16.5% minimum).
3/ For WB, you have to tranfer for a shorter time than for mid or high MW proteins as short peptide may go through the PVDF or nitrocellulose membrane. One option for a first trial could be to put 2 membrane one behind the other and to reveal the two membranes
However, if you would like to detect yur peptide routinely, one good option should but to shift from Western blot to an ELISA type test which are more appropriate to detect small peptides.
All the Best ;)
Western-blot is not the easiest way to detect short peptides but if you have the right antibody or probe, it could be possible.
1/ You have to load an high amount of your peptide compared to a regular mid or high MW protein. I guess you have to load µg of your peptide if you want to detect it with a classical ECL system.
2/ For electrophoresis, you can use highly concentrated gradient gels (eg 4-20%) or Tricine PAGE type gels (16.5% minimum).
3/ For WB, you have to tranfer for a shorter time than for mid or high MW proteins as short peptide may go through the PVDF or nitrocellulose membrane. One option for a first trial could be to put 2 membrane one behind the other and to reveal the two membranes
However, if you would like to detect yur peptide routinely, one good option should but to shift from Western blot to an ELISA type test which are more appropriate to detect small peptides.
All the Best ;)
Another way if ELISA or WB don't work for you, then you could resort to mass spec if available.
- Badges
- Science method