5% bio-lyte ampholyte
(3-10, Bio-Rad), bromophenol blue (a few grains).
, Pharmalyte pH 6-8 and pH 7-9, were obtained from Pharmacia.
The isoelectric focusing (IEF) was performed without ampholytes
and glycerol according to the manufacturer's instructions for peptide focusing.
Although gel-based techniques for protein separation are tremendously powerful, the observation of protein bands reflects a complex set of interactions between protein, ampholytes
, buffer composition, gel composition, and pore size.
The isoelectric focusing of proteins can be determined using immobilized pH gradients or ampholytes
5:1, 9 M urea, 2% ampholytes
(pH range 3-10), and 1% Triton X-100.
Proteins are ampholytes
, exhibiting amphoteric behavior owing to the presence of both positively- and negatively-charged side chain groups distributed along the length of the molecule and at its ends.
0 ml ampholytes
(pH gradient 4-5; Crescent Chemical, Hauppa, NY), 4.
1] Millipore 2D optimized carrier ampholytes
5; marine byssal precursors, in contrast, are highly basic - many with pIs exceeding the effective resolving range of available ampholytes
4% (v/v) carrier ampholytes
(pH 3-10, BioRad, USA) and 50 L of protease inhibitor cocktail (Catalogue No.