USE OF DISCRETE SPACERS FOR THE SEPARATION OF PROTEINS BY GEL ISOTACHOPHORESIS

Discrete amphoteric spacers can generate a linear pH gradient concomitant with a conductivity gradient on isotachophoresis. The design of the system requires a mixture of discrete spacers calculated to generate an electrophoretic mobility-pH or charge-pH relationship as linear as possible and a mixture of counter ions calculated to provide a buffer capacity proportional to the charge of the mixture of spacers over the expected pH gradient. The separation of human blood plasma proteins in agarose gels is used as an example. The spacers were amphoteric ions: amino acids, amino acid derivatives and sulphonic compounds. The results obtained using mixtures of discrete spacers resemble those obtained using commercially available ampholyte mixture as judged from the separation of the C3-complement proteins. The major advantage of the use of discrete spacers in ITP lies in the possibility of designing separation systems for specific proteins.