Enhanced resolution of phosphoglucomutase (PGM 1 ) by the addition of a separator to ultrathin isoelectric focusing gels

Enhanced separation of phosphoglucomutase (PGM 1 ) isoenzymes has been achieved by addition of the separator N‐(2‐Hydroxyethyl)piperazine‐N′‐3‐propanesulphonic acid (EPPS) to pH 5–7 Ampholines in ultrathin and 1 mm isoelectric focusing gels. The resultant pH gradient ranges between pH 5.6 to pH 5.9 and effectively spreads PGM 1 isoenzymes across the whole width of the gel. The technique has been shown to be especially useful for separating some of the rare PGM 1 variants. The use of different electrolyte combinations has also been investigated and it has been shown that strong electrolytes, e. g. sodium hydroxide and orthophosphoric acid, result in a small but statistically significant increase in the gradient range compared with weaker electrolytes such as threonine/histidine. In addition, the effect of temperature on the rate of migration of PGM 1 isoenzymes has been investigated.