Reproducibility of immobilized pH gradients after seven months of storage

Abstract Recently we described a procedure for pouring immobilized pH gradients at ambient temperature in the presence of a neutral buffer (0.1 M Tris‐phosphate pH 6.8) (Altland and Rossmann, Electrophoresis 1985, 6 , 314–325). After washing in plain water and in 1 % glycerolin water, the gels were dried, sealed in polyethylene bags and stored at –20°C until use. The reproducibility of a batch of 20 gels with a linear pH 6–10 gradient, poured under computer control, was studied by rehydrating the dried gels in a solution containing 8 M urea, 50 m M dithiothreitol and 0.5 % carrier ampholytes (pH 6–10) and by running human red cell lysates as samples. A comparison of the globin patterns obtained at various time intervals within a range of 7 months revealed that there was no detectable shift in the pH gradient within this period and that gels stored for 7 months can be used to clearly identify all human globins from newborns and adults. Since the tested pH gradient was prepared from Immobilines claimed as being the most sensitive to degradation by hydrolysis ( e. g. Immobiline p K a 8.5 and 9.3), it is concluded that storage of dry gels, rehydrated prior to use, is a realistic alternative to the preparation of fresh gels from stored stock solutions for achieving reproducible results in long‐term comparative studies.