Isoelectric focusing in acrylamide gels: Use of amphoteric dyes as internal markers for determination of isoelectric points

Synthetic oligopeptides as isoelectric point markers for capillary isoelectric focusing with ultraviolet absorption detection

“…Instead of proteinaceous macromolecular pI markers, the use of low molecular mass ampholytes as pI markers has been proposed, examples of which are amphoteric dyes [4], phenanthroline iron complexes [5], and substituted aminomethylphenols [6]. Among these, the substituted aminomethylphenols showed practical importance as pI markers, since they cover a relatively wide pH range (5.3±10.4), although suitable markers are not available for pH 8.4±10.1.…”

Evidence for identical subunits in pea seed nucleoside diphosphate kinase

“…lsoelectric focusing of pea seed NDP kinase in polyacrylamide gel was performed according to ConwayJacobs and Lewin [12]. Pea seed NDP kinase, equine heart myoglobin (two times crystallized from Calbiochem), human serum albumin (Kabi) and ovalburain, grade V, (Sigma) were added, separately or in mixture, to the gel solution.…”

“…The final concentration was 0.1 mg per ml of each protein. A few microliters of a Patent blue V solution (2 mg/ml) were also added [12]. The gels were allowed to polymerize in bright light, and subjected to electrophoresis at 110 V for 16 hr in the cold room (+ 5°C).…”

“…The gels were allowed to polymerize in bright light, and subjected to electrophoresis at 110 V for 16 hr in the cold room (+ 5°C). Clearly visible precipitation discs were formed by all proteins tested, and the isoelectric point of NDP kinase was calculated as described [ 12].…”

Isoelectric focusing of proinsulin and intermediate in polyacrylamide gel

“…Acrylamide gels were prepared according to Conway-Jacobs and Lewin [3], merely, the volume of the solution was doubled and 200 ~1 of glycerol/ tube were incorporated. The protein samples, 0.1 mg per gel in glass tubes 12 X 0.6 cm inner diameter, were electrofocused for 9 hr under cooling by circulat- tions are biosynthetic intermediates or not and further studies will be required particularly for the detection of endogenous intermediates.…”