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cals are described below. See Separation procedures and Development
procedures for detailed instructions for using these products.
PhastGel IEF media
PhastGel IEF media are homogeneous (5% T, 3% C) polyacrylamide
gels containing Pharmalyte
®
carrier ampholytes. Pharmalyte
generates stable, linear pH gradients with a smooth conductivity
profile across the entire pH range, which means that high field
strengths of 500 volts/cm and above can be used. Three different
PhastGel IEF media are available: PhastGel IEF 3-9, 4-6.5 and 5-8.
PhastGel IEF media are run without buffer strips.
The histogram shown here illustrates the pH ranges of PhastGel IEF
media with respect to the pI distribution of 800 proteins. (See
Technical data, chapter 8, and Separation technique file No. 100,
chapter 9, for further details.)
PhastGel electrophoresis media
PhastGel electrophoresis media are used together with PhastGel
buffer strips. Buffer strips, made of high quality agarose with low
electroendosmosis and high purity reagents, serve as buffer reservoirs
to generate discontinuous buffer systems in the gels during a run.
During a separation, proteins are first concentrated in a porous
stacking gel zone, they then move into the separation gel zone where
they are separated according to size. The migration distance of a
protein is related to the logarithm of its molecular weight (MW).
Molecular weights are easily estimated using one of the
GE Healthcare
molecular weight calibration kits. (See Evaluation and
presentation of data, chapter 6, for instructions.)
Seven different gels for electrophoresis are available, three for gradient
gel electrophoresis and four for homogeneous gel electrophoresis. The
three gradient gels are PhastGel gradient 10-15 with a continuous
gradient from 10 to 15% polyacrylamide, PhastGel gradient 8-25
with a continuous gradient from 8 to 25%, polyacrylamide and
PhastGel gradient 4-15 with a continuous gradient from 5-15% total
polyacrylamide and a 1-2% gradient cross linker. Three of the homo-
geneous gels are PhastGel homogeneous 7.5, PhastGel homogeneous
12.5 and PhastGel homogeneous 20, with a concentration of 7.5,
12.5 and 20% polyacrylamide respectively. The fourth homogeneous
gel is PhastGel high density which has a polyacrylamide concentration
of 20% and a 30% concentration of ethylene glycol. (See Technical
data, chapter 8, and Separation technique files 111, 112, 120, 121
and 130, chapter 9, for further discussion and details.)
3. Description of the system
