Bio-Rad Aurum Ion Exchange Mini Kits - Manual (Page 7)

Fig. 1. Separation of three test proteins on Aurum AEX and CEX
columns.
The Aurum CEX column was equilibrated in 20 mM sodium acetate buffer, pH 5.0. The mixture of
three proteins (lane 3, 0.4 ml at 30 mg/ml total protein) was added to the column. The unbound
protein containing ovalbumin was collected (lane 4). The bound fraction containing conalbumin and
cytochrome c (lane 5) was eluted with 2 x 300 µl washes with 20 mM sodium acetate buffer, pH 5.0,
containing 1 M NaCl. The eluates from the washes were desalted and buffer exchanged using Micro
Bio-Spin 6 desalting columns (catalog #732-6221) and applied to an Aurum AEX column. The
unbound fraction containing cytochrome c was collected (lane 6), while the bound fraction containing
conalbumin (lane 7) was recovered with 2 x 300 µl washes of 20 mM Tris, pH 8.3, containing 1 M
NaCl. The fractions were assayed for protein using the Bio-Rad protein assay to determine protein
recovery. Purity was estimated from SDS-PAGE analysis on a 4–20% linear gradient Criterion
™
Tris-HCl gel (catalog #345-0033). The gel was stained with Coomassie Blue. Bio-Rad Precision Plus
Protein
™ standards (catalog #161-0373) are shown in lane 1.
5
1 2 3 4 5 6 7
Conalbumin
Ovalbumin
Cytochrome c
250 kD
150
100
75
50
37
25
20
15
10
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