GE HEALTHCARE AKTAprime plus, Quick Reference Instructions

Page: 21 / 40

11-0027-48 AC, 2007-09 • p21
Ordering information
Product Quantity Code No.
MBPTrap HP 5 × 1 ml 28-9187-78
HiTrap Desalting 5 × 5 ml 17-1408-01
100 × 5 ml
*
11-0003-29
HiPrep 26/10 Desalting 1
(53 ml)
17-5087-01
4
(53 ml)
17-5087-02
Superloop 10 ml 1 18-1113-83
Superloop 50 ml 1 18-1113-84
Superloop 150 ml 1 18-1023-85
Troubleshooting
High backpressure:
•
Column clogged
– Clean the column according
to instructions. Make sure the sample has been
centrifuged and/or filtered through a 0.45 µm filter.
•
System clogged – Replace the column with a piece
of tubing. Check pressure. If backpressure > 0.3 MPa,
clean system according to manual.
No binding:
•
Regenerate the column with 3 column volumes (CV)
water, 3 CV 0.5 M NaOH, 3 CV water, 5 CV binding
buffer before starting the run.
•
Check that the correct column is used.
•
Check that the inlet tubing from each buffer is
connected to the correct inlet port.
•
Check that the composition and pH of the buffers are
correct.
•
Check that the sample has been adjusted to the
binding buffer conditions.
•
Check that your sample contains target protein.
No elution:
•
Check that the inlet tubing from each buffer is
connected to the correct inlet port.
•
Check that the composition and pH of the buffers are
correct.
•
Use alternative elution conditions according to the
column instructions.
•
Check that your sample contains target protein.
*
Pack size available by special order
0.5
1.0
1.5
2.0
2.5
AU
280 nm
20
40
60
80
100
%B
0 5 10 15 20 25 30 min
Sample: Clarified lysate of E. coli expressing MBP-tagged protein.
Column: MBPTrap™ HP 1 ml
Binding buffer (port A1): 20 mM Tris-HCl, 200 mM NaCl, 1 mM EDTA, 1 mM DTT,
pH 7.4
Elution buffer (port B): 20 mM Tris-HCl, 200 mM NaCl, 1 mM EDTA, 1 mM DTT,
10 mM maltose, pH 7.4
0
0
5
Typical result