BioHPLC Column Selection Guide Cover - Agilent Technologies
BioHPLC Column Selection Guide Cover - Agilent Technologies
BioHPLC Column Selection Guide Cover - Agilent Technologies
Create successful ePaper yourself
Turn your PDF publications into a flip-book with our unique Google optimized e-Paper software.
Method Development<br />
Bio Ion-Exchange <strong>Column</strong> Methods<br />
Bio MAb Monoclonal Antibodies<br />
Mobile Phase: A: 20 mM sodium<br />
phosphate, pH 5.5<br />
B: Buffer A plus<br />
400 mM NaCl<br />
Gradient: 10-35% B in 50 min<br />
Sample Size: 2 mg/mL;<br />
5 µL injection<br />
Temperature: Ambient<br />
Detection: UV 220 nm<br />
02 www.agilent.com/chem/biohplc<br />
Cation-Exchange (SCX, WCX)<br />
Monoclonal Antibodies, Proteins,<br />
Peptides, and Glycoproteins<br />
Recommended Initial Conditions<br />
Mobile Phase:<br />
A: 20 mM sodium<br />
phosphate, pH 5.0<br />
for WCX or pH 6.0<br />
for SCX<br />
B: Buffer A plus<br />
500 mM NaCl<br />
Gradient: 1-100% B in 30 min<br />
for 50 mm columns;<br />
60 min for 250 mm<br />
columns<br />
Sample Size: 2 mg/mL;<br />
5 µL injection<br />
Temperature: Ambient<br />
Detection: UV 220/280 nm<br />
Mobile Phase: A: 10 mM Tris buffer,<br />
pH 8 for WAX<br />
or pH 9.0 for SAX<br />
B: Buffer A plus<br />
400 mM NaCl<br />
Select Flow Rate Based on <strong>Column</strong> Diameter and Particle Size<br />
2.1 mm id <strong>Column</strong>s 4.6 mm id <strong>Column</strong>s<br />
Anion-Exchange (SAX, WAX)<br />
Oligonucleotides, Proteins<br />
Gradient: 1-100% B in 30 min<br />
for 50 mm columns<br />
and 60 min for<br />
250 mm columns<br />
Sample Size: 2 mg/mL;<br />
5 µL injection<br />
Temperature: Ambient<br />
Detection: UV 220/280 nm<br />
Particle Size, µm Flow Rate, mL/min Particle Size, µm Flow Rate, mL/min<br />
5 0.1-0.5 1.7 0.1-0.3<br />
1.7 0.1-0.8 3 0.1-0.5<br />
5 0.1-0.8<br />
10 0.1-1.0
Change language