Hu IgG2k aggregates on BioSep3000

LC Conditions

Column

BioSep 5 µm SEC-s3000 290 Å, LC Column 300 x 7.8 mm, Ea

Brand

Biosep

Part No

00H-2146-K0

Phase Name

SEC-s3000

Elution Type

Isocratic

Mobile Phase

A: 100mM Sodium Phosphate pH 6.8

Gradient Profile

Step No.

Time(min)

%A

%B

1

0

100

0

Flow Rate

1.00 mL/min

Temperature

22°C

Detection

UV-Vis Abs.-Variable Wave.(UV) @ (22°C)

Detection Info

Sample Notes

Application Focus: An example of Ig-G aggregate testing on BioSep 3000 Analyzing Ig-G based therapeutics for aggregates is probably the most common application that is performed on a GFC column. Most users are attempting to get best resolution possible between the Ig-G monomer peak and any associated dimers in the sample. In this example human Ig-G2 is injected on a BioSep 3000 column (300x7.8 mm) and dimer and aggregate is quantitated. Results show that the dimer peak of Ig-G is well resolved from the monomer peak, and that there appears to be two different dimer forms that are partially resolved by the BioSep 3000 column. In addition, there appears to be aggregate at the total excluded void of the column and the appearance of a possible trimer peak ahead of the dimer peak. Finally, there is a fragment peak that elutes after the Ig-G monomer peak, most likely attributed to an Ig-G that is missing one of its Fab fragment. These results show the utility of using BioSep 3000 for antibody aggregate analysis.