Comparison of the Rapid Clear® Cap 3000 vs GE Capsule Filters for clarifying monospecific IgG

Introduction

Two methods to clarify 10L of post transfected IgG harvested from culture are compared. The culture was harvested on day 4 with 75% viability and a yield of 3.44e6/mL VCD.

Equipment

Method

Rapid Clear® Cap Filtrations

10L culture was split in half and run through 2 Rapid Clear® Cap 3000’s

GE Filtration ULTA Prime GF 5 µm 10” Capsule Filter and ULTA Prime CG 0.2µm 5” Capsule Filter

Two GE capsule filter columns are used in tandem. Culture is pumped through the columns and collected in a flask. ULTA Prime GF 5 µm 10” Capsule Filter and ULTA Prime CG 0.2 µm 5” Capsule Filter, p/n 12410247 was utilized.

Purification & Analysis

All filtered material was analyzed by HPLC to determine quantity and purity. A 25mL Mab Select Sure proA capture column was used on an AKTA system. Two washes with buffer were performed prior to elution with pH 3.5 buffer to neutralize. Total recovery for both clarification systems was equivalent at 115mg/mL for the Rapid Clear® Cap and 100mg/L for the GE method for post column purification comparison:

* Note: this IgG is known from prior work to have < 90% purity post proA

Results

Protein Yield Post Filtration & Final

fig.1Protein Yield Post Filtration (left) Post Purification (right) - For data see table 1 below

HPLC comparison of post purified Rapid Clear® Cap and GE column was performed using a proA column purification was performed for both the Rapid Clear® System and the GE clarified solutions. The Rapid Clear® System yielded a slightly higher quantity of intact IgG, see fig 2, than the GE even though there was a lower volume of clarified culture, see table 1.

fig.2HPLC comparison of the purified IgG. The Rapid Clear® Cap is the red line and GE the blue line.

fig.2Both GE and Rapid Clear® clarified IgG were run on an SDS Page gel for comparison: Lanes 1 & 4 are molecular weight standard ladder; Lanes 2&3 are non-reduced; lanes 5&6 are reduced.

Conclusion

Using the Thomson Rapid Clear® Cap 3000 was quick and easy. It needed less bench space and equipment increasing efficiency and minimizing waste. The Rapid Clear® Cap 3000 resulted in high quality Mab with higher yield per recovered volume over the GE, Table 1. The Rapid Clear Cap® 3000 had 20% higher recovery of intact protein than the GE capsule columns as calculated by % purity by HPLC divided by the total recovered volume.

Thomson Instrument Company is not affiliated with GE Life Sciences, Biorad or their products