Cellufine™ MAX AminoButyl
Cellufine™ MAX AminoButyl is developed mainly for concentration of molecules having strong hydrophobicity such as VLPs (virus like particles). Cellufine™ MAX AminoButyl is designed on the basis of optimized Cellufine™ MAX Butyl to improve recovery of target molecules.
Ligand structure of Cellufine™ MAX AminoButyl is described in Figure 1.
Specification and characteristics of Cellufine™ MAX AminoButyl is described in Table 1 and 2.
| Item | |
|---|---|
| N contents (μmol/ml) | 20 - 33 |
| Elution volume (ml) α-chymotrypsinogen A (HIC mode) Pepsin (IEX mode) |
12.0 - 17.0 12.0 - 17.0 |
| Microscopic test (%) | < 5 |
Table 1Specification of Cellufine™ MAX AminoButyl
| Matrix | Highly cross-linked cellulose |
|---|---|
| Particle size | 90μm (40 – 130 μm) |
| Ligand | Butyl + primary Amine |
| Protein adsorption (1) in 2M (NH4)2SO4 | α-Chymotripsinogen A; ＋ Ribonuclease A; － Lysozyme; － |
| Protein adsorption (2) in 20 mM Tris-HCl (pH7.5) | Transferrin; － BSA; + Pepsin; + |
+; adsorption, －; no adsorption
Table 2Characteristics of Cellufine™ MAX AminoButyl
As base matrix is highly cross-linked cellulose particles, Cellufine™ MAX AminoButyl show superior flow property like figure 2.
(Condition; Column; 2.2 cm ID x 20 cm, Mobile phase; Pure water (24±1℃)
Application of Cellufine™ MAX AminoButyl
Purification of r-HBsAg (recombinant Hepatitis B surface antigen) VLPs from yeast with Cellufine™ AminoButyl
Partial purified r-HBsAg VLPs solution was loaded on packed Cellufine™ MAX AminoButyl column (16 mm I.D. x 500 mm H) and then column was washed by 20 mM phosphate buffer (pH 7.0) sufficiently. At first, phosphate buffer (pH 7.0) containing 0.1 % Triton X was used as elution solution (Elution 1). And then molecules was eluted by 2 M NaCl (Elution 2). Each fraction was recovered and analyzed.
The figure below showed a chromatogram of purification of r-HBsAg VLPs with Cellufine AminoButyl. r-HBsAg VLPs was detected by ELISA assay.
| VLP | Protein | |||
|---|---|---|---|---|
| nU | % | ug | % | |
| Load | 4,260 | 100 | 2,320 | 100 |
| Flow through | 480 | 11 | 350 | 13 |
| Elution 1 | 2,060 | 48 | 770 | 30 |
| Elution 2 | 172 | 4 | 1,190 | 46 |
Table below showed the results in this test.
Most r-HBsAg was obtained in elution 1. Results of protein assay in fraction suggested further purified r-HBsAg was concentrated by detergent elution. The results suggested Cellufine™ MAX AminoButyl is useful to purify VLPs.
Chemical Stability and Cleaning-In-Place
Cellulose is well-known as a natural product having chemical and physical stability. Thus, since Cellufine™ is derived from cellulose, it also is stable to chemicals and caustic and acidic solutions. CIP of all Cellufine™ media can be carried out with 0.5 M NaOH solution. Used media should be stored in 20 % ethanol at 2-25 ℃ after cleaning.
