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Cellufine ETclean Affinity Chromatography Media
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Introduction
The Cellufine ETclean is poly(ε-lysine) immobilized
Cellufine (cellulose spherical beads). The beads bind and remove endotoxin from your sample solution. The poly(ε-lysine)
is a microbial poly(amino acid) that consist of 30-35 lysine residues produced by Streptomyces albulus.
The poly(ε-lysine) was immobilized onto chloromethyloxirane-activated cellulose beads. The beads are a stable affinity beads that are resistant against the
cleanup solutions, which include 0.2 M sodium hydroxide and 2 M sodium
The Cellufine ETclean can remove endotoxin from a cellular product solution at physiological pH, ionic strength of μ = 0.02-1.0, and 0oC - 25oC.
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Electron micrograph of
Cellufine ETclean-S beads. |
| Cellufine ET clean S
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| Cellufine ET clean L
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| Partial Structure |
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| Name |
Supplied |
Wet Bead Diameter |
Pore Size* |
| Cellufine ETclean-S |
a slurry in 20 % ethanol |
ca. 40-130 μm |
Mlim 2000 |
| Cellufine ETclean-L |
a slurry in 20 % ethanol |
ca. 40-130 μm |
>Mlim 2x106 |
| *The pore size (molecular weight exclution; Mlim) of the beads was estimated from calibration curves obtained by size exclusion chromatography. Pullulan and maltose were used for the Mlim determination. |
| Selective adsorption of endotoxin (LPS) from a bovine serum albumin (BSA) solution by Cellufine ETclean beads. |
| Selective adsorption of endotoxin was determined using a batchwise method with 0.2 g of the wet beads and 2 ml of a sample solution (BSA: 500 μg/ml, E. coli O111: B4 LPS: 100 ng/ml, pH 7.0, ionic strength of μ = 0.05-0.8 ). |
| Selective removal of endotoxin from a protein solution by Cellufine ETclean beads. |
| Sample Solution |
Cellufine ETclean-S |
Cellufine ETclean-L |
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Compound
pI |
Concentration of endotoxin before treatment
(pg/ml) |
(μ = 0.05, pH 7.0) |
(μ = 0.05, pH 7.0) |
| Concentration of endotoxin after treatment
(pg/ml) |
Recovery of protein after treatment
(%) |
Concentration of endotoxin after treatment
(pg/ml) |
Recovery of protein after treatment
(%) |
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28,000 |
81 |
99 |
<10 |
95 |
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32,000 |
45 |
99 |
<10 |
97 |
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4,500 |
18 |
99 |
<10 |
98 |
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5,600 |
20 |
99 |
<10 |
97 |
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1,500 |
15 |
99 |
<10 |
98 |
| The removal of endotoxin was determined by a batchwise method with 0.3 ml of wet adsorbent and 2 ml of a protein solution (1 mg/ml) containing natural endotoxin. |
| ET removal Exp.
BSA / ETclean L |
| Column chromatography |
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Column size : 1 X 1.1 cm (I.D.) (1.1ml) |
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Flow rate 0.17 ml / min (10cm / h) |
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Buffer 50 mM PB, pH 7 + 0.15 mol NaCl aq |
| Assay |
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Protein Abs. at 280 nm |
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ET LAL rate assay |
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| - Injection sample (150 ml)
- BSA 1 mg/ml ET 100 EU/ml |
| Lysozyme / ETclean L |
| Column chromatography |
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Column size 10 x 0.9 cm (I.D.) (9.6 ml) |
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Flow rate 0.5 ml / min (47 cm / h) |
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Buffer 1 mM Tris-HCl, pH 7.3 |
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Gradient 0 - 1.0 mol / l NaCl aq. |
| Assay |
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Protein Abs. at 280 nm |
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ET LAL rate assay |
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| - Injection sample (1ml) : 14 mg / ml |
| Insulin chain A / ETclean-L |
| - Injection sample (1ml) : 13 mg / ml, 309 EU / ml |
| Tranceferrin / ETclean L |
| - Injection sample (1ml) : 13 mg / ml, 2982 EU / ml |
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