Endotoxin Removal Kit

Instruction manual

FOR IN VITRO AND RESEARCH USE ONLY
NOT FOR USE IN CLINICAL DIAGNOSTIC PROCEDURES

First Edition (Revised on April, 2016)

[ DESCRIPTION ]

Lipopolysaccharide (LPS) is a bacterial endotoxin, and a major constituent of the outer leaflet of the outer membrane of Gram-negative bacteria (e.g., E. coli). Endotoxins are shed in small amounts during active bacterial growth and released in large amounts upon bacterial death. During lysis of bacterial cells for plasmid preparations, endotoxin molecules are released from the outer membrane into the lysate. So, the removal of these endotoxins is highly necessary for downstream processes.

[ INTENDED USE ]

The Endotoxin Removal Kit can quickly and effectively reduce endotoxins level to <0.1 EU/mL in solutions containing proteins via the immobilized polymyxin B, which is well-established for capturing endotoxin and mitigating toxic effects.

[ KEY FEATURES ]

Appearance	Agarose beads suspension in 20% ethanol
Matrix	Sepharose 4B
Ligand	Modified PMB (Polymyxin B)
Resin content	2mg/mL cross-linked agarose
Mean Bead Size	90μm
Target Molecule	LPS
Binding Capacity	≤2,000,000EU/mL resin
pH Stability	pH 6-10
Reusability	Up to five times with proper regeneration

[ REAGENTS AND MATERIALS PROVIDED ]

Component	Quantity(50T-48S)
Equilibration Buffer	50mL
Regeneration Buffer	50mL
Endotoxin Removal Agarose	1mL in column



Instruction Manual	1 copy

[ STORAGE OF THE KITS ]

Store at 2-8℃ for up to one year. Do not freeze.

[ ASSAY PROCEDURE ]

1.Wash the column with 5mL of Equilibration Buffer at a flow rate of ≤1 mL/min, and allow the column to drain completely.

2.Apply sample at a flow rate of ≤0.3mL/min and allow it to drain completely, collect the flow-through and detect the endotoxin level of the collected samples.

3.Wash the column with 10 ml of Regeneration Buffer at a flow rate of ≤1 mL/min, and add 20% ethanol for storage.

[ TROUBLESHOOTING ]

Problems	Possible Cause	Solutions
Low removal efficiency	The pH of the sample is not within pH 7-8 range.	Adjust sample pH to 7-8.
	The contact time between sample and the resin is insufficient.	Increase the contact time by decreasing the flow rate.
	The removal or detection system is contaminated by exogenous LPS.	Use endotoxin-free labware.
	LPS binds to target protein strongly.	1. Optimize the pH of sample buffer for disaggregation. 2. Increase the contact time by decreasing the flow rate.
High sample loss	Non-specific binding of sample to resin.	Increase the concentration of NaCl in the sample and Equilibration Buffer.
	Target protein aggregates with LPS and is co-removed.	1. Optimize the pH of sample buffer for disaggregation. 2. Increase the contact time by decreasing the flow rate.