Endotoxin Removal Kits- For DNA

Overview

Reduce endotoxin levels to 0.1 EU/µg DNA or less
Available in 3 formats: Mini, Midi, and Maxi to suit your desired input volume
Remove endotoxins from up to 1 mg of DNA with the Maxi format kit
Fast and easy processing using a rapid spin-column format

These kits are designed for the rapid spin column removal of endotoxins from previously purified DNA. Norgen's spin columns bind DNA while endotoxins, salts and other contaminants are washed away. These kits reduce endotoxins to 0.1 EU/μg DNA or less providing plasmid DNA that is immediately ready for transfections or other endotoxin-sensitive applications. Typical recovery of DNA is >90% of the starting sample.

Endotoxin Removal Kit (Mini)

This kit is designed for the rapid spin column removal of endotoxins from up to 25 µg of previously purified DNA. The convenient spin column procedure can be completed in approximately 20 minutes.

Endotoxin Removal Kit (Midi)

This kit is designed for the rapid spin column removal of endotoxins from up to 200 μg of previously purified DNA. The convenient spin column procedure can be completed in approximately 30 minutes.

Endotoxin Removal Kit (Maxi)

This kit is designed for the rapid spin column removal of endotoxins from up to 1 mg of previously purified DNA. The convenient spin column procedure can be completed in approximately 30 minutes.

About Endotoxins

Endotoxins (also called lipopolysaccharides), are cell-membrane components of Gram-negative bacteria such as E. coli. Endotoxins are released during the lysis step of plasmid purification and significantly reduce transfection efficiencies in endotoxin sensitive cell lines. Therefore, the removal of endotoxins from plasmid preparations is often necessary prior to the use of the DNA in such downstream applications.

Details

Supporting Data

Figure 1. Efficiently Remove Endotoxins. An input of 11 µg of a plasmid in a 50 µL volume with an endotoxin level of over 8 EU/µg was subjected to clean-up by Norgen's Endotoxin Removal Kit. One microliter of the input and the 50 µL eluted DNA was diluted 1000-fold or 10 fold, respectively for a 100 µL end-point Limulus Amebocyte Lysate test for endotoxin level. The endotoxin levels of the plasmid DNA sample were reduced from over 8 EU/µg to less than or equal to 0.1 EU/µg of plasmid DNA using Norgen's Endotoxin Removal Kit (Mini), with high DNA recovery (see Figure 2).

Figure 2. High DNA Recovery. An input of 11 µg of a plasmid in a 50 µL volume with an endotoxin level of over 8 EU/µg was subjected to clean-up by Norgen's Endotoxin Removal Kit in triplicate. The 50 µL DNA elution was quantified by spectrophotometry. There is very little loss of DNA associated with using Norgen's Endotoxin Removal Kit (Mini). Recoveries of plasmid DNA are greater than 90% of the input amount when using this kit with efficient endotoxin removal (see Figure 1.)

Kit Specifications
Maximum DNA Input	25 μg
Final Endotoxin Level	≤ 0.1 EU/µg DNA
Size of DNA Purified	Up to 13,000 bp
Maximum DNA Volume Input	100 μL
Average Recovery	> 90%
Time to Complete 10 Purifications	20 minutes

Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. This kit is stable for 2 years after the date of shipment.

Component	Cat. 22700 (25 preps)	Cat. 52200 (10 preps)	Cat. 21900 (4 preps)
Buffer SK	15 mL	60 mL	60 mL
Wash Solution H	18 mL	18 mL	18 mL
Elution Buffer I	6 mL	12 mL	12 mL
Endotoxin Removal Solution	1.5 mL	1.5 mL	1.5 mL
Precipitation Solution	-	1.5 mL	1.5 mL
Spin Columns (assembled with Collection Tubes)	-	10	4
Spin Columns	25	-	-
Collection Tubes	25	-	-
Elution Tubes (1.7 mL)	25	-	-
Elution Tubes (15 mL)	-	10	-
Elution Tubes (50 mL)	-	-	4
Product Insert	1	1	1

Documentation

PROTOCOLS

(22700) Endotoxin Removal Kit (Mini) - Protocol (25 prep)

(52200) Endotoxin Removal Kit (Midi) - Protocol (10 prep)

(21900) Endotoxin Removal Kit (Maxi) - Protocol (4 preps)

SAFETY DATA SHEETS

(21900) Endotoxin Removal Maxi Kit - SDS

(22700) Endotoxin Removal Mini Kit - SDS

(52200) Endotoxin Removal Midi Kit - SDS

FLYERS

Flyer

FAQs

Mini, Midi, Maxi

Why do I have poor DNA recovery?

Poor DNA recovery could be due to one or a combination of the following factors:

DNA did not bind properly to the column.
Ensure that the Buffer SK does not contain any precipitates. Warm and mix gently if necessary.
The appropriate amount of ethanol was not added to Wash Solution H.
Wash Solution H has been specifically designed to contain the appropriate amount of components. Ensure that Wash Solution H is prepared using the correct amount of ethanol.
Incomplete removal of Wash Solution H.
Ensure that the column is spun for specified time minutes after the wash step, in order to completely dry the column. Traces of ethanol may remain in the eluted sample otherwise, and interfere with subsequent enzymatic reactions.
The appropriate amount of Buffer SK was not added.
Ensure that 5 volumes of Buffer SK is added for every 1 volume of DNA processed. The DNA volume must not exceed the specified volume for each kit.

Why is the purified DNA not performing well in downstream applications?

If the DNA does not perform well in downstream applications, it may be due to one or more of the following:

DNA was not washed with the provided Wash Solution H.
Ensure that the column is washed with the appropriate amount of Wash Solution H. This solution has been specifically designed to remove all traces of ethanol and endotoxin.
Proper Elution Buffer was not used.
The provided Elution Buffer I has been optimized for endotoxin-free recoveries. If endotoxin-free water is used for the elution, ensure that the pH is between 7 and 8.
A different Elution Buffer was used.
The provided Elution Buffer I has been optimized for endotoxin-free recoveries. The endotoxin-free properties of the eluted DNA will be compromised if another elution buffer is used. If a different elution buffer other than the one provided is used, the buffer should also be checked for any components that may interfere with the application. Common components that are known to interfere are high salts (including EDTA), detergents, and other denaturants. Check the compatibility of your elution buffer with the intended use.

Why are endotoxin levels in the eluted DNA slightly higher than 0.1 EU/μg DNA?

Endotoxin levels in the eluted DNA could become slightly higher than 0.1 EU/μg DNA due to the following:

A different Elution Buffer was used.
The provided Elution Buffer I has been optimized for endotoxin-free recoveries. The endotoxin-free properties of the eluted DNA will be compromised if another elution buffer is used. If a different elution buffer other than the one provided is used, the buffer should also be checked for endotoxin levels.
The endotoxin levels of the input were extremely high.
If the initial input DNA had extremely high endotoxin levels, the levels may not be completely reduced to 0.1 EU/μg of DNA or less. In this case, the eluted DNA could be applied to a second column and the procedure repeated in order to further reduce the endotoxin levels.

Citations

Title	Precision multidimensional assay for high-throughput microRNA drug discovery
Citation	Nature Communications 2023.
Authors	Benjamin Haefliger1 , Laura Prochazka1 , Bartolomeo Angelici1 & Yaakov Benenson

Title	Rational design and construction of multi-copy biomanufacturing islands in mammalian cells
Citation	Nucleic Acids Research 2023.
Authors	Altamura R, Doshi J, Benenson Y.

Title	Transplantation of prokaryotic two-component signaling pathways into mammalian cells
Citation	PNAS 2023.
Authors	J Hansen, E Mailand, KK Swaminathan, J Schreiber, B Angelici, Y Benenson