Profos changed the way to remove
endotoxins from aqueous
solutions
EndoTrap has become the chromatography
system of choice for removing endotoxin from biological solutions
during the last years. Since 2003 the Profos AG
successfully provides products to remove endotoxins from aqueous
solutions for the bioresearch and bioprocess markets. EndoTrap
eliminates common limitations connected with present methods
e.g. ultrafiltraion, ion exchange
chromatography, or two phase extraction.
Advantages
and disadvantages of current
methods for endotoxin removal: Many different
processes have been developed for the removal of LPS from proteins
based on the unique molecular properties of the LPS molecules. The
success of the techniques in separating LPS from proteins depends
strongly on the properties of the target molecule (e.g. net charge,
hydrophobicity) and the solution conditions (e.g. pH, ionic
strength). Only affinity chromatography is LPS
specific.
Current methods for
endotoxin removal |
Advantages /
Disadvantages |
ultrafiltration |
advantages: - easy
handling - suitable for small
proteins disadvantages: - not
LPS specific - not suitable for large proteins
|
two
phase extraction |
advantage: - cheap
procedure disadvantages: - not
LPS specific - traces of solvents have to be removed from
the target sample as they > interfere with high
resolution mass spectrometry > interact with
eukaryotic cell membranes > and have toxic
effects
|
heat
sterilization
|
advantages: - easy
handling disadvantages: - not
LPS specific - not applicable to bioreagents like
recombinant proteins that are destroyed at high temperatures
as well
|
ion exchange
chromatography |
advantages: - easy handling
- reusable - no limitation considering the molecular
weight disadvantages: - not
LPS specific - negatively charged
proteins may co-adsorb onto the matrix and cause a
significant loss of target protein -
positively charged proteins could form complexes with
LPS, causing the proteins to drag LPS along the column and
minimizing the LPS removal efficiency
|
affinity
chromatography |
advantages: - easy
handling - specific LPS
capture - reusable - no limitation
considering the molecular
weight disadvantage: - not
suitable for viscous
solutions
| Affinity chromatography is based on
the specific adsorption of a molecule to a ligand. Almost all
biological molecules can be purified on the basis of specific
interaction between their chemical or biological structure and a
suitable affinity ligand. EndoTrap affinity chromatography is one
principal method for cleaning biological solutions from endotoxin
contaminations. The EndoTrap ligand is a protein by nature, which
binds to the bead-matrix by stable covalent bonds. However, leakage of minute of amounts of ligand is a matter of
fact for all affinity materials and testing on these
contaminants is often required for regulatory purpose. Profos
provides leakage data for EndoTrap blue and EndoTrap red and a
Leakage ELISA for EndoTrap HD.
Advantages and disadvantages
of current affinity chromatography
methods for endotoxin removal:
Affinity
chromatography
|
Advantages /
Disadvantages |
Ligand: Polymyxin
B *
|
advantages: - specific LPS
capture (Lipid-A) - easy handling - many
customer publications
available disadvantages: -
Polymyxin B is an antibiotic
substance (you have to consider
that you will contaminate your target molecule with
an antibiotic substance!) - most
products based on Polymyxin B have low LPS binding
capacity - pH limitation - temperature limitation -
the removal procedure needs often incubation times -
regeneration often with DOC which is toxic for cells - no
leakage data available - not suitable for viscous
solutions
|
Ligand: "solid phase
reagent" *
|
advantages: - specific LPS
capture - nearly no temperature
limitation disadvantages: -
low LPS binding capacity - pH limitation - for the
removal procedure incubation step is recommended -
regeneration with 1M NaOH, therefore intensive washing steps
necessary - additional equipment necessary: peristaltic
pump - sample must be filtered before being applied - no
leakage data available - not suitable for viscous
solutions
|
Ligand: polylysin
* |
advantages: - high LPS
binding capacity - nearly no temperature
limitation disadvantages: - pH
limitation - complex protocol for the removal
procedure - regeneration limitation (only 5-times) - no
leakage data available - not suitable for viscous
solutions - no customer publications available
|
Ligand: peptides
*
|
advantages: - high LPS
binding capacity (Lipid-A) - nearly no temperature
limitation - nearly no pH limitation -
reusable disadvantages: - only
for small sample volume (1 ml sample / 1 ml resin) -
regeneration with DOC which is toxic for cells - no leakage
data available - not suitable for viscous solutions - no
customer publications available
|
Ligand: EndoTrap blue
or EndoTrap
red
(protein-based LPS-binding
ligand)
|
advantages: - high LPS
binding capacity (inner core region of LPS) - up
to 50 ml sample / 1 ml resin - nearly no temperature
limitation - nearly no pH limitation - best sample recovery (compared to
competitive products) -
best LPS removal (compared to competitive
products) - reusable - low ligand
leakage - leakage data available - many customer
publications available (link) disadvantages: -
not suitable for viscous solutions
|
Ligand: EndoTrap
HD
launched June 2008
|
advantages: - can be used in
early or late biomanufacturing process step - high
LPS binding capacity (inner core region of LPS) -
nearly no temperature limitation - nearly no pH
limitation - best sample recovery - best LPS
removal - reusable - robust and
chemically defined matrix -
excellent chromatographic characteristics - low
ligand leakage - Leakage ELISA available - Regulatory
Support File
available disadvantages: - not
suitable for viscous solutions
| * Product information according website or protocol. We can
not give any guarantee for this information.
EndoTrap
represents a benchmark in the field of endotoxin
removal technology based on affinity chromatography (e.g.
binding capacity) in the meantime. Of course there exist
various products for the removal of endotoxin but we are not aware
of any other product that combines all important advantages like
EndoTrap. Through its efficiency and its favourable and unique price
performance ratio EndoTrap is the first choice for the removal of
endotoxins during research and biomanufacturing process.
EndoTrap Customer
Feedback
|
Customer
Feedback (EndoTrap blue, EndoTrap
red) |
- "Probably the best system of recent
years." - "... We are not aware of any other method or
commercially available product, which allows the removal of
endotoxins from native protein samples as the last step of the
protein purification protocol with the same efficiency and recovery rates as we have
experienced with the EndoTrap-family products. It is
also worth mentioning the adequate, competent, and immediate
support from the Profos AG team. The way of interaction is
very straight forward, uncomplicated, and solution
oriented." - "... We compared the EndoTrap red columns with
several other LPS removal procedures and found this to be the
best system in our hands." |
Our EndoTrap customers among others
are from ... *
|
Biotechnology Companies
(like AstraZeneca, Cytos Biotechnology, ESBATech,
Integrated DNA Technologies and Micromet), Big Pharma (like Baxter and Eli
Lilly), Research Institutes
(like DKFZ, INSERM, NCI-Frederick, RKI and The Scripps
Research Institute) and over 400 universities (like Charité, Johns Hopkins University and
TU Munich) world-wide.
| *
EndoTrap customers who agreed to mention them as
reference.
Within only four years (customer
publications considered: November 2004 - October 2008) more than 90 papers were published
by the R&D market (universities, research institutes,
biotech companies). The newest product within the EndoTrap-family
was launched in June 2008 (EndoTrap HD for biomanufacturing
processes) and underlines that Profos has taken the next step
to become a global market leader in the field of endotoxin removal
systems and high-quality endotoxin-free products.
Overview
about the differences between the
EndoTrap products:
|
EndoTrap
blue |
EndoTrap
red |
EndoTrap
HD |
LPS binding capacity |
2.000.000 EU/ml |
2.000.000 EU/ml |
5.000.000 EU/ml |
Support matrix |
highly cross-linked 4% agarose |
highly cross-linked 4% agarose |
hydrophilic,cross-linked methacrylic polymer |
pH stability |
4-9
|
6-9 |
4-9 |
Ionic strength
|
up to 600 mM NaCl |
up to 250 mM NaCl |
up to 1.000 mM NaCl
|
Tested kind of substances which can be
applied |
proteins, peptides, enzymes, antibodies, plasmid DNA,
phage particles, buffers |
proteins, peptides, enzymes, antibodies, phage particles,
buffers |
proteins, peptides, enzymes, antibodies, plasmid DNA,
phage particles, buffers |
Package size |
ready-to-use columns (gravity flow) or slurry
|
ready-to-use columns (gravity flow) or slurry |
only available as slurry |
Intended use |
Laboratory scale, Research use only
|
Laboratory scale, Research use only |
For biomanufacturing & large scale process,
Biomanufacturing & Research use.
|
Note |
need 100 µM Ca2+ for efficient binding of
LPS |
|
need 100 µM Ca2+ for efficient binding of
LPS |
|
|
|
Leakage ELISA & Regulatory Support
File available |
Profos is located in the BioPark I and II in
Regensburg, one of the most important biotechnology centers in
Germany. With more than 1700 sqm of office, lab and production
space the facility houses state of the art equipment and
instrumentation for biochemistry and
molecular biology research. Profos AG is certified
according to ISO 9001:2000 and 13485, the
internationally recognized quality management system standard
developed by the International Organization for
Standardization (ISO), for its Regensburg facilities of
Research & Development, Production and Marketing &
Sales. |
|
Profos AG Josef-Engert-Str. 11 93053
Regensburg Germany
email: mailto:stephanie.steck@profos.de homepage:
http://www.profos.de/
This newsletter is a service of Profos AG.
Sitz der Gesellschaft: Regensburg Registergericht: Regensburg
HRB 8175 Aufsichtsratvorsitzender: Dr. Hellmut
Beckstein Vorstand: Dr. Wolfgang Mutter (Vorsitzender); Dr. Bernd
Buchberger; Dr. Stefan Miller; Oliver Glück
|