product brochure CANDOR - CANDOR Bioscience GmbH

Transcrição

BUFFERS AND
SOLUTIONS
PACKAGE SIZES – SUMMARY
LowCross-Buffer®
CONTACT
Antibody Stabilizer “TRIS”
Sample Buffer
50 mL order number 100 050
The Blocking Solution
Antibody Stabilizer “PBS”
Stripping Buffer
SmartBlockTM
Liquid Plate Sealer®
Coating Buffer “pH 7.4” 10×
BSA-Block
LowCross® HRP-Stab
HRP-Protector
TM
Coating Buffer “pH 9.6” 10×
Washing Buffer “PBS” 10×
Washing Buffer “PBS IHC” 10×
Washing Buffer “TRIS” 10×
Washing Buffer “TRIS IHC” 10×
Starter Package
small
order number 600 001
large
Starter Package “IHC”
small
large
CANDOR Bioscience GmbH
Simoniusstrasse 39
88239 Wangen, Germany
+49 (0) 75 22 - 7952 70
www.candor-bioscience.com
www.readytector.com
Ordering
[email protected]
Fax: +49 (0) 7522 - 7952 729
Technical support
[email protected]
General inquiries
[email protected]
Blocking Sampler Package
small
large
Please find more information at
PREFACE
CONTENT
True results for your measurements.
Immunoassays are an important tool
in everyday work for many scientists
and lab technicians. But regardless
whether we are experts in drug
discovery, medical research, veterinary
diagnostics, biotechnological quality
control, environmental analysis or
whatsoever – we do not perform
immunoassays only because it is a
funny amusement. We just and only
perform immunoassays to get the
correct answer to our specific scientific
questions. If this is the case for you,
what is your opinion? Shouldn’t all of
your assay results be accurate?
Sincerely yours Dr. Peter Rauch
Yes. We do think so. And that is the
reason why our team at CANDOR
works for reliable science. From the
very beginning – more than a decade
ago – we have focused on better tools
for better immunoassays. We believe
it is not acceptable, that assays do
not show true results – regardless
whether a matrix is challenging or
not. The great success of CANDOR
solutions in the last years was due
to so many lab technicians and
scientists, who only accept reliable
and convenient assays with true
results.
Sincerely yours Dr. Tobias Polifke
CANDOR solutions are “Made in
Germany” under very strict and ISO
certified quality control. CANDOR not
only supplies life science research
laboratories worldwide, but is also
trusted supplier for many industrial
kit manufacturers for human and
veterinary
diagnostics,
food
and
feed diagnostics and environmental
analysis. We have set new industrial
standards for assay reliability and
stability. With this brochure we would
like to showcase our current products
for professional users in modern
research laboratories. Enjoy reading!
3
Preface
3
Methods
4
Products
7
Optimizers
7
Blockers
11
Stabilizers
17
Buffer Solutions
27
Packages
32
ReadyTector®
34
METHODS
ELISA is the abbreviation for EnzymeLinked Immuno Sorbent Assay.
The Western blotting technique is
In immunohistochemistry or im-
Protein arrays are miniaturized assays
Immuno-PCR combines ELISA tech-
mixtures. First, the proteins are sep
structures are stained by using anti-
allel. They use proteins or antibodies
enhancement of a PCR (polymerase
used to detect proteins from crude
The principle is the binding of a la-
arated by gel-electrophoresis into
beled specific antibody to the target
bands. Then all proteins are trans
antigen (called analyte). The signal of
ferred to a membrane, where the
the enzyme labeled antibody is the
target protein band is detected with
conversion of a substrate, which can
primary antibodies specific to the
be detected and quantified.
target protein.
The most important assay formats
Such primary antibodies on the
are: Sandwich ELISA – Antibody Coat
membrane are detected with secon
ed, Sandwich ELISA – Antigen Coated
dary antibodies, which are either
and Competitive ELISA – Antibody
enzymatic or fluorescence labeled.
Coated.
munocytochemistry tissue or cell
bodies. A very good local resolution
can be achieved, if preparation of
tissue slices is optimized and block
ing of nonspecific binding is effec
tive. Thus not only presence, but
also localization of an analyte can be
visualized.
Detection is done commonly either
by enzymatic substrate-conversion
or by fluorescence-labeling.
4
for detection of many proteins in par
immobilized on surfaces such as glass
slides or beads. A single target protein
is detected either by localization of a
signal at a specific point in the array
or by differences in the specific beads,
which can be individually coded e.g.
nique with the exponential signal
chain reaction). The specific antibodies are labeled with a DNA marker.
By using the amplification steps of
the PCR, the sensitivity can be better compared to the “classic” ELISA.
by color or fluorescence.
Due to the fact that signals from
The major advantage of this technique
enhanced by the PCR-reaction, false
is that many analytes can be detected
at the same time from one sample. But
cross-talk between the multiplexed
assays has to be avoided to enable sufficient reliability of results.
nonspecific binding are significantly
positive results are very common. As
a consequence effective blocking of
the plate surface and avoiding of interference are extremely important
for Immuno-PCR.
5
OPTIMIZERS
Optimizers are a class of advanta
geous and multifunctional solutions
for immunoassays.
They are outstanding compared to
other diluents and buffers. LowCrossBuffer® is the first solution that
combines several functionalities of
diluents in one product and is not
only an assay diluent. Additionally it
LowCross®
HRP-Stab
is
another
The Blocking Solution shows out-
product with several functionalities.
standing
This product enables convenient
combined with a reduced back-
blocking
characteristics
one step incubation assays without
ground and a high sensitivity of the
negatively affecting the reliability of
immunoassay.
the results. This is due to both the
optimizing and stabilizing properties
Please find more information about
of the solution. Due to its stabilizing
The Blocking Solution in the chapter
properties, LowCross® HRP-Stab is
Blockers.
described in the chapter Stabilizers.
can reduce HAMA-interference and
furthermore helps to avoid matrix
The casein based surface blocker
effects.
“The Blocking Solution” is also part of
the class of optimizers.
7
ELISA
LOWCROSS-BUFFER
®
Western Blot
1.2
Antibody and sample diluent for
In addition matrix effects, coming for
cross-reactivities and matrix effects
specimen are reduced.
1.0
That means that matrix effects
0.8
nonspecific
binding,
in immunoassays.
example from blood sera or plasma
LowCross-
no longer exacerbate detection of
Buffer® helps to reduce interference
analytes. Minimization of all these
in
quality
negative effects upgrades the quality
of detection can be significantly
of the assay and improves the
improved. Nonspecific binding of
reliability of the results.
The
formulation
immunoassays.
of
Thus
B/B0
minimizing

1:750 LowCross-Buffer®
1:750 PBS/BSA Tween
0.6
0.4
the antibodies, negative effects of
0.2
interfering substances and low or
medium affinity cross-reactivities
0
0.01
of the antibodies are minimized.
0.1
1
10
100
1000
Analyte concentration [ng/mL]
10000
Calibration curve of an ELISA. The analyte was diluted 1:750 in PBS/BSA Tween
or 1:750 in LowCross-Buffer®. The ELISA with PBS/BSA Tween shows, despite
the high dilution factor, a very high coefficient of variation (error bar) due to
interference. The precision of the ELISA with LowCross-Buffer® as assay diluent
is significantly better.
8
A
B
C
D
E
F
G
H
A
B
C
D
E
F
G
H
1 2 3 4 5 6 7 8 9 10 11 12
Immunohistochemistry
false positives
Protein Array
Standard assay
diluent
Immuno-PCR
false positives
1 2 3 4 5 6 7 8 9 10 11 12
LowCrossBuffer®
Package sizes
Elimination of false positive binding in an ELISA
Control of specifity in (A1-12) and blanks (H1-H12)
show false positive binding.
(Data from Dr. C. Specht, vivo Science GmbH,
Gronau, Germany)
9
BLOCKERS
The surfaces of ELISA plates or
Without surface blocking, assay
CANDOR offers three blockers with
antibodies, tracers, analytes or other
different formulations and blocking
optimized for efficient attachment
components of the sample would
characteristics. BSA-Block is a classic
bind to the surface. This can lead
BSA-based blocker for many applica-
to false results or high background
tions. SmartBlock™ is a very efficient
signals and therefore it has to be

peptide based blocker. The Blocking
prevented in order to get reliable
Solutionis a premium casein based
assays. Blocking is necessary to
blocker for routine applications as well
saturate
as for challenging immunoassays.
Western blotting membranes are
of proteins and antibodies.
This enables optimal assays with a
high density of capture antibodies or
capture molecules on the surfaces. As
a consequence of this optimization
of the surfaces, many other proteins
or peptides and other molecules
from the sample can attach on the
surface, too.
free
binding
capacities
of the respective surfaces. This
saturation
can
be
achieved
by
covering the surface with a dense
layer of molecules without gaps in
this layer.
11
ELISA
THE BLOCKING SOLUTION
Western Blot
Casein based blocker for minimizing
nonspecific binding to surfaces.
Additionally, no components of the
sample can bind nonspecifically to
The Blocking Solution is based on
membranes or other protein binding
chemically modified and fragmented
surfaces and prevents nonspecific
highly purified casein.
Coefficient of variation [%]
The Blocking Solution saturates free
binding to surfaces.
The
production
process
of The
This minimizes unwanted binding of
Blocking Solution has been developed
the analyte or the detection antibody
and up-scaled by CANDOR. This
to the surface and so background
proprietary process guarantees an
can be avoided and sensitivity of the
extremely good lot-to-lot consistency
assay can be improved.
which can normally be obtained
although
the
casein
is
production of immunodiagnostics
by several international kit manu-
8
facturers.
6
The Blocking Solution is not com-
4
able or home-made casein based
2
dissolving and filtration of casein.
0
only from synthetic raw materials
originally from natural sources.
Protein Array
Immuno-PCR
parable to other commercially availsolutions which are produced by just
The Blocking Solution
BSA blocker
Decrease of the coefficient of variation. Compared to standard blockers, The
Blocking Solution impressively lowered the coefficient of variation as shown
in this assay validation of a competitive ELISA (n = 96, measured at the
maximum B0).
derived
used as a blocker in industrial scale
10
the surface, if it is effectively blocked.
binding sites on plastic surfaces,
The Blocking Solution is routinely
12
Package sizes
12
13
ELISA
SMARTBLOCK
TM
ELISA
BSA-BLOCK
Western Blot
BSA-free blocker for saturating free
SmartBlock™
other protein binding surfaces.
priced assays with good reliability
binding sites on plastic surfaces or
sets
e.g. for diagnostics, pharmaceutical
research,
based
biotechnological
improve
peptides.
the
blocking
To
further
efficiency,
peptides were chemically modified
in
food
pharmaceutical
process
diagnostics,
BSA-Block, we recommend using The
nonspecific binding even when using
Blocking Solution.
Protein Array
BSA-Block is a well-established, versatile blocker for many applications.
control,
environmental
If you experience background due to
Protein Array
or
diagnostics or research.
Standard blocker for saturating free
binding sites on plastic surfaces or
the development of economically
SmartBlock™ is a modern blocker
on
standards
Western Blot
Immuno-PCR
Immuno-PCR
The high purity and control of the
raw materials enable good lot-to-lot
in a specific process. The result is
a peptide solution with very good
For many assays SmartBlock™ is an
consistency for this blocker based
blocking characteristics. At the same
economical and useful alternative to
on BSA. Therefore a high reliability of
time SmartBlock™ has a very good
BSA containing solutions.
the assay over years is obtained even
when different lots are used.
lot-to-lot consistency, up until now
obtained only by synthetic blockers.
Package sizes
Package sizes
15
STABILIZERS
Immunoassay stabilizers are needed
CANDOR offers stabilizers to protect
CANDOR stabilizers can be used for
proteins, antibodies and conjugates
preserving activity of all biological
of proteins and to protect them from
in solution as well as stabilizers
components of an immunoassay.
to preserve the native conformation
losing their functionality.
for antibodies coated on surfaces.
The portfolio ranges from Antibody
CANDOR stabilizers are routinely
Stabilizers for immunoassays allow
Stabilizer over HRP-Protector™ and
used for production of commercial
long-term storage of antibodies,
LowCross® HRP-Stab to Liquid Plate
ELISA kits with shelf lives of 2 or more
conjugates (e.g. HRP-labeled or AP-
Sealer®.
years.
labeled) and antigens in solutions
as well as coated onto surfaces like
To always get the same reliable results
microtiter plates or others.
in immunoassays, it is essential to
preserve standards for calibration
curves, coated plates and antibodies
during storage.
17
ELISA
LOWCROSS HRP-STAB
®
Western Blot
Assay diluent for long-term storage
HRP-Stab gives peroxidase-antibody
120
nonspecific binding, cross-reactivities
stability over several years, even when
100
and matrix effects in immunoassays.
LowCross®
reliability
stands
by
for
superior
neutralizing
assay
conjugates the crucial long-term
stored as a ready-to-use solution.
The result is an assay diluent and
storage
buffer
for
peroxidase
HAMA-
conjugates in one solution with high
derived ones. The LowCross-effect
reliability and convenience due to
interferences
minimizes
including
matrix
effects, cross-
Conjugate activity [%]
of HRP conjugates and for minimizing
ready-to-use dilution.
reactivities and nonspecific binding.
day 0
day 104
80
Protein Array
day 251
60
day 378
40
20
Thus several washing steps like in
sequential assays are not necessary
0
and one-step-incubation protocols
without multiple washing can be
LowCross® HRP-Stab
Shelf life of an HRP-labeled antibody is measured as signal (= antibody
binding + enzymatic activity) in an ELISA after storage under heat stress at
45°C. The concentration of the stored antibody is 400 ng / mL.
done by using this assay diluent.
Standard assay diluent
Package sizes
18
19
ELISA
ANTIBODY STABILIZER
Western Blot
proteins or antibodies at 2 – 8°C.
Antibody
Stabilizer
is
used
for
120
storage of assay antibodies and for
as calibration standards or controls
structure of proteins and antibodies
in immunoassays.
100
storage of proteins, which are used
Antibody Stabilizer conserves the
Stabilizer for long-term storage of
preventing them from losing func
tionality due to storage.
Just dissolve proteins or antibodies
with Antibody Stabilizer and store in
the refrigerator.
day 0
day 104
80
Protein Array
day 251
60
day 378
Immuno-PCR
40
20
0
Antibody Stabilizer “TRIS”
Shelf life of an antibody is measured as binding in an ELISA after
storage under heat stress at 45°C. The concentration of the stored
antibody is 400 ng/mL.
20
Package sizes “TRIS”
Package sizes “PBS”
21
ELISA
LIQUID PLATE SEALER
®
Protein Array
Fig. 1: BSA-Block
Stabilizer for coated antibodies and
It allows long-term storage of coated
surfaces.
room temperature.
4
plates under dry conditions even at
After immobilization of the antibodies/
The recommended optimum storage
antigens and blocking, Liquid Plate
temperature is 2 – 8°C. But most
Sealer® seals the plates with a uniform
stabilized plates can also be stored
stabilizing layer, demonstrating good
dry at ambient temperature for many
solubility and without affecting the
months without loss in activity.
immunochromatographic test strips
(lateral flow assays), affinity chro
protein arrays. Plates treated with
matography columns, protein arrays
Liquid Plate Sealer® can be stored for
and for similar applications.
2
1.5
0
0.1
long periods after drying.
It is used as a coating stabilizer for
industrial ELISA kit production.
2.5
0.5
for stabilizing coated ELISA plates,
on polystyrene- or glass-surfaces for
day 84
1.0
Liquid Plate Sealer® can be used
antibodies and antigens after coating
day 0
3
assay afterwards.
It can also be used for stabilizing
Immuno-PCR
3.5
OD 450 nm
antigens on polystyrene- or glass-
Fig. 2: BSA-Block + Liquid Plate Sealer®
22
1
10
100
1000
Calibration curves of an ELISA are shown, which was either blocked only
with BSA (fig. 1) or blocked with BSA and additionally stabilized with Liquid
Plate Sealer® (fig. 2). Calibration curves at the starting day are identical. The
calibration curve of fig. 1 after storage shows that there is no remaining
binding activity of the capture antibodies.
0.1
1
10
100
The stabilized plate does not show significant decrease in
sensitivity after 84 days of incubation at 37°C.
1000
Package sizes
23
ELISA
HRP-PROTECTOR
TM
Western Blot
Diluent for long-term storage of HRP
conjugates.
Typical ready-to-use concentrations
120
for detection are between 40 – 100
500 ng/mL.
If you have background or inter
(HRP) coupled to antibodies or Neu-
ference – for example derived from
travidin/Streptavidin. HRP-Protector™
cross-reactivities – we recommend
gives peroxidase conjugates the
using LowCross® HRP-Stab as long-
crucial long-term stability over sev
term stabilizer for the conjugate.
HRP-Protector™ is a long-term stabilizer for horseradish peroxidase
eral years even when stored in low
concentrated ready-to-use dilutions.
day 0
day 104
80
Protein Array
day 251
60
day 378
40
20
HRP-Protector™ can be used directly
as an assay buffer for incubating the
0
HRP conjugates.
HRP-ProtectorTM
Shelf life of an HRP-labeled antibody is measured as signal (= antibody
binding + enzymatic activity) in an ELISA after storage under heat stress at
45°C. The concentration of the stored antibody is 400 ng/mL.
Package sizes
24
25
BUFFER SOLUTIONS
CANDOR Buffer Solutions comprise
the solutions. Therefore all products
centrated buffer solutions for easy
areaslike GLP laboratories (good
says.
kit production.
convenient ready-to-use or 10× con-
and fast working with immunoas-
can be used directly in regulated
laboratorypractice) or for diagnostics
All of these solutions are manufac-
The quality from lot-to-lot is always
tured in our own facility, which is
the same and conforms to highest
certified according to current DIN EN
requirements. In this chapter, buffers
ISO 9001 standards. This includes e.g.
such as Washing Buffer, Coating
inprocess-controls, batch testing and
Buffer, Sample Buffer and Stripping
traceability of every component of
Buffer are summarized.
27
ELISA
SAMPLE BUFFER
Western Blot
STRIPPING BUFFER
Western Blot
Sample and antibody dilution buffer
Sample Buffer is designed for trouble
Protein Array
Buffer for stripping antibodies from
membranes for multiple reprobing.
for immunoassays.
Stripping Buffer removes reaction
free assays, which are not affected by
solution and primary and secondary
interference derived from samples.
antibodies from Western blotting
Immuno-PCR
membranes.
If you have background or false
positives – for example derived from
After stripping, the membrane can be
cross-reactivities or matrix effects –
used for repeated detection (reprob
we recommend using LowCross-
ing) with antibodies.
Buffer® as the assay diluent.
Stripping Buffer does not contain
mercapto-ethanol nor DTT.
Package sizes
Package sizes
29
ELISA
COATING BUFFER 10×
ELISA
WASHING BUFFER 10×
Protein Array
Buffer for adsorptive immobilization of
testing the two Coating Buffers side
faces (for example microtiter plates)or
well as on proteins or antibodies the
Immuno-PCR
Western Blot
Washing steps are needed to remove
For immunohistochemistry (IHC) we
which are able to interfere with the
and detergents.
unbound and excessive components
proteins and antibodies on plasticsur-
by side. Depending on the surface as
required incubation time can vary.
Depending on the capture protein or
tion of the incubation procedure is
Washing Buffer is optionally available
recommended.
based on TRIS or PBS.
10× concentrate – simply dilute with
Washing Buffer is suitable for au-
water!
tomatic washers (Please note the
the capture antibody the pH of the
Coating Buffer is crucial for efficient
immobilization. Select from Coating
Buffer pH 7.4 and pH 9.6, as the pH-
assay.
Consequently, individual optimiza-
value can have an influence on the
specifications of your device and fol-
steric structure of proteins or anti-
low the instruction manual of your
bodies, thus affecting their immobilization.
For the optimization of the coating
procedure of a newly developed immunoasay we strongly recommend
washer!).
Package sizes
pH 7.4 10×
Package
sizes
500 mL order
number
120 500
pH 9.6 10×
125 mL
125
125 mL order
order number
number 120
121 125
order
number
120
500
offer Washing Buffer without Tween
Protein Array
10× concentrate – simply dilute with
water!
Immuno-PCR
Package sizes
PBS 10×
PBS IHC 10×
TRIS 10×
TRIS IHC 10×
31
ELISA
PACKAGE
CANDOR Starter Package
PACKAGE
CANDOR Starter Package IHC
ELISA
PACKAGE
CANDOR Blocking Sampler Package
Western Blot
Western Blot
Starter Package IHC small
Blocking Sampler Package small
50 mL LowCross-Buffer
50 mL The Blocking Solution
Starter Package small
50 mL SmartBlock™
125 mL Washing Buffer TRIS IHC 10×
50 mL BSA-Block
50 mL LowCross-Buffer®
Starter Package IHC large
Blocking Sampler Package large
125 mL Washing Buffer TRIS 10×
125 mL LowCross-Buffer
Starter Package large
125 mL Washing Buffer TRIS IHC 10×
For a first test, which solution is most
Protein Array
suitable for a specific assay, we offer
contains:
different sampler packages.
Immuno-PCR
contains:
contains:
contains:
Protein Array
Immuno-PCR
contains:
125 mL SmartBlock™
125 mL BSA-Block
contains:
125 mL LowCross-Buffer®
125 mL Washing Buffer TRIS 10×
Package sizes
Package sizes
Package sizes
small order number 600 001
large
large
large
33
READYTECTOR®
Easy, quick and clear
Western Blot
ReadyTector® is the all-in-one detection solution for Western blotting.
The next step is just washing using
Standard protocol
ReadyTector® protocol
the special ReadyTector® Wash buffer.
ReadyTector® contains everything
Despite the fast, all-in-one incuba-
you need for fast, all-in-one immuno
tion process, ReadyTector® reduces
detection. Only a specific primary
background, allowing users to gener
antibody has to be added. All-in-one
ate clear, distinct bands suitable for
means that one solution contains
publishing.
Comparable results obtained with standard
Western blotting protocol or ReadyTector®
protocol and ReadyTector® Chemiluminescent
Substrate
Lanes 1– 5 contain 22, 11, 5.5, 2.8, 1.4 ng Alpha-1Antitrypsin spiked in cell lysates, respectively.
Protein detection: Mouse anti-A1AT Clone 1AT
(Biotrend), 0.2 µg / mL on nitrocellulose.
everything you need, and the entire
process takes place in a single work
ReadyTector® for Western blotting –
step – blocking and binding of the
easy, quick and clear
primary and secondary antibody
occur simultaneously.
– easy – simple to use
– quick – very fast procedure with
very quick results
– clear – no background
ReadyTector® protocol
Washing
3 × 5 min
Incubation
ReadyTector® 15 min – 1 h
Washing
3 – 4 × 5 min
ReadyTector® saves several washing and incubation steps. Therefore
a time saving of several hours can be obtained.
Run your gel and blot and detect the membrane in a single morning.
You get the same great results with ReadyTector® – simply much faster.
Standard protocol
Washing
3 × 5 min
Please find more information at
www.readytector.com
Blocking
1 – 2 h
Incubation primary
antibody 1 – 2 h
Washing
3 × 5 min
Incubation secondary
antibody 1 – 2 h
Washing
3 × 5 min
35
08.2015 | www.designconnection.eu
CANDOR Bioscience GmbH
Simoniusstrasse 39
88239 Wangen, Germany
Phone +49 (0) 75 22 - 7952 70
Fax +49 (0) 7522 - 7952 729
[email protected]