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Activated Carrier Proteins: Adar's Maleimide-Activated OvA
/Product Detail : Cat. 1026-Bulk Activated Carrier Proteins: Adar's Maleimide-Activated OvA
Product name : Activated Carrier Proteins: Adar's Maleimide-Activated OvA
Catalog number : Cat. 1026-Bulk
Supplier name : Adar
Data sheet : Ask more or other datasheet now !
Activated KLH, BSA and Ovalbumin (OVA), are commonly used as protein
carriers for haptens such as peptides in order to enable the immune response to
small molecules. In many cases KLH conjugate is used for the immunization, while
BSA or OVA conjugate is used in immunoassys of the resulting antibodies. OVA
conjugates are used in order to filter out the antibodies directed to the KLH.
However in some cases OVA or other proteins are preffered as carrier proteins
because of a variety of other reasons.
The Maleimide activated KLH and OVA produced by Adar Biotech are
preactivated with a heterobifunctional cross-linker (GMBS). These activated
proteins may be reacted with biomolecules that contain a free sulfhydryl groups, to
form a stable thioether bond.
Maleimide-Activated OVA characteristics.
Activation method: GMBS.
Binding capacity: ~2-4 mg of peptide (average MW of 1000-2500) per 2 mg OVA
Protein concentration: 4 mg/ml (0.5 ml)
Storage buffer: PBS pH 7.5
Transportation conditions: Ambient temperature up to 7 days.
Storage condition: -20°C.
A. Procedure for Peptide Conjugation
1. Dissolve the sulfhydryl-containing hapten in a volume of water (or suitable
buffer) equals to x0.4 to x1 times the volume of OVA. For example dissolve 2 mg
of peptide in 200-500 μl of buffer for addition to 2 mg of activated OVA in 500 μl.
Note: For haptens with limited solubility, DMSO may be used for solubilization. Use .30%
DMSO in the final conjugation solution or the carrier protein may irreversibly denature.
Alkaline pH values (above 8.5) may hydrolyze the maleimide group or generate side reactions
with amines. Haptens must contain cysteine or a sulfhydryl group in the reduced state in order to
react efficiently with the maleimide group.
2. Thaw the Maleimide Activated OVA at room temperature.
Note: Do not vortex or heat the activated OVA.
3. Mix the peptide with activated OVA adding peptide stepwise (1/4th of quantity
each time in 5 minutes intervals) and watching for possible appearance of turbidity
in solution. If slight turbidity forms – stop adding peptide to KLH solution.
Reaction is done for 2 hours at room temperature under occasional tapping of tube.
4. Peptide-conjugated OVA can be purified by gel filtration or dialysis to remove
Note: If the immunogen is to be stored for more than a few days it is recommended to store
frozen at -20°C.
5. The coupling efficiency of conjugation can be determined by assaying the
content of free sulfhydryl groups in the unreacted peptide using DTNB reagent.
The activated-OVA should be stored frozen until use.
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