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Supplemental Product Information:

Product Information

Volume/Quantity : 0.2 mg x 3 vial
Conjugation : Peroxidase from Horseradish (HRP) was conjugated to NHS Ester.
Product Form : Horseradish Peroxidase NHS Ester Conjugate. Lyophilized in 10 mM sodium citrate, 15 mM NaCl, 50 mM sucrose, pH 6.0.
Preservative : None

Description

HRP NHS Ester Conjugate is designed to facilitate the conjugation of the HRP to proteins, peptides and ligands that contain an amino group (NH₂). It can be used to label antibodies with a simple mixing.

Storage

Store at −20 °C to −80 °C (Preferably at −80 °C). Reconstitute this reagent immediately before use.
Do not prepare stock solutions for storage. Discard any unused reconstituted reagent.

Protocol

1. Dissolve the amine compound to be conjugated at 2 - 10 mg/mL in a buffer.
   Note: Small molecules containing azide or amino groups such as sodium azide need to be removed. Tris buffer and other amine containing buffers will also interfere with the labeling reaction.
   We recommend using the amine free buffer (e.g PBS, Phosphate buffer, Borate buffer, Bicarbonate buffer) pH range 7 - 9.
   The amount of antibody used for labeling ideally should correspond to molar ratios between 1:4 and 1:1 antibody to HRP. This vial contains 0.2 mg of HRP NHS and can effectively modify 0.2 - 0.8 mg of antibody.
2. Equilibrate the vial to room temperature.
3. Reconstitute the contents of the vial with 20 - 100 μL of a buffer to obtain a 2 - 10 mg/mL solution of HRP NHS.
   Note: Do not vortex or heat the HRP NHS.
4. Immediately mix the amine-containing molecule and HRP NHS solutions and incubate for 2 hours at 37 °C.
   Note: Alternatively, the reaction may be initiated by adding amine-containing molecule solution to HRP-NHS vial.
5. To quench the reaction, add 200 μL of 100 mM Tris-HCl Buffer pH 7.5 and incubate for 1 hour at 37 °C.
6. If desired, desalt, dialyze or purify the conjugate.

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