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Amplite® Fluorimetric Goat Anti-Rabbit IgG-HRP Conjugate ELISA Assay Kit
Red Fluorescence
Horseradish Peroxidase (HRP) is a small molecule (MW ~40 KD) that is widely used in a variety of biological detections. HRP conjugates are extensively used as secondary detection reagents in ELISAs, immuno-histochemical techniques, Northern, Southern and Western blot analyses. Due to its small size, it rarely causes steric hindrance problem with antibody/antigen complex formation. It is usually conjugated to an antibody in a 4:1 ratio. Additionally, HRP is inexpensive compared to other labeling enzymes. The major disadvantage associated with peroxidase is their low tolerance to many preservatives such as sodium azide that inactivates peroxidase activity even at low concentration. Our Amplite® Fluorimetric ELISA Assay Kit contains all the essential components including our fluorogenic Amplite® Red HRP substrate for ELISA detection. The kit provides an optimized assay protocol that is compatible with HTS liquid handling, as little as 10 pg of a Polyclonal antibody in the well of a microplate can be detected. Its signal can be easily read by either fluorescence microplate reader or absorbance microplate reader. It can be used for the assays that detect goat anti-rabbit IgG as the secondary detection agent.
Detection of rabbit total IgG using the Amplite® Fluorimetric ELISA Kit. Rabbit IgG was diluted into 1 µg/mL and made 1 to 3 serial dilutions in 0.2 M sodium bicarbonate buffer at pH 9.4. 100 µL/well serial dilutions were coated into a black 96-well plate at 4°C overnight, and blocked with 3% milk in PBS and 0.02% Tween 20 at 4°C overnight. The wells were washed, and assayed using the reagents. 1 to 6000 dilutions of goat anti-rabbit IgG-HRP conjugate were used. The reactions were incubated for 15 to 60 minutes and then measured for fluorescence at Ex/Em = 540/590 nm with Gemini fluorescence microplate reader (Molecular Devices).
Detection of rabbit total IgG using the Amplite® Fluorimetric ELISA Kit. Rabbit IgG was diluted into 1 µg/mL and made 1 to 3 serial dilutions in 0.2 M sodium bicarbonate buffer at pH 9.4. 100 µL/well serial dilutions were coated into a black 96-well plate at 4°C overnight, and blocked with 3% milk in PBS and 0.02% Tween 20 at 4°C overnight. The wells were washed, and assayed using the reagents. 1 to 6000 dilutions of goat anti-rabbit IgG-HRP conjugate were used. The reactions were incubated for 15 to 60 minutes and then measured for fluorescence at Ex/Em = 540/590 nm with Gemini fluorescence microplate reader (Molecular Devices).
protocol
Protocol
sds
SDS
COA
CatalogSize
Price
Quantity
115411000 Tests
Price
 
Spectral properties
Excitation (nm)571
Emission (nm)584
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
UNSPSC12171501
Instrument settings
Absorbance microplate reader
Absorbance576 ± 5 nm
Recommended plateClear bottom
Fluorescence microplate reader
Excitation540 nm
Emission590 nm
Cutoff570 nm
Recommended plateSolid black
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Page updated on October 16, 2025