New 3-HRP conjugate
e-Proteins is developing a new cutting edge conjugate using HRP (HorseRadish peroxidase). The new conjugate, 3-RHP, enables detection and quantification.
There are 2 main challenges to achieve this high performance . The first one is the production of a the mix of HRP with only one biotin to avoid the HRP polymerisaiton. The second one is the selection of the conjugate with 3 HRP per binding protein (strepavidin, Neutralite avidin,avidin) . This specific ratio enable a more efficient detection (+ 50 %) but also a direct quantification of the antibody.
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To proceed such a conjugate , e-proteins requests from its supplier the highest quality of enzyme that is not avaible on the world market. The major suppliers are not able to supply the HRP as it must be to proceed the perfect mix 3 HRP/1 binding proteins. For this reason, e-Proteins has tested several purification method with its supplier in order to reach the level of required quality. After 4 R&D years , a new 3 steps purification protocol that enable the extraction from the plant and the purification of the enzyme has been implemented. Our next challenge is to improve the production yield in order to give the opportunity for each of our customers to move to this new conjugate without significant additionnal cost.
HRP can be used in ChemiLuminescent ImmunoAssays (CLIA) . This technology combines the advantages of chemiluminescence sensitivity with the specificity of an immunoreaction. In HRP converts a substrate into a chemiluminescent signal that is emitted as photons of light in the visible or near visible range. The amount of light produced correlates to the amount of target analyte present in the sample. Because the 3HRP is homogeneous , a direct quantification is possible.
Compared with the Elisa Method, the main advantage of CLIA is much more sensitive due to signal amplification and multiplication but it works without incubation time and without stopping reagents.
Horseradish peroxidase is one of the most important enzymes obtained from a plant source. It continues to attract the attention of researchers from a variety of disciplines because of its practical and commercial applications. Advances in understanding the structure and catalytic mechanism of horseradish peroxidase have been made using protein engineering and other techniques. The physiological role of the enzyme is now being investigated in the context of new information on the plant peroxidase gene family of Arabidopsis thaliana.