Invitrogen™ BACE1 (β-Secretase) FRET Assay Kit, red

BACE1 FRET Assay (ß-Secretase) Kit, Red

BACE1 (β-secretase) is a key enzyme involved in the production of Amyloid beta-peptides (Abeta) found in extracellular amyloid plaques of Alzheimers disease (AD). In some cases, early onset familial AD can be attributed to a Swedish mutation in the amyloid precursor protein (APP), dramatically enhancing the cleavage of this protein by BACE1. This and other genetic and pathological evidence has led to therapeutic approaches focusing on the inhibition of BACE1 and other APP-cleaving enzymes, such as gamma-secretase. The BACE1 fluorescence resonance energy transfer (FRET) Assay Kit provides a sensitive and efficient method for screening potential BACE1 inhibitors. This kit uses purified baculovirus-expressed BACE1 and a new red FRET peptide substrate based on the Swedish mutant.

The BACE1 FRET Assay Kit offers:

Red emission to reduce compound interference

Excitation at 545 nm and read emission at 585 nm

An easy-to-use, rapid, homogeneous assay that is performed in solution

Versatile formatting for high-throughput screening

Stable results—measurements can be made for up to 24 hours

The principle of the BACE1 FRET assay is as follows: The peptide substrate is synthesized with two fluorophores, a fluorescent donor (a rhodamine (Rh) derivative), and a proprietary quenching acceptor. The distance between these two groups has been selected so that upon light excitation, the donor (D) fluorescence energy is significantly quenched by the acceptor (A) through a quantum mechanical phenomenon known as resonance energy transfer (Figure 1). Upon cleavage by the protease, the fluorophore is separated from the quenching group, restoring the full fluorescence yield of the donor. Thus, a weakly fluorescent peptide substrate becomes highly fluorescent upon enzymatic cleavage; the increase in fluorescence is linearly related to the rate of proteolysis.

Source of BACE1 Enzyme:
A cDNA sequence encoding amino acids 1-460 of human BACE1, corresponding to the ectodomain, was expressed in recombinant, baculovirus-infected insect cells. Purified BACE1 exists as the proBACE1 form having an apparent molecular weight of 55 kDa and an N-terminal sequence of TQHGIRLPLR.

Content And Storage

Components of the BACE1 FRET Assay Kit are listed in Table 1. Sufficient reagent is supplied for 400 high-throughput kinetic or endpoint assays. The BACE1 Enzyme is also available separately. Store BACE1 Enzyme at -80°C. Store BACE1 Substrate and BACE1 Product Standard at -20°C. Avoid subjecting these components to repeated freeze-thaw cycles. The BACE1 Assay Buffer and BACE1 Stop Solution can be thawed upon receipt and stored at room temperature.

Assay

β-Secretase Assay

Product Type

FRET Assay Kit

Substrate Properties

Peptide-Based Substrate

Substrate Type

Beta-Secretase Substrate

For Use With (Application)

FRET

Quantity

400 x 40 μL Assays

Shipping Condition

Dry Ice

Target

Beta Secretase (BACE1)

For Research Use Only. Not for use in diagnostic procedures.