100X RELAY^TR Europium/Cy5 Ubiquitin Mix

Time resolved (TR)-FRET offers a sensitive, low background, stable and homogenous assay for determination of protein ubiquitination. Europium-ubiquitin and Cy5-ubiquitin are a pair of modified ubiquitin suitable for TR-FRET assays to assess formation of polyubiquitin chains, thereby reporting the activity of E2 and E3 enzymes in the reaction. The concentration and molar ratio of europium-ubiquitin and Cy5-ubiquitin in the RELAYTR Europium/Cy5 Ubiquitin Mix are optimized to achieve excellent signal-to-background ratio in ubiquitination reactions. Optimization of protein concentrations is often necessary to achieve a desirable signal-to-background ratio. Typical protein concentration ranges are: UbE1 (10-30 nM), E2 (25-250 nM) and E3 (10-250 nM).

Reaction time is usually 1-3 hours in kinetic or end point assay.

Additional Information

Product Name:

Also Known As:

Eu-Ub; Eu-Ubiquitin; Cy5-Ub, Cy5-Ubiquitin

Catalog No.:

T3501-2K

Size:

2,000 x 20 µl Reactions

Molecular Weight:

Eu-Ub (9.5 kDa); Cy5-Ub (9.1 kDa)

Species:

Human

Source:

Bacterial recombinant

Stock:

Concentration:

See tube label

Quality Assurance:

~ 95% by SDS-PAGE; Validated by RELAY^TR ubiquitination assays.

Storage:

Store at -80°C; avoid multiple freeze-thaw cycles

PDF Data Sheet:

PDF Datasheet

NCBI RefSeq:

Image(s):

Images:
(Click image to enlarge)

A. Kinetic mode monitoring 25 nM CHIP autoubiquitination. RELAY^TR Europium/Cy5 Ubiquitin Mix (Catalog# T3501) was used in all reactions.
B. CHIP concentration-dependent autoubiquitination monitored by RELAY^TR FRET assays with Eu-Ub and Cy5-Ub. S655/S620 ratios from reactions with ATP were positive Signal (S), and without ATP were Backgrounds (B). The signal-to-background ratio was calculated by using the formula of (S-B)/B.
C. Itch concentration-dependent autoubiquitination, similar to B.
D. Nedd4 concentration-dependent autoubiquitination, similar to B..

Shipping Method:

Dry ice shipping

References:

1. https://www.bmglabtech.com/en/tr-fret/

Details

Time resolved (TR)-FRET offers a sensitive, low background, stable and homogenous assay for determination of protein ubiquitination. Europium-ubiquitin and Cy5-ubiquitin are a pair of modified ubiquitin suitable for TR-FRET assays to assess formation of polyubiquitin chains, thereby reporting the activity of E2 and E3 enzymes in the reaction. The concentration and molar ratio of europium-ubiquitin and Cy5-ubiquitin in the RELAYTR Europium/Cy5 Ubiquitin Mix are optimized to achieve excellent signal-to-background ratio in ubiquitination reactions. Optimization of protein concentrations is often necessary to achieve a desirable signal-to-background ratio. Typical protein concentration ranges are: UbE1 (10-30 nM), E2 (25-250 nM) and E3 (10-250 nM).
Reaction time is usually 1-3 hours in kinetic or end point assay.

100X RELAYTR Europium/Cy5 Ubiquitin Mix

Additional Information

Details

100X RELAY^TR Europium/Cy5 Ubiquitin Mix