SouthBayBio

Cullin4a/Rbx1/DDB1/CRBN E3 Ligase Complex TR-FRET Kit

CHF 1'250.00

In stock

SBB-KF0120-KI011 KitCHF 1'250.00

Figure 1: % Signal to Background of Continuous Real-Time TR-FRET Cullin4a/Rbx1/DDB1/CRBN titration (autoubiquitination): Serial dilutions of Cullin4a/Rbx1/ DDB1/CRBN from 75nM to 18.75nM mixed with UBA1, UBE2D1, and, TRF-Ub mix. Reaction was initiated with addition of Mg-ATP.

Figure 2: Estimated Z-primes of Continuous Real-Time TR-FRET Cullin4a/Rbx1/DDB1/CRBN titration (autoubiquitination): Serial dilutions of Cullin4a/Rbx1/ DDB1/CRBN from 75nm to 18.75nM mixed with UBA1, UBE2D1, and TRF-Ub mix. Reaction was initiated with addition of Mg-ATP.

Specifications / Handling

More Information
Product Details
Product Type	Kit
Properties
Application Set	Compound Screening
Specificity	Readout: Endpoint/Kinetic Label or Dye: Europium-Cryptate & Cy5 Detection Method: TR-FRET Substrate Properties: Protein-based Substrate. Typical experimental concentration is 1x. Excitation/Emission (nm): 304/620 & 304/665
Crossreactivity	Human
Quantity	Sufficient Reagents for: 400 x 30μl reactions
Kit Contains	100x UBA1 (150μl) 100x UBE2D1 (150μl) 100x Cul4a/Rbx1 Complex (neddyl.) (150μl) 100x DDB1/CRBN (150μl) 10x Reaction Buffer (1ml) 10x Mg-ATP (1ml) 1 x 384-well white low-volume microplate
Other Product Data	The kit uses ubiquitin labeled with either Europium-Cryptate or Cy5 as FRET pair donor and acceptor fluorophores respectively, completely eliminating the need for antibody based detection setups. Enzymatic incorporation of the labeled ubiquitins into chains conjugated onto CRBN (or substrates) leads to an increase in fluorescence emission at 665nm (Em-Acceptor) and decrease at emission wavelength 620nm (Em-Donor). Click here for a Typical Lot-specific Product Datasheet from the Original Manufacturer Our product description may differ slightly from the original manufacturers product datasheet.
Declaration	Manufactured by South Bay Bio.
Shipping and Handling
Shipping	DRY ICE
Short Term Storage	-20°C
Long Term Storage	-80°C
Handling Advice	Aliquot to avoid freeze/thaw cycles. Avoid freeze/thaw cycles.
Documents
Manual	No
MSDS	Inquire
Product Specification Sheet
Datasheet	Download PDF

Product-specific References

Description

South Bay Bio’s Cullin4a/Rbx1/DDB1/CRBN Neddylated E3 Ligase TR-FRET Kit provides a fast and sensitive method monitoring ubiquitin conjugation onto the E3 complex in solution, resulting from an enzymatic ubiquitin cascade without the need of running and staining an SDS gel. The kit enables continuous TR-FRET detection of ubiquitin chain formation onto CRBN (or substrates) in a real-time detection setup, or in an end-point configuration if desired. TR-FRET uses the extended fluorescence emission decay lifetimes typical of rare-earth lanthanides to impart a short time-delay between FRET donor excitation and emission. This delay provides a means to separate “true” signal from short-lived background fluorescence, and reduce interference from compound fluorescence and other assay artifacts.

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