Millipore Sigma Vibrant Logo
Attention: We have moved. Merck Millipore products are no longer available for purchase on MerckMillipore.com.Learn More

CBA020 K-LISA™ Checkpoint Activity Kit

CBA020
  
Purchase on Sigma-Aldrich

Přehled

Tabulka spec. kláve

Detection Methods
Colorimetric
Description
Overview

This product has been discontinued.





The K-LISA™ Checkpoint Activity Kit is a rapid, sensitive, ELISA-based activity assay suitable for measuring the kinase activity of purified or partially purified Chk1 and Chk2 preparations, in vitro Chk1 and Chk2 inhibitor screening, and for assessing the regulation of Chk1 and Chk2 in cell signaling. The assay utilizes a biotinylated peptide substrate (KKKVSRSGLYRSPSMPENLNRPR) that is phosphorylated on the third serine by Chk1 and Chk2. Biotinylated Chk substrate and sample containing Chk1 or Chk2 are incubated in the presence of ATP in wells of the streptavidin-coated 96-well plate, which allows for phosphorylation and substrate capture in a single step.

• Highly versatile: can detect kinase activity in purified or partially purified Chk1 and Chk2 preparations.
• Highly sensitive: can detect kinase activity at picogram (Chk1) and nanogram levels (Chk2).
• Highly convenient: 96-well format, non-radioactive detection.
• Highly useful: enables high throughput screening of kinase inhibitors.

Catalogue NumberCBA020
Brand Family Calbiochem®
SynonymsK-LISA™ Chk1/Chk2 Activity Kit
Application Data
The activity of HisTag®, Human, Recombinant Chk1 (Cat. No. 220479) (17-2200 pg) was determined using protocol A as indicated in the Detailed Protocol section. Assay range: 34 pg-1100 pg (740 Units/mg).

The activity of Human, Recombinant Chk2 (0.06-60 ng) was determined using protocol A in the Detailed Protocol section. Assay range: 200 pg-20 ng (1283 Units/mg).
Materials Required but Not Delivered Chk1 or Chk2-containing sample
Anti-Chk1, Human (Sheep) (for immunoprecipitating Chk1) Cat. No. PC423
Anti-Chk2, Human (Rabbit) (for immunoprecipitating Chk2) Cat. No. DR1036
Protein A Agarose Suspension (Cat. No. IP02), Protein G-Plus Agarose Suspension (Cat. No. IP04), or Protein A/Protein G-Plus Agarose Suspension (Cat. No. IP05) (for immunoprecipitation of Chk1 and Chk2)
1X PBS
RIPA Buffer (Tris-HCl: 50 mM, pH 7.4, NP-40: 1%, Na-deoxycholate: 0.25% NaCl: 150 mM, EDTA: 1 mM)
PhosphoSafe™ Extraction Buffer (Cat. No. 71296) or equivalent for preparation of cell lysates
Wash bottle or multichannel dispenser for washing
Spectrophotometer capable of measuring absorbance in 96-well plates at 450nm, preferably in the dual wavelength mode at 450nm against a reference wavelength of 540 or 595 nm.
References
ReferencesBartek, J., and Lukas, J. 2003. Cancer Cell 3, 421.
O'Neill T., et al. 2002. J Biol. Chem. 277, 16102.
Jackson, J.R., et al. 2000. Cancer Res. 60, 566.
Liu, Q., et al. 2000. Genes Dev. 14, 1448.
Product Information
Detection methodColorimetric
Form96 Tests
Format96-well plate
Kit containsBiotinylated Chk1 Substrate, Phosphoserine Detector Antibody, Streptavidin Coated 96-Well Plate Strips, HRP-Conjugated Secondary Antibody, Serine/Threonine Kinase Assay Buffer, ATP/MgCl₂, Chk1 Inhibitor, Wash Buffer, TMB Substrate, Stop Solution, Plate Sealers, and a user protocol. Purified Chk1 is available separately (Cat. No. 220479).
Biological Information
Assay range25-725 µU (Chk1) and 260 µU - 26 mU (Chk2)
Assay time3 h
Sample TypeCell lysates, tissue extracts, purified enzymes
Physicochemical Information
Sensitivity25 pg (0.018 mU) purified Chk1
Product Usage Statements
Intended useThe K-LISA™ Checkpoint Activity Kit is suitable for measuring the kinase activity of purified or partially purified Chk1 and Chk2 preparations, in vitro Chk1 and Chk2 inhibitor screening, and for assessing the regulation of Chk1 and Chk2 in cell signaling.
Storage and Shipping Information
Ship Code Dry Ice Only
Toxicity Multiple Toxicity Values, refer to MSDS
Hazardous Materials Attention: Due to the nature of the Hazardous Materials in this shipment, additional shipping charges may be applied to your order. Certain sizes may be exempt from the additional hazardous materials shipping charges. Please contact your local sales office for more information regarding these charges.
Storage -20°C
Storage ConditionsUpon arrival store the Streptavidin-Coated 96-Well Plate at 4°C and the remaining components of the kit at -20°C.
Avoid freeze/thaw Avoid freeze/thaw
Do not freeze Ok to freeze
Supplemental Information
Kit containsBiotinylated Chk1 Substrate, Phosphoserine Detector Antibody, Streptavidin Coated 96-Well Plate Strips, HRP-Conjugated Secondary Antibody, Serine/Threonine Kinase Assay Buffer, ATP/MgCl₂, Chk1 Inhibitor, Wash Buffer, TMB Substrate, Stop Solution, Plate Sealers, and a user protocol. Purified Chk1 is available separately (Cat. No. 220479).
Global Trade Item Number
Katalogové číslo GTIN
CBA020 0