For research use only. Not for use in diagnostic procedures.
Cleavable ICAT® Reagents are a cysteine-specific, protein-based labeling strategy designed to compare two different sample states. These reagents provide you with more complete protein identification and quantification data than is possible with 2-D gels.
Incorporates carbon-13, rather than deuterium, into the heavy reagent, resulting in the co-elution of protein pairs labled with ICAT® Reagents from the reversed-phase column for improved peptide quantification.
Incorporates an acid-cleavable site in the reagent, allowing for removal of the biotin portion of the ICAT® Reagent tag prior to MS and MS/MS analysis.
Reduces the overall tag size enabling the analysis of larger peptides.
Reduces tag fragmentation, thereby improving MS/MS data quality and significantly increasing the number of identified proteins for higher confidence per experiment.
Improved reagent design reduces formation of ICAT/TCEP reaction by-products for overall MS data quality improvement.
The mass difference (9 dalton) between the heavy and light reagents avoids possible confusion between oxidized methionine and two cysteines labeled with ICAT® Reagents in LC/ESI/MS workflows.
ICAT® is a registered trademark of the University of Washington and is exclusively licensed to AB Sciex Pte. Ltd.
Increase proteome coverage: By labeling protein cysteinyl groups with the cleavable ICAT® Reagent, you can increase proteome coverage, identify and quantify proteins of interest, and accelerate your discovery efforts. The specific, cleavable label enables quantitation and identification of medium- to low-abundance proteins and reduces sample complexity. Incorporating the mass difference through tagging enables true, expression-dependent experiments, where quantitation is achieved in MS mode, and only those proteins that exhibit mass changes are further analyzed via MS/MS for identification.
Improved performance: Cleavable ICAT® Reagents incorporate a number of key technological advances that significantly improve protein-expression analysis. Incorporation of an acid-cleavable linker into the ICAT® Reagent molecule allows removal of the biotin affinity tag before MS and MS/MS analysis. This improves MS/MS performance and significantly increases the number of proteins identified and quantified, so that higher confidence scores can be provided in a single experiment. Incorporation of carbon-13 into the heavy ICAT® Reagent molecule promotes co-elution of protein pairs labeled with heavy and light isotope-coded tags in reversed-phase chromatography, thereby increasing accuracy of quantification measurements by LC/ESI/MS, LC/MALDI-TOF-MS, and MALDI-TOF-MS. These significant improvements in the reagent design, along with the availability of ICAT® Software for data analysis, enable researchers to fully exploit the potential of this technology in protein-expression profiling studies.
ICAT® Reagent composition: The cleavable ICAT® Reagent is composed of four moities: 1) a protein reactive group (iodocetamide) that covalently links the isotope-coded affinity tag to the protein through cysteine alkylation; 2) a biotin affinity tag that simplifies labeled-peptide analysis through affinity chromatography, specifically selecting for cysteine-containing peptides; 3) either a C13-labeled linker chain (heavy) or non-labeled chain (light, 9 dalton mass difference) that allows for mass spectrometric comparison of heavy- and light-labled peptides and provides a ratio of the concentration of the proteins in the original sample; and 4) an acid cleavage site allows for the removal of the biotin label following affinity purification of the labeled peptides to reduce the overall tag mass for improvement in peptide fragmentation efficiency.
The cleavable ICAT® Reagents are available through the Methods Development Kit, 10-Assay Kit formats, or as bulk reagents.
ICAT Starter Kit - Hardware ICAT Starter Kit contains hardware required for operation of Cation exchange and Avidin affinity cartridges used in the ICAT reagent kit protocol, and operation of Cation exchange cartridges for the iTRAQ reagent kit protocol. Includes Needleport Adapter (4326689), Outlet Connector (4326690), and Cartridge Holder (4326688).
Cartridge Holder Cartridge holder to operate cation exchange cartridges and avidin affinity cartridges.
Needleport Adaptor Used to manually operate the cation exchange or avidin affinity cartridges included in the ICAT Reagents kit. Used with: ICAT Starter Kit.
Outlet Connector Outlet tubing used for manual operation of cation exchange or avidin affinity cartridges used in the ICAT Reagent kit protocol. Used with: ICAT Cation Exchange Cartridge (5 Pack), ICAT Avidin Affinity Cartridge (5 Pack), ICAT Cartridge Holder.
Avidin (5 pack) Package of five 0.2-ml avidin affinity cartridges used in ICAT Kit protocol. Cartridges are operated with cartridge holder (sold separately).
Cation Exchange (5 pack) Package of five 0.2-ml cation exchange cartridges used in ICAT Kit protocol. Cartridges are operated with cartridge holder (sold separately).
ICAT Avidin Affinity Buffer Pack Contains individual 100 ml bottles of loading buffer, wash#1 buffer, wash #2 buffer, elution buffer and storage buffer. The buffer pack also includes one 0.2-ml avidin affinity cartridge.
ICAT Cation Exchange Buffer Pack Contains individual 100 ml bottles of loading buffer, elution buffer, cleaning buffer and storage buffer. The buffer pack also includes one 0.2-ml cation exchange cartridge.
Cleaving Reagent - A Contains 10 ml of reagent which is enough to perform approximately 100 x 1 unit assays.
Cleavable ICAT Reagent Methods Development Kit Contains everything required to start protein expression analysis studies using the new Cleavable ICAT reagent technology. The Methods Development kit contains 3 units of heavy reagent and 6 units of light reagent. Each unit of reagent will label approximately 100ug of protein. Included: ICAT Starter Kit - Hardware, Cation Exchange Buffer Pack, Affinity Buffer Pack, ICAT reagents, and protocol.
Cleavable ICAT Reagent – 10-Assay Kit Cleavable ICAT Reagent 10-Assay Kit contains all of the necessary reagents, buffers, cartridges, and instructions to start protein expression analysis studies using the new, cleavable ICAT Reagent technology. The 10-assay kit contains 10 units of heavy reagent and 11 units of light reagent. Each unit of reagent will label approximately 100 μg of protein.
Cleavable ICAT Reagent Bulk Kit (10 Units) Contains 10 units of both heavy and light reagent along with cleaving reagent A and operating instructions. Each unit of reagent will label approximately 100 μg of protein.
Cleavable ICAT Reagent Bulk Kit (100 Units) Contains 100 units of both heavy and light reagent along with cleaving reagent A and operating instructions. Each unit of reagent will label approximately 100 μg of protein.
Cleaving Reagent - B Contains 500 μl of cleaving reagent, enough to perform 100 x 1 unit assays.
Trypsin, TPCK-treated, 10-Pack A modified (TPCK treated) serine protease that specifically hydrolyzes peptide bonds at the carboxyl side of lysine and arginine residues. Contains calcium chloride which may improve consistency in digestion. Modified trypsin is treated with N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) to inactivate extraneous chymotryptic activity. Each package contains 10 vials containing 25 μg in each vial, sufficient for 12 duplex when using 100 ug of sample.
Trypsin, TPCK-treated, 8-Pack A modified (TPCK treated) serine protease that specifically hydrolyzes peptide bonds at the carboxyl side of lysine and arginine residues. Modified trypsin is treated with N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) to inactivate extraneous chymotryptic activity. Each package contains 8 vials containing 25 μg in each vial, sufficient for 10 duplex . Note: Does not contain calcium chloride.