PRODUCT AND SERVICES CATALOG. Labeling reagents. Cryptates, XL665, DNP Cryptate labeling kit PAGE 63

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1 Labeling reagents Cryptates, XL665, DNP Cryptate labeling kit 63

Labeling reagents HTRF fluorophores for do-it-yourself labeling. Description HTRF labeling reagents are a do-it-yourself solution for labeling s with HTRF fluorophores.

2 Labeling reagents HTRF fluorophores for do-it-yourself labeling. Description HTRF labeling reagents are a do-it-yourself solution for labeling s with HTRF fluorophores. For maximum labeling flexibility, multiple functionalizations are available for both the donor and the acceptor Learn more about training for HTRF fluorophores. labeling on page 74 Donor fluorophores: The Europium Trisbipyridine cryptate (Eu 3+ -TBP cryptate) is available with 3 different functional groups: K-NH 2 : Di-amino cryptate (Eu 3+ -TBP-NH 2 cryptate), for use in combination with bifunctional reagents such as carbodiimides and SMCC/SPDP, for a flexible labeling solution. K-NHS: N-hydroxysuccinimide activated cryptate (Eu 3+ -TBP-NHS cryptate), reacts with primary amines. This specific cryptate is available either by itself or in the cryptate labeling kit (see next section for more details). K-MP: Maleimide activated cryptate (Eu 3+ -TBP-MP cryptate), enabling specific labeling of thiol groups. Acceptor fluorophore The cross-linked allophycocyanin XL665 is available either with an amine group (XL665-NH 2 ) or a maleimide group (XL665-maleimide) for easier handling. Functionalized chemical tag DNP (2,4-dinitrophenyl) is a small organic tagging alternative to biotin. It can be detected by the anti-dnp antibody (labeled with cryptate or XL665, see page 54). The DNP-NHS labeling reagent enables conjugation of DNP by targeting primary aminogroups (N-terminus or lysine residues) under very mild conditions. Features Large variety of labeling reagents available for complete assay design (donor, acceptor, DNP) Long shelf life for labeling reagents Choice of functional groups for simplified labeling THE ADVANTAGES 64 A selection of HTRF fluorophores is available for experienced users familiar with labeling and chromatography. The availability of various functional groups on Eu 3+ cryptate and XL665 adds significant flexibility to labeling strategies and assay design.

3 Labeling reagents Use and definitions HTRF fluorophore labeling reagents are for use with standard protein chemistry protocols, such as SMCC/SPDP coupling. As shown beside, preactivation of both the label and the may be required. Recovery of labeled material can be achieved using conventional chromatographic methods, such as gel filtration or HPLC. The devices required for recovering labeled material are not delivered with these reagents. SPDP activation activated Eu 3+ cryptate DDT cryptate SMCC activation activated reduction cryptate maleimide Thiol Ordering information Eu 3+ -TBP-NH2 cryptate Eu 3+ -TBP-NHS cryptate Eu 3+ -TBP-MP cryptate XL665-NH2 XL665-Maleimide DNP-NHS Designation Label qty mg (nmoles) 0.1 (68) 1 (680) 0.1 (69) 1 (690) 0.1 (66) 1 (660) 1 (10) 10 (100) 1 (10) 10 (100) 0.2 (500) 1 (2,500) Target molecule* (mg) * Quantity of target molecule that can be labeled is based on a MW 150 kda (antibody) Cat# 65EU2ABA 65EU2ABB 65EUSABA 65EUSABB 65EUMABA 65EUMABB 65XL2ABB 65XL2ABC 65XLMABB 65XLMABC 65DNSABD 65DNSABB Frozen -20 C Dessicated -20 C Dessicated -20 C 4 C in saturated ammonium sulfate Frozen -80 C Dessicated 4 C coupling step conjugate purified cryptate conjugate Eu 3+ -TBP-NH2 cryptate labeling of a using the SPDP-SMCC method SELECTED BIBLIOGRAPHY This poster can be downloaded from Tokuda C, Jacquemart L, Preaudat M. Development of a Homogeneous Heparanase Assay using HTRF Technology. SBS 10th Annual Conference. September 2004, Orlando (USA). Ouled-Diaf J, Russello SV, Rourke K, Degorce F, Bunker C, Claret E. Use of Europium cryptate-labeled phospho-specific antibodies for high throughput kinase assays. Miptec. May 2003, Basel (CH). Préaudat M, Ouled-Diaf J, Alpha-Bazin B, Mathis G, Mitsugi T, Aono Y, Takahashi K, Takemoto H. A homogeneous caspase 3 activity assay using HTRF technology. J Biomol Screening. 2002; 7: Amoravain M, Ouled-Diaf J, Trinquet E, Gregor N, Bazin H, Blériot A, Degorce F, Seguin P, Mathis G. New cryptate Labeling Kit, a convenient way to label s for HTRF assays and High Throughput Screening. SBS 7th Annual Conference. September 2001, Baltimore (USA). Trinquet E, Bazin H, Bleriot A, Piccio V, Autiero H, Mathis G. New europium cryptates to probe molecular interactions using HTRF. SBS 6th Annual Conference. September 2000, Vancouver (Canada). Degorce F, Beaumont F, Cougouluègne F, Alpha-Bazin B, Bazin H, Seguin P, Mathis G. Use of 2,4-dinitrophenyl-N Hydroxysuccinimide ester(dnp-nhs) for protein labeling and its detection by anti-dnp HTRF generic reagents. SBS 5th Annual Conference. September 1999, Edinburgh (UK). 65

Labeling kit A quick and easy-to-use cryptate labeling method. Description The cryptate labeling kit is suitable for labeling peptides, proteins, and oligonucleotides, as well as small molecules.

4 Labeling kit A quick and easy-to-use cryptate labeling method. Description The cryptate labeling kit is suitable for labeling peptides, proteins, and oligonucleotides, as well as small molecules. N-hydroxysuccinimide activated cryptate (Eu 3+ -TBP-NHS cryptate) is supplied in the kit and reacts with primary amines for simplified conjugation. The kit includes all necessary materials for the labeling procedure (except dialysis and concentration devices): Eu 3+ cryptate-nhs Elution buffer Separation column This kit is designed for very small and fast labeling needs, such as determining the best antibody among several antibodies available for HTRF assay development. Eu 3+ -TBP- NHS cryptate is also available in bulk size (see Labeling Reagents section on page 73) for larger labelings. OUTSOURCE YOUR BIOMOLECULE LABELING You may outsource your labeling to Cisbio. See our service offer on page 73 Features Ready-to-use kit with all the necessary materials Labeling of molecules from MW 15 to 200 kda Labeling procedure takes less than 1 hour Primary amine targeting for simplified conjugation Typically provides 1,000 to 5,000 tests (20 µl format) No chromatographic equipment required THE ADVANTAGE A straightforward labeling procedure as a first easy step in assay development 66

5 Labeling kit Use and definitions The cryptate labeling kit includes all the necessary materials for rapid cryptate labeling of peptides, proteins, oligonucleotides, or small molecules containing amino-groups (N-terminus or lysine residues). The labeling procedure requires only a few steps and is performed under very mild conditions. A column and buffer for purifying the conjugate are also included in the kit, as well as a calibrator to define the ideal working dilution of the labeled material in HTRF assays. Depending on the molecular weight of the target molecule, each kit enables the labeling of 5 µg (protein-mw 15 kda) to 50 µg (antibody-mw 150 kda). add 50 µl of protein at 6.67 µm directly into the cryptate vial (i.e. 50 µg for an antibody) incubate 15 min at RT add the reaction mixture to the column collect the cryptate-conjugate fraction ( approx. 20 min) Ordering information Designation Label qty mg (nmoles) Target molecule* Cat# Cryptate labeling kit 0.007(5) 50 µg 62EUSPEA * Quantity of target molecule that can be labeled is based on a MW 150 kda (antibody) SELECTED BIBLIOGRAPHY This poster can be downloaded from Ouled-Diaf J, Russello SV, Rourke K, Degorce F, Bunker C, Claret E. Use of Europium cryptate-labeled phospho-specific antibodies for high throughput kinase assays. Miptec. May 2003, Basel (CH). Amoravain M, Ouled-Diaf J, Trinquet E, Gregor N, Bazin H, Blériot A, Degorce F, Seguin P, Mathis G. New cryptate Labeling Kit, a convenient way to label s for HTRF assays and High Throughput Screening. SBS 7th Annual Conference. September 2001, Baltimore (USA). 67

6 68 PRODUCT AND SERVICES CATALOG

Alkaline Phosphatase Labeling Kit-NH2

Alkaline Phosphatase Labeling Kit-NH2 Catalog Number KA0001 1 Kit Version: 02 Intended for research use only www.abnova.com Table of Contents Introduction... 3 Background... 3 Principle of the Assay...