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	48-602MAG	Buffer Detection Kit for Magnetic Beads	1 Kit

PK02 Raytide™ Substrate

Raytide™ Substrate: SDB (Sicherheitsdatenblätter), Analysenzertifikate und Qualitätszertifikate, Dossiers, Broschüren und andere verfügbare Dokumente.

Analysenzertifikat
Literatur
Broschüren
Data Sheet

PK02

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Übersicht

Replacement Information

Key Spec Table

Purity
>98% by SDS-PAGE

Description
Overview	Modified gastrin analog that serves as a general substrate for tyrosine kinases. Contains one phosphorylation site per molecule.
Catalogue Number	PK02
Brand Family	Calbiochem®

References
References	Guan, K.L., et al. 1991. Nature 350, 359.

Product Information
Form	Lyophilized
Formulation	Lyophilized in a volatile buffer, BSA.
Positive control	Protein-Tyrosine Kinase, (p60^c-src), Cat. No. PK03, and Raytide™, Cat. No. PK02, a substrate for tyrosine kinases, can serve as positive controls in experiments measuring protein phosphorylation.

Applications
Application Notes	Kinase Assay
Application Comments	Activity of p60^c-src can be measured as described below using Raytide™, other peptide substrates and many naturally occurring proteins. This preparation of p60^c-src has no detectable Ser or Thr kinase activity. Suggested Kinase Reaction Protocol Materials required: [γ-³²P]ATP (10 mCi/ml). Kinase Assay Buffer: 50 mM HEPES, pH 7.5 containing 0.1 mM EDTA and 0.015% Brij 35. Kinase Dilution Buffer: kinase assay buffer containing 0.1 mg/ml BSA and 0.2% β-mercaptoethanol (note: β-mercaptoethanol is required for src kinase activity). ATP Mix: 0.15 mM ATP, 30 mM MgCl₂ and 200 mCi/ml [γ-³²P]ATP in kinase assay buffer. Phosphoric acid Acetone P81 Ion Exchange Chromatography paper (i.e. Whatman No. 3698915). Procedure: 1. Dilute p60^c-src stock solution to 0.2 units/μl in Kinase Dilution Buffer. 2. Mix 10 μl diluted p60^c-src with 10 μl Raytide™ (1 mg/ml solution in Kinase Assay Buffer). 3. Start the reaction by adding 10 μl of ATP Mix. 4. Incubate at 30°C for 30 minutes. 5. Stop the reaction by adding 120 μl of 10% phosphoric acid. Vortex and apply 120 μl onto a square (2 cm x 2 cm) of P81 paper. 6. Wash papers extensively in 0.5% phosphoric acid. Wash once with acetone. Dry and quantitate by liquid scintillation counting. (Phosphorylated Raytide™ will stick, while ATP will wash off the papers). Notes: Each unit of p60^c-src enzyme catalyzes the incorporation of one pmol phosphate per minute from ATP into tyrosyl residues. In addition to phosphorylating Raytide™, the p60^c-src protein-tyrosine kinase can be used to phosphorylate many naturally occurring proteins. Natural substrates phosphorylated on tyrosyl residues may be purified from complex mixtures using the Immunoaffinity System for purification of phosphotyrosyl proteins (Cat. No. PTS01). In addition to being phosphorylated by p60^c-src, Raytide™ can be used as a substrate for many naturally occurring tyrosine kinases. Raytide™ contains BSA. Raytide™ control peptide (without a tyrosine, Cat. No. PK05) is also available. To use Raytide™ in a dephosphorylation assay with tyrosine phosphatases, see Nature 350, 359-62, 1991.

Applications

Application Notes

Kinase Assay

Application Comments

Activity of p60^c-src can be measured as described below using Raytide™, other peptide substrates and many naturally occurring proteins. This preparation of p60^c-src has no detectable Ser or Thr kinase activity. Suggested Kinase Reaction Protocol Materials required: [γ-³²P]ATP (10 mCi/ml). Kinase Assay Buffer: 50 mM HEPES, pH 7.5 containing 0.1 mM EDTA and 0.015% Brij 35. Kinase Dilution Buffer: kinase assay buffer containing 0.1 mg/ml BSA and 0.2% β-mercaptoethanol (note: β-mercaptoethanol is required for src kinase activity). ATP Mix: 0.15 mM ATP, 30 mM MgCl₂ and 200 mCi/ml [γ-³²P]ATP in kinase assay buffer. Phosphoric acid Acetone P81 Ion Exchange Chromatography paper (i.e. Whatman No. 3698915). Procedure: 1. Dilute p60^c-src stock solution to 0.2 units/μl in Kinase Dilution Buffer. 2. Mix 10 μl diluted p60^c-src with 10 μl Raytide™ (1 mg/ml solution in Kinase Assay Buffer). 3. Start the reaction by adding 10 μl of ATP Mix. 4. Incubate at 30°C for 30 minutes. 5. Stop the reaction by adding 120 μl of 10% phosphoric acid. Vortex and apply 120 μl onto a square (2 cm x 2 cm) of P81 paper. 6. Wash papers extensively in 0.5% phosphoric acid. Wash once with acetone. Dry and quantitate by liquid scintillation counting. (Phosphorylated Raytide™ will stick, while ATP will wash off the papers). Notes: Each unit of p60^c-src enzyme catalyzes the incorporation of one pmol phosphate per minute from ATP into tyrosyl residues. In addition to phosphorylating Raytide™, the p60^c-src protein-tyrosine kinase can be used to phosphorylate many naturally occurring proteins. Natural substrates phosphorylated on tyrosyl residues may be purified from complex mixtures using the Immunoaffinity System for purification of phosphotyrosyl proteins (Cat. No. PTS01). In addition to being phosphorylated by p60^c-src, Raytide™ can be used as a substrate for many naturally occurring tyrosine kinases. Raytide™ contains BSA. Raytide™ control peptide (without a tyrosine, Cat. No. PK05) is also available. To use Raytide™ in a dephosphorylation assay with tyrosine phosphatases, see Nature 350, 359-62, 1991.

Biological Information
Purity	>98% by SDS-PAGE

Physicochemical Information

Dimensions

Materials Information

Toxicological Information

Safety Information according to GHS

Safety Information

Product Usage Statements

Storage and Shipping Information
Ship Code	Blue Ice Only
Toxicity	Standard Handling
Storage	+2°C to +8°C
Avoid freeze/thaw	Avoid freeze/thaw
Do not freeze	Ok to freeze
Special Instructions	Following reconstitution, aliquot and freeze (-20°C) or refrigerate (4°C) with 0.1% azide. Stock solutions are stable for up to 3 months at -20°C. Avoid freeze/thaw cycles of solutions.

Packaging Information

Transport Information

Supplemental Information

Specifications

Global Trade Item Number
Bestellnummer	GTIN
PK02	0

Documentation

Raytide™ Substrate Analysenzertifikate

Titel	Chargennummer
PK02	Chargen-Nr.

Literatur

Übersicht
Guan, K.L., et al. 1991. Nature 350, 359.

Broschüre

Titel
Protein Phosphatases Technical Bulletin

Datenblatt

Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

Revision	18-September-2008 RFH
Application	Kinase Assay
Description	Raytide™ is a modified gastrin analog. There is one tyrosyl phosphorylation site per Raytide™ molecule. The carboxyl end of the peptide contains acidic and hydrophilic amino acid residues, while the amino terminus contains basic amino acids. In phosphoric acid the amino end is positively charged and binds to phosphocellulose paper (p81 Whatman). The distance between the two charged ends of the peptide prevents neutralization. Additionally, the peptide maintains an unfolded state and functions as an excellent substrate for protein-tyrosine kinases and tyrosine phosphatases (Nature 350, 359-62, 1991). Raytide™ EL, Cat. No. PK04, which has a higher labeling efficiency when used as either a kinase or a phosphatase substrate is also available. Note: the exact sequence of Raytide™ and Raytide™ EL is proprietary to Calbiochem and cannot be released.
Background	Raytide™ is a modified gastrin analog. There is one tyrosyl phosphorylation site per Raytide™ molecule. The carboxyl end of the peptide contains acidic and hydrophilic amino acid residues, while the amino terminus contains basic amino acids. In phosphoric acid the amino end is positively charged and binds to phosphocellulose paper (p81 Whatman). The distance between the two charged ends of the peptide prevents neutralization. Additionally, the peptide maintains an unfolded state and functions as an excellent substrate for protein-tyrosine kinases and tyrosine phosphatases (Nature 350, 359-62, 1991). Raytide™ EL, Cat. No. PK04, which has a higher labeling efficiency when used as either a kinase or a phosphatase substrate is also available. Note: the exact sequence of Raytide™ and Raytide™ EL is proprietary to Calbiochem and cannot be released.
Positive control	Protein-Tyrosine Kinase, (p60^c-src), Cat. No. PK03, and Raytide™, Cat. No. PK02, a substrate for tyrosine kinases, can serve as positive controls in experiments measuring protein phosphorylation.
Form	Lyophilized
Formulation	Lyophilized in a volatile buffer, BSA.
Recommended reaction conditions	Suggested Kinase Protocol Materials required: • [γ-³²P]ATP (10 mCi/ml). • Kinase Assay Buffer: 50 mM HEPES, pH 7.5 containing 0.1 mM EDTA and 0.015% Brij^®-35 detergent. • Kinase Dilution Buffer: kinase assay buffer containing 0.1 mg/ml BSA and 0.2% β-mercaptoethanol (note: β-mercaptoethanol is required for src kinase activity). • ATP Mix: 0.15 mM ATP, 30 mM MgCl₂ and 200 mCi/ml [γ-³²P]ATP in kinase assay buffer. • Phosphoric acid • Acetone • P81 Ion Exchange Chromatography paper (i.e. Whatman No. 3698915). Procedure: 1. Dilute p60^c-src stock solution to 0.2 units/µl in Kinase Dilution Buffer. 2. Mix 10 µl diluted p60^c-src with 10 µl Raytide™ (1 mg/ml solution in Kinase Assay Buffer). 3. Start the reaction by adding 10 µl of ATP Mix. 4. Incubate at 30°C for 30 min. 5. Stop the reaction by adding 120 µl of 10% phosphoric acid. Vortex and apply 120 µl onto a square (2 cm x 2 cm) of P81 paper. 6. Wash papers extensively in 0.5% phosphoric acid. Wash once with acetone. Dry and quantitate by liquid scintillation counting. (Phosphorylated Raytide™ will stick, while ATP will wash off the papers). Notes: In addition to being phosphorylated by p60^c-src, Raytide™ can be used as a substrate for many naturally occurring tyrosine kinases. Raytide™ control peptide (without a tyrosine, Cat. No. PK05) is also available. To use Raytide™ in a dephosphorylation assay with tyrosine phosphatases, see Nature 350, 359-62, 1991.
Purity	>98% by SDS-PAGE
Solubility	Aqueous Buffers (1 mg/ml)
Comments	Activity of p60^c-src can be measured as described below using Raytide™, other peptide substrates and many naturally occurring proteins. This preparation of p60^c-src has no detectable Ser or Thr kinase activity. Suggested Kinase Reaction Protocol Materials required: [γ-³²P]ATP (10 mCi/ml). Kinase Assay Buffer: 50 mM HEPES, pH 7.5 containing 0.1 mM EDTA and 0.015% Brij 35. Kinase Dilution Buffer: kinase assay buffer containing 0.1 mg/ml BSA and 0.2% β-mercaptoethanol (note: β-mercaptoethanol is required for src kinase activity). ATP Mix: 0.15 mM ATP, 30 mM MgCl₂ and 200 mCi/ml [γ-³²P]ATP in kinase assay buffer. Phosphoric acid Acetone P81 Ion Exchange Chromatography paper (i.e. Whatman No. 3698915). Procedure: 1. Dilute p60^c-src stock solution to 0.2 units/μl in Kinase Dilution Buffer. 2. Mix 10 μl diluted p60^c-src with 10 μl Raytide™ (1 mg/ml solution in Kinase Assay Buffer). 3. Start the reaction by adding 10 μl of ATP Mix. 4. Incubate at 30°C for 30 minutes. 5. Stop the reaction by adding 120 μl of 10% phosphoric acid. Vortex and apply 120 μl onto a square (2 cm x 2 cm) of P81 paper. 6. Wash papers extensively in 0.5% phosphoric acid. Wash once with acetone. Dry and quantitate by liquid scintillation counting. (Phosphorylated Raytide™ will stick, while ATP will wash off the papers). Notes: Each unit of p60^c-src enzyme catalyzes the incorporation of one pmol phosphate per minute from ATP into tyrosyl residues. In addition to phosphorylating Raytide™, the p60^c-src protein-tyrosine kinase can be used to phosphorylate many naturally occurring proteins. Natural substrates phosphorylated on tyrosyl residues may be purified from complex mixtures using the Immunoaffinity System for purification of phosphotyrosyl proteins (Cat. No. PTS01). In addition to being phosphorylated by p60^c-src, Raytide™ can be used as a substrate for many naturally occurring tyrosine kinases. Raytide™ contains BSA. Raytide™ control peptide (without a tyrosine, Cat. No. PK05) is also available. To use Raytide™ in a dephosphorylation assay with tyrosine phosphatases, see Nature 350, 359-62, 1991.
Storage	Avoid freeze/thaw +2°C to +8°C
Do Not Freeze	Ok to freeze
Special Instructions	Following reconstitution, aliquot and freeze (-20°C) or refrigerate (4°C) with 0.1% azide. Stock solutions are stable for up to 3 months at -20°C. Avoid freeze/thaw cycles of solutions.
Toxicity	Standard Handling
References	Guan, K.L., et al. 1991. Nature 350, 359.

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Beschreibung

PK04

Raytide™ EL Substrate

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