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CBA097 InnoZyme™ DPP IV Activity Assay Kit

InnoZyme™ DPP IV Activity Assay Kit MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.

Synonyms: Dipeptidyl Peptidase IV Activity Assay Kit

CBA097
  
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Overview

Replacement Information

Key Spec Table

Detection Methods
Fluorometric
Description
Overview

This product has been discontinued.





The DPP IV Soluble Activity Assay is designed to measure purified human DPP IV activity and for screening DPP IV inhibitors.
Catalogue NumberCBA097
Brand Family Calbiochem®
SynonymsDipeptidyl Peptidase IV Activity Assay Kit
Application Data
Human recombinant DPP IV activity was measured as outlined in the Detailed Protocol above using a 1-h incubation at 37°C. The data is presented as Relative Fluorescence Units.
Materials Required but Not Delivered• Distilled H2O
• Pipettors or multi-channel pipettor carefully calibrated to the target volume
• 37°C incubator
• Fluorescent plate reader capable of measuring fluorescence at an excitation wavelength of 360 nm and an emission wavelength of 460 nm
References
ReferencesMcIntosh, C.H. 2008. Front. Biosci. 13, 1753.
Thompson, M.A., et al. 2007. Mini Rev Med Chem. 7, 253.
Demuth, H.U., et al. 2005. Biochim Biophys Acta. 1751, 33.
Gallwitz, B. 2005. Rev Diabetic Stud. 2, 61.
Pro, B., and Dang, N.H. 2004. Histol Histopathol. 19, 1345.
Durinx, C., et al. 2000. Eur J Biochem. 267, 5608.
Product Information
Detection methodFluorometric
Form96 Tests
Format96-well plate
Kit contains96-Well Plate, Human Recombinant DPP IV Control, Substrate, Assay Buffer, 10X Sample Buffer, Plate Sealer, and a user protocol.
Applications
Biological Information
Assay range0.31-20 ng/ml
Physicochemical Information
Dimensions
Materials Information
Toxicological Information
Safety Information according to GHS
Safety Information
R PhraseR: 36/37/38

Irritating to eyes, respiratory system and skin.
S PhraseS: 26-36/37/39-56

In case of contact with eyes, rinse immediately with plenty of water and seek medical advice.
Wear suitable protective clothing, gloves and eye/face protection.
Dispose of this material and its container at hazardous or special waste collection point.
Product Usage Statements
Intended useThe Calbiochem® InnoZyme™ DPP IV Soluble Activity Assay is designed for screening DPP IV inhibitors or for measuring DPP IV activity in purified samples.
Storage and Shipping Information
Ship Code Dry Ice Only
Toxicity Irritant
Storage -20°C
Storage ConditionsUpon arrival store the unopened kit at -20°C. All kit components, once opened, can be stored for up to 3 months under the following conditions:

Table 1: Storage Conditions


Note: Following initial use the Control should be dispensed into aliquots and stored at -20°C or -70°C. Avoid freeze/thaw cycles.
Do not freeze Ok to freeze
Packaging Information
Transport Information
Supplemental Information
Kit contains96-Well Plate, Human Recombinant DPP IV Control, Substrate, Assay Buffer, 10X Sample Buffer, Plate Sealer, and a user protocol.
Specifications
Global Trade Item Number
Catalogue Number GTIN
CBA097 0

Documentation

InnoZyme™ DPP IV Activity Assay Kit Certificates of Analysis

TitleLot Number
CBA097

References

Reference overview
McIntosh, C.H. 2008. Front. Biosci. 13, 1753.
Thompson, M.A., et al. 2007. Mini Rev Med Chem. 7, 253.
Demuth, H.U., et al. 2005. Biochim Biophys Acta. 1751, 33.
Gallwitz, B. 2005. Rev Diabetic Stud. 2, 61.
Pro, B., and Dang, N.H. 2004. Histol Histopathol. 19, 1345.
Durinx, C., et al. 2000. Eur J Biochem. 267, 5608.
User Protocol

Revision12-May-2008 JSW
SynonymsDipeptidyl Peptidase IV Activity Assay Kit
Form96 Tests
Format96-well plate
Detection methodFluorometric
StorageUpon arrival store the unopened kit at -20°C. All kit components, once opened, can be stored for up to 3 months under the following conditions:

Table 1: Storage Conditions


Note: Following initial use the Control should be dispensed into aliquots and stored at -20°C or -70°C. Avoid freeze/thaw cycles.
Intended useThe Calbiochem® InnoZyme™ DPP IV Soluble Activity Assay is designed for screening DPP IV inhibitors or for measuring DPP IV activity in purified samples.
BackgroundDipeptidyl peptidase IV (DPP IV), also known as CD26, gp110 DPP 4, or adenosine deaminase-binding protein (ADAbp), is a serine exopeptidase. Together with fibroblast activation protein α (FAPα), it belongs to the propyl oligopeptidase family, S9. DPP IV is structurally unrelated to dipeptidyl peptidase II (DPP II) and atractin, but possesses similar substrate specificity. DPP IV is a membrane-bound, homodimeric, type II protein with a molecular weight of 110-150 kDa per subunit. The enzyme is bound to the membrane by a 22-amino acid transmembrane sequence. In addition to its role as a protease, the DPP IV has a long extracellular domain (738 residues) that exhibits receptor properties and binds to the extracellular matrix (ECM) via collagen or fibronectin. In human, the DPP IV activity is ubiquitous in almost all organs and tissues, with the highest expression in kidney, small intestine and placenta; moderate expression in lung, spleen, and liver; and a low levels of expression in heart, pancreas, and brain. A soluble form of DPP IV that can be found in human serum has been characterized as a proteolytic derivative of the membrane-bound form of DPP IV. DPPIV exhibits specificity toward a vast array of physiological substrates including chemokines, cytokines, endomorphins, hormones, and the pancreatic polypeptide of the PACAP/glucagons peptide family. This broad specificity suggests a role for DPPIV in many vital physiological processes that regulate pain, the immune system, the cardiovascular system, and metabolism. Recently DPPIV was the subject of intense research in diabetes due to its role in regulating incretin hormones, GLP-1 and GIP, which have a pleiotropic effect on glucose metabolism. Both hormones are deactivated simultaneously by DPP IV, enabling a multivalent method of treatment for diabetes and glucose metabolism disorders via the inhibition of this protease. In addition, DPP IV is not regulated by limited proteolysis or endogenous inhibitors, therefore specific DPP IV inhibitors should possess a strong therapeutic potential in various pathological processes.
Principles of the assayThe Calbiochem® InnoZyme™ DPP IV Soluble Activity Assay uses a fluorogenic substrate, Gly-Pro AMC, to measure DPP IV activity. Cleavage of the peptide bond releases the free AMC group, resulting in an increase in fluorescence, which is measured using an excitation wavelength of 360 nm and an emission wavelength of 460 nm.
Materials providedThe kit provides all components required to perform the assay.

• 96-Well Plate (Kit Component No. JA7666-1EA): 1 plate, 96-wells, supplied as six 2x8 well strips
• Control, Human Recombinant DPP IV (Kit Component No. JA9436-30UL): 1 vial, 30 µl supplied at 1 µg/ml
• Substrate (Kit Component No. JA9437-120UL): 1 vial, 120 µl, supplied as 20 mM Gly-Pro AMC in DMSO
• Assay Buffer (Kit Component No. JA9438-20ML): 1 bottle, 20 ml
• Plate Sealer: 2 each
Materials Required but not provided• Distilled H2O
• Pipettors or multi-channel pipettor carefully calibrated to the target volume
• 37°C incubator
• Fluorescent plate reader capable of measuring fluorescence at an excitation wavelength of 360 nm and an emission wavelength of 460 nm
Reagent preparationWarm all reagents to room temperature (15-25°C) just prior to use. • Diluted Control: Dilute the Control, Human Recombinant DPP IV 25-50-fold with chilled Assay Buffer. Maintain the Diluted Control on ice at all times. Important: Following initial thaw the Control, Human Recombinant DPP IV should be dispensed into aliquots and stored at -20°C. Avoid freeze/thaw cycles. • 1X Substrate: Dilute the Substrate 1:50 with Assay Buffer. For example, to prepare enough 1X Substrate for the entire 96-well plate, add 100 µl Substrate to 4.90 ml Assay Buffer.
Detailed protocolIt is recommended that all samples and standards be assayed in duplicate.

1. Remove the desired number of strips from the 96-Well Plate, return the remaining strips to foil pouch, and close the re-sealable edge.
2. Add 50 µl Diluted Control or purified sample enzyme to designated wells. Prepare Blank wells by adding 50 µl Assay Buffer to designated wells.
3. Add 50 µl 1X Substrate to each well. Cover the plate with a Plate Sealer and incubate for 1 h at 37°C. Protect from light.
4. Read the fluorescence at an excitation wavelength of 360 nm and an emission wavelength of 460 nm.
CalculationsCorrect the fluorescence value of all Diluted Controls and samples by subtracting the value of the blank from each reading. Calculate the mean fluorescence value for each duplicate Diluted Control and sample. Plot the data as Relative Fluorescence Units (RFU) as show in the figure below.
Assay characteristics and examples

Figure 1: Activity of Recombinant Human DPP IV

Human recombinant DPP IV activity was measured as outlined in the Detailed Protocol above using a 1-h incubation at 37°C. The data is presented as Relative Fluorescence Units.
Assay Range0.31-20 ng/ml
Assay Range NotesThe assay was standarized against recombinant, human DPP IV expressed in S. frugiperda insect cells.
Registered TrademarksCalbiochem® is a registered trademark of EMD Chemicals, Inc.
InnoZyme™ and Interactive Pathways™ are trademarks of EMD Chemicals, Inc.