Human DPPIV/CD26 DuoSet ELISA Summary
* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.
Product Features
- Optimized capture and detection antibody pairings with recommended concentrations save lengthy development time
- Development protocols are provided to guide further assay optimization
- Assay can be customized to your specific needs
- Economical alternative to complete kits
Kit Content
- Capture Antibody
- Detection Antibody
- Recombinant Standard
- Streptavidin conjugated to horseradish-peroxidase (Streptavidin-HRP)
Other Reagents Required
PBS: (Catalog # DY006), or 137 mM NaCl, 2.7 mM KCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, pH 7.2 - 7.4, 0.2 µm filtered
Wash Buffer: (Catalog # WA126), or equivalent
Reagent Diluent: 1% BSA in PBS, pH 7.2-7.4, 0.2 m filtered (R&D Systems Catalog # DY995). Quality of BSA is critical (see Technical Hints).
Blocking Buffer: 1% BSA in PBS, pH 7.2-7.4, 0.2 m filtered (R&D Systems Catalog # DY995). Quality of BSA is critical (see Technical Hints).
Substrate Solution: 1:1 mixture of Color Reagent A (H2O2) and Color Reagent B (Tetramethylbenzidine) (Catalog # DY999)
Stop Solution: 2 N H2SO4 (Catalog # DY994)
Microplates: R&D Systems (Catalog # DY990), or equivalent
Plate Sealers: ELISA Plate Sealers (Catalog # DY992), or equivalent
Scientific Data
Product Datasheets
Preparation and Storage
Background: DPPIV/CD26
DPPIV/CD26 is a serine exopeptidase that is expressed as a noncovalent homodimer on the surface of epithelial cells, endothelial cells, and activated lymphocytes. It regulates multiple aspects of immune and endocrine function by cleaving Xaa-Pro or Xaa-Ala dipeptides from the N-terminus of a wide variety of chemokines (CCL4 and 5, CXCL6, 9, 10, 11, and 12), growth factors (GM-CSF, IL-3), and peptide hormones (Glucagon, Glucagon-like Peptides 1 and 2, GIP, GHRH, Procalcitonin, Neuropeptide Y, and Substance P). DPPIV interacts in cis with Adenosine Deaminase on T cells, in trans with Caveolin-1 on antigen presenting cells, and it serves as a cell entry coreceptor for HIV and coronavirus. It provides costimulatory proliferation and activation signals to both CD4+ and CD8+ T cells. A soluble form of DPPIV can be proteolytically shed from adipocytes, leading to insulin resistance in adipocytes and skeletal muscle.
Assay Procedure
GENERAL ELISA PROTOCOL
Plate Preparation
- Dilute the Capture Antibody (to the working concentration stated in the product datasheet ) in PBS without carrier protein. Immediately coat a 96-well microplate with 100 µL per well of the diluted Capture Antibody. Seal the plate and incubate overnight at room temperature.
- Aspirate each well and wash with Wash Buffer, repeating the process two times for a total of three washes. Wash by filling each well with Wash Buffer (400 µL) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each step is essential for good performance. After the last wash, remove any remaining Wash Buffer by aspirating or by inverting the plate and blotting it against clean paper towels.
- Block each well of the microplate as recommended in the product datasheet. Incubate at room temperature for a minimum of 1 hour.
Note: The recommended Reagent Diluent typically contains 1% BSA. Some DuoSet Development Kits require alternative blocking agents, or for plates to be blocked overnight with a higher percentage of BSA, please see the product datasheet for details.
- Repeat the aspiration/wash as in step 2. The plates are now ready for sample addition.
PRECAUTION
The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face and clothing protection when using this material.
Assay Procedure
- Add 100 µL of sample or standards in Reagent Diluent, or an appropriate diluent, per well. Cover with an adhesive strip and incubate 2 hours at room temperature.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- Add 100 µL of the Detection Antibody, diluted in Reagent Diluent (as recommended in the product datasheet), to each well. Cover with a new adhesive strip and incubate 2 hours at room temperature.
- Repeat the aspiration/wash as in step 2 of Plate Preparation.
- Add 100 µL of the working dilution of Streptavidin-HRP to each well. Cover the plate and incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
- Repeat the aspiration/wash as in step 2.
- Add 100 µL of Substrate Solution to each well. Incubate for 20 minutes at room temperature. Avoid placing the plate in direct light.
- Add 50 µL of Stop Solution to each well. Gently tap the plate to ensure thorough mixing.
- Determine the optical density of each well immediately, using a microplate reader set to 450 nm. If wavelength correction is available, set to 540 nm or 570 nm. If wavelength correction is not available, subtract readings at 540 nm or 570 nm from the readings at 450 nm. This subtraction will correct for optical imperfections in the plate. Readings made directly at 450 nm without correction may be higher and less accurate.
Citations for Human DPPIV/CD26 DuoSet ELISA
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
11
Citations: Showing 1 - 10
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Circulating Concentrations of Cathelicidin Anti-Microbial Peptide (CAMP) Are Increased during Oral Glucose Tolerance Test
Authors: Höpfinger, A;Karrasch, T;Schäffler, A;Schmid, A;
International journal of molecular sciences
Species: Human
Sample Types: Serum
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Distinctive CD26 Expression on CD4 T-Cell Subsets
Authors: OJ Cordero, C Rafael-Vid, R Varela-Cal, C Calviño-Sa, B Malvar-Fer, S García, JE Viñuela, JM Pego-Reigo
Biomolecules, 2021-10-02;11(10):.
Species: Human
Sample Types: Cell Culture Supernates
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Regulation of Fibroblast Activation Protein by Transforming Growth Factor Beta-1 in Glioblastoma Microenvironment
Authors: E Krepela, Z Vanickova, P Hrabal, M Zubal, B Chmielova, E Balaziova, P Vymola, I Matrasova, P Busek, A Sedo
International Journal of Molecular Sciences, 2021-01-21;22(3):.
Species: Human
Sample Types: Cell Lysates
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Investigation of CD26, a potential SARS-CoV-2 receptor, as a biomarker of age and pathology
Authors: AA Raha, S Chakrabort, J Henderson, E Mukaetova-, S Zaman, J Trowsdale, R Raha-Chowd
Biosci Rep, 2020-12-23;40(12):.
Species: Human
Sample Types: Plasma
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Plasma levels of DPP4 activity and sDPP4 are dissociated from inflammation in mice and humans
Authors: LL Baggio, EM Varin, JA Koehler, X Cao, Y Lokhnygina, SR Stevens, RR Holman, DJ Drucker
Nat Commun, 2020-07-28;11(1):3766.
Species: Human
Sample Types: Plasma
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The associations between some biological markers, obesity, and cardiovascular risk in Slovenian children and adolescents
Authors: NM Varda, M Medved, L Ojsteršek
BMC Pediatr, 2020-02-21;20(1):81.
Species: Human
Sample Types: Whole Blood
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CD26-Related Serum Biomarkers: sCD26 Protein, DPP4 Activity, and Anti-CD26 Isotype Levels in a Colorectal Cancer-Screening Context
Authors: L De Chiara, M Páez de la, J Rodríguez-, MC Alvarez-Pa, MC Pardiñas-A, R Varela-Cal, OJ Cordero
Dis. Markers, 2020-01-21;2020(0):4347936.
Species: Human
Sample Types: Cell Culture Supernates
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Systemic DPP4 activity is reduced during primary HIV-1 infection and is associated with intestinal RORC+ CD4+ cell levels: a surrogate marker candidate of HIV-induced intestinal damage
Authors: MJ Ploquin, A Casrouge, Y Madec, N Noël, B Jacquelin, N Huot, D Duffy, SP Jochems, L Micci, C Lécuroux, F Boufassa, T Booiman, T Garcia-Tel, M Ghislain, RL Grand, O Lambotte, N Kootstra, L Meyer, C Goujard, M Paiardini, ML Albert, M Müller-Tru
J Int AIDS Soc, 2018-07-01;21(7):e25144.
Species: Human
Sample Types: Plasma
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Heterogeneity of molecular forms of dipeptidyl peptidase-IV and fibroblast activation protein in human glioblastomas.
Authors: Matrasova I, Busek P, Balaziova E, Sedo A
Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub, 2017-04-26;161(3):252-260.
Species: Human
Sample Types: Tissue Homogenates
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Intraindividual changes of dipeptidyl peptidase-IV in peripheral blood of patients with rheumatoid arthritis are associated with the disease activity.
Authors: Sromova L, Busek P, Sedova L, Sedo A
BMC Musculoskelet Disord, 2015-09-09;16(0):244.
Species: Human
Sample Types: Plasma
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Shedding of dipeptidyl peptidase 4 is mediated by metalloproteases and up-regulated by hypoxia in human adipocytes and smooth muscle cells.
Authors: Rohrborn D, Eckel J, Sell H
FEBS Lett, 2014-09-12;588(21):3870-7.
Species: Human
Sample Types: Cell Culture Supernates
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