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NPY Kit ELISA

NPY Reactivité: Humain, Souris, Rat Colorimetric Competition ELISA 0.1-1.000 ng/mL Cell Culture Supernatant, Plasma, Serum
N° du produit ABIN1979113
  • Antigène Voir toutes NPY Kits ELISA
    NPY (Neuropeptide Y (NPY))
    Reactivité
    • 7
    • 7
    • 5
    • 5
    • 3
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
    • 1
    Humain, Souris, Rat
    Méthode de détection
    Colorimetric
    Type de méthode
    Competition ELISA
    Gamme de detection
    0.1-1.000 ng/mL
    Seuil minimal de détection
    0.1 ng/mL
    Application
    ELISA
    Fonction
    Human/Mouse/Rat Neuropeptide Y EIA Kit optimized for serum, plasma and cell culture supernatants. Competition-based ELISA on a 96-well strip plate.
    Type d'échantillon
    Plasma, Serum, Cell Culture Supernatant
    Analytical Method
    Quantitative
    Specificité
    This EIA kit is designed to detect human, mouse, and rat active NPY (1-36).
    Réactivité croisée (Details)
    This ELISA kit shows no cross-reactivity with any of the cytokines tested: Ghrelin, Nesfatin, Angiotensin II and APC. This kit was designed to recognize the C-terminus of NPY and therefore will detect full-length NPY. However, it does not recognize active forms including NPY 1-36, 2-36, 3-36 or 3-35.
    Sensibilité
    0.2 ng/mL
    Attributs du produit
    • Strip plates and additional reagents allow for use in multiple experiments
    • Quantitative protein detection
    • Establishes normal range
    • The best products for confirmation of antibody array data
    Ingrédients
    • Pre-Coated 96-well Strip Microplate
    • Wash Buffer
    • Standard Peptide
    • Assay Diluent(s)
    • Biotinylated Peptide
    • HRP-Streptavidin
    • TMB One-Step Substrate
    • Stop Solution
    • Assay Diagram
    • Positive Control Sample
    • Capture Antibody
    • User Manual
    Matériel non inclus
    • Distilled or deionized water
    • Precision pipettes to deliver 2 μL to 1 mL volumes
    • Adjustable 1-25 mL pipettes for reagent preparation
    • 100 mL and 1 liter graduated cylinders
    • Tubes to prepare standard and sample dilutions
    • Orbital shaker
    • Aluminum foil
    • Saran Wrap
    • Absorbent paper
    • Microplate reader capable of measuring absorbance at 450nm
    • SigmaPlot software (or other software that can perform four-parameter logistic regression models)
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  • Indications d'application
    Recommended Dilution for serum and plasma samplesHuman: 2X / Mouse: 2X / Rat: 8X
    Volume d'échantillon
    100 μL
    Durée du test
    5 h
    Plaque
    Pre-coated
    Protocole
    1. Prepare all reagents, samples and standards as instructed.
    2. Add 100 μL detection antibody to each well.
    3. Incubate 1.5 h at RT or O/N at 4 °C.
    4. Add 100 μL standard or sample to each well.
    5. Incubate 2.5 h at RT.
    6. Add 100 μL prepared streptavidin solution.
    7. Incubate 45 min at RT.
    8. Add 100 μL TMB One-Step Substrate Reagent to each well.
    9. Incubate 30 min at RT.
    10. Add 50 μL Stop Solution to each well.
    11. Read plate at 450 nm immediately.
    Préparation des réactifs
    1. Keep kit reagents on ice during steps. Equilibrate plate to room temperature before opening the sealed pouch.
      2. Briefly centrifuge the NPY Antibody vial (Item N) and reconstitute with 5 µL of ddH2O before use. Add 50 µL of 1x Assay Diluent E into the vial to prepare a detection antibody concentrate. Pipette up and down to mix gently.
      3. The antibody concentrate should then be diluted 100-fold with 1x Assay Diluent E. This is your anti-Neuropeptide Y antibody working solution, which will be used in step 2 of the Assay Procedure. NOTE: the following steps may be done during the antibody incubation procedure (step 2 of Assay Procedure).
      4. Briefly centrifuge the vial of biotinylated Neuropeptide Y peptide (Item F) and reconstitute with 20 µL of ddH2O before use. Add 10 µL of Item F to 5 m 1X Assay Diluent E. Pipette up and down to mix gently. The final concentration of biotinylated Neuropeptide Y will be 20 ng/mL. This solution will only be used as the diluent in step 5 of Reagent Preparation.
      5. Preparation of Standards: Label 6 microtubes with the following concentrations: 1000 ng/mL, 100 ng/mL, 10 ng/mL, 1 ng/mL, 100 pg/mL and 0 pg/mL. Pipette 450 µL of biotinylated Neuropeptide Y solution into each tube, except for the 1000 ng/mL (leave this one empty). It is very important to make sure the concentration of biotinylated Neuropeptide Y is 20 ng/mL in all standards. a. Briefly centrifuge the vial of standard Neuropeptide Y peptide (Item C) and reconstitute with 10 µL of ddH2O. In the tube labeled 1000 ng/mL, pipette 8 µL of Item C and 792 µL of 20 ng/mL biotinylated Neuropeptide Y solution (prepared in step 4 above). This is your Neuropeptide Y stock solution (1000 ng/mL Neuropeptide Y, 20 ng/mL biotinylated Neuropeptide Y). Mix thoroughly. This solution serves as the first standard. b. To make the 100 ng/mL standard, pipette 50 µL of Neuropeptide Y stock solution the tube labeled 100 ng/mL. Mix thoroughly. c. Repeat this step with each successive concentration, preparing a dilution series as shown in the illustration below. Each time, use 450 µL of biotinylated Neuropeptide Y and 50 µL of the prior concentration until 100 pg/mL is reached. Mix each tube thoroughly before the next transfer. d. The final tube (0 pg/mL Neuropeptide Y, 20 ng/mL biotinylated Neuropeptide Y) serves as the zero standard (or total binding).
      6. Prepare a 10-fold dilution of Item F. To do this, add 2 µL of Item F to 18 µL of 1X Assay Diluent E. This solution will be used in steps 7 and
      9.
      7. Positive Control Preparation: Briefly centrifuge the positive control vial and reconstitute with 100 µL of ddH2O before use (Item M). To the tube of Item M, add 101 µL 1x Assay Diluent E. Also add 4 µL of 10-fold diluted Item F (prepared in step 6) to the tube. This is a 2-fold dilution of the positive control. Mix thoroughly. The positive control is a cell culture medium sample that is meant to be a system control (to verify that the detection & kit components are working). It may be diluted further if desired, but be sure the final concentration of biotinylated Neuropeptide Y is 20 ng/mL.
      8. If Item B (20X Wash Concentrate) contains visible crystals, warm to room temperature and mix gently until dissolved. 1000 100 10 1 100 0 ng/mL ng/mL ng/mL ng/mL pg/mL pg/mL 50 µL 50 µL 50 µL 50 µL Dilute 20 µL of Wash Buffer Concentrate into deionized or distilled water to yield 400 µL of 1X Wash Buffer.
      9. Sample Preparation: Use 1X Assay Diluent E + biotinylated NPY to dilute samples, including serum/plasma, cell culture medium and other sample types. It is very important to make sure the final concentration of the biotinylated Neuropeptide Y is 20 ng/mL in every sample.
      Example: to make a 4-fold dilution of sample, mix together 5 µL of 10-fold diluted Item F (prepared in step 6), 182.5 µL of 1X Assay Diluent E, and 62.5 µL of your sample, mix gently. The total volume is 250 µl, enough for duplicate wells on the microplate. Do not use Item F diluent from Step 5 for sample preparation. If you plan to use undiluted samples, you must still add biotinylated Neuropeptide Y to a final concentration of 20 ng/mL.
      Example: Add 5 µL of 10-fold diluted Item F to 245 µL of sample.
      10. Briefly centrifuge the HRP-Streptavidin vial (Item G) before use. The HRP-Streptavidin concentrate should be diluted 400- fold with 1X Assay Diluent E.
    Préparation de l'échantillon

    Use 1X Assay Diluent E + biotinylated NPY to dilute samples, including serum/plasma, cell culture medium and other sample types. It is very important to make sure the final concentration of the biotinylated Neuropeptide Y is 20 ng/mL in every sample. EXAMPLE: to make a 4-fold dilution of sample, mix together 5 µL of 10-fold diluted Item F (prepared in step 6), 182.5 µL of 1X Assay Diluent E, and 62.5 µL of your sample, mix gently. The total volume is 250 µl, enough for duplicate wells on the microplate. Do not use Item F diluent from Step 5 for sample preparation. If you plan to use undiluted samples, you must still add biotinylated Neuropeptide Y to a final concentration of 20 ng/mL. EXAMPLE: Add 5 µL of 10-fold diluted Item F to 245 µL of sample.

    Procédure de l'essai
    1. Keep kit reagents on ice during reagent preparation steps. It is recommended that all standards and samples be run at least in duplicate.
      2. Add 100 µL anti-Neuropeptide Y antibody (see Reagent Preparation step 3) to each well. Incubate for 1.5 hours at 0 room temperature with gentle shaking (1-2 cycles/sec). You may also incubate overnight at 4 degrees C.
      3. Discard the solution and wash wells 4 times with 1x Wash Buffer (200-300 µL each). Washing may be done with a multichannel pipette or an automated plate washer. Complete removal of liquid at each step is essential to good assay performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
      4. Add 100 µL of each standard (see Reagent Preparation step 5), positive control (see Reagent Preparation step 7) and sample (see Reagent Preparation step 9) into appropriate wells. Be sure to include a blank well (Assay Diluent only). Cover wells and incubate for 2.5 hours at room temperature with gentle shaking (1-2 cycles/sec) or overnight at 4 °C.
      5. Discard the solution and wash 4 times as directed in Step
      3.
      6. Add 100 µL of prepared HRP-Streptavidin solution (see Reagent Preparation step 10) to each well. Incubate with gentle shaking for 45 minutes at room temperature. It is recommended that incubation time should not be shorter or longer than 45 minutes.
      7. Discard the solution and wash 4 times as directed in Step
      3.
      8. Add 100 µL of TMB One-Step Substrate Reagent (Item H) to each well. Incubate for 30 minutes at room temperature in the dark with gentle shaking (1-2 cycles/sec).
      9. Add 50 µL of Stop Solution (Item I) to each well. Read absorbances at 450 nm immediately. 1
    Calcul des résultats

    Calculate the mean absorbance for each set of duplicate standards, controls and samples, and subtract the blank optical density. Plot the standard curve using SigmaPlot software (or other software which can perform four-parameter logistic regression models), with standard concentration on the x-axis and percentage of absorbance on the y-axis. Draw the best-fit curve through the standard points.

    Précision du teste
    Intra-Assay: CV < 10 %
    Inter-Assay: CV < 15 %
    Restrictions
    For Research Use only
  • Conseil sur la manipulation
    Avoid repeated freeze/thaw cycles.
    Stock
    -20 °C
    Stockage commentaire
    Standard, Biotinylated ProPPY peptide, and Positive Control should be stored at -20°C after arrival. Avoid multiple freeze-thaws. The remaining kit components may be stored at 4°C. Opened Microplate Wells and antibody (Item N) may be stored for up to 1 month at 2° to 8°C. Return unused wells to the pouch containing desiccant pack and reseal along entire edge.
    Date de péremption
    6 months
  • Kaya, Orbak, Polat, Polat, Gümüşdere: "Leptin and neuropeptide Y levels in newborns." dans: Journal of pediatric endocrinology & metabolism : JPEM, Vol. 29, Issue 1, pp. 21-5, (2016) (PubMed).

    Symons, ElGhazi, Reilly, Barney, Hanson, Panoskaltsis-Mortari, Armitage, Wilcox: "Can biomarkers differentiate pain and no pain subgroups of nonverbal children with cerebral palsy? A preliminary investigation based on noninvasive saliva sampling." dans: Pain medicine (Malden, Mass.), Vol. 16, Issue 2, pp. 249-56, (2015) (PubMed).

    Wysoki?ski: "Fasting serum levels of neuropeptide y in patients with schizophrenia on clozapine monotherapy." dans: Clinical neuropharmacology, Vol. 38, Issue 1, pp. 18-22, (2015) (PubMed).

    Bilir, Engin, Can, Temi, Demirtas: "The prognostic role of inflammation and hormones in patients with metastatic cancer with cachexia." dans: Medical oncology (Northwood, London, England), Vol. 32, Issue 3, pp. 56, (2015) (PubMed).

    Gao, Li, Zhu, Zhang, Hou, Sun, Liu, Peng, Gu: "Chronic administration of methamphetamine promotes atherosclerosis formation in ApoE-/- knockout mice fed normal diet." dans: Atherosclerosis, Vol. 243, Issue 1, pp. 268-77, (2015) (PubMed).

    Bae, Oh, Park, Noh, Noh, Kim, Lee: "Progressive muscle relaxation therapy for atopic dermatitis: objective assessment of efficacy." dans: Acta dermato-venereologica, Vol. 92, Issue 1, pp. 57-61, (2012) (PubMed).

    Wu, Xia, Xu, Zhai, Zhang, Zhang, Li, Li, Gu, Ma, Lu: "Acute effects of different glycemic index diets on serum motilin, orexin and neuropeptide Y concentrations in healthy individuals." dans: Neuropeptides, Vol. 46, Issue 3, pp. 113-8, (2012) (PubMed).

    Ilhan-Mutlu, Wagner, Wöhrer, Jungwirth, Marosi, Fischer, Preusser: "Blood alterations preceding clinical manifestation of glioblastoma." dans: Cancer investigation, Vol. 30, Issue 9, pp. 625-9, (2012) (PubMed).

    Matic, Matthews, Kizivat, Igwe, Marijanovic, Ruohonen, Savontaus, Adams, Kalajzic: "Bone-specific overexpression of NPY modulates osteogenesis." dans: Journal of musculoskeletal & neuronal interactions, Vol. 12, Issue 4, pp. 209-18, (2012) (PubMed).

    Vijay-Kumar, Aitken, Carvalho, Cullender, Mwangi, Srinivasan, Sitaraman, Knight, Ley, Gewirtz: "Metabolic syndrome and altered gut microbiota in mice lacking Toll-like receptor 5." dans: Science (New York, N.Y.), Vol. 328, Issue 5975, pp. 228-31, (2010) (PubMed).

    Kuo, Kitlinska, Tilan, Li, Baker, Johnson, Lee, Burnett, Fricke, Kvetnansky, Herzog, Zukowska: "Neuropeptide Y acts directly in the periphery on fat tissue and mediates stress-induced obesity and metabolic syndrome." dans: Nature medicine, Vol. 13, Issue 7, pp. 803-11, (2007) (PubMed).

    King, Widdowson, Doods, Williams: "Regulation of neuropeptide Y release by neuropeptide Y receptor ligands and calcium channel antagonists in hypothalamic slices." dans: Journal of neurochemistry, Vol. 73, Issue 2, pp. 641-6, (1999) (PubMed).

    Pomonis, Levine, Billington: "Interaction of the hypothalamic paraventricular nucleus and central nucleus of the amygdala in naloxone blockade of neuropeptide Y-induced feeding revealed by c-fos expression." dans: The Journal of neuroscience : the official journal of the Society for Neuroscience, Vol. 17, Issue 13, pp. 5175-82, (1997) (PubMed).

    Dryden, Pickavance, Frankish, Williams: "Increased neuropeptide Y secretion in the hypothalamic paraventricular nucleus of obese (fa/fa) Zucker rats." dans: Brain research, Vol. 690, Issue 2, pp. 185-8, (1996) (PubMed).

    Hanson, Dallman: "Neuropeptide Y (NPY) may integrate responses of hypothalamic feeding systems and the hypothalamo-pituitary-adrenal axis." dans: Journal of neuroendocrinology, Vol. 7, Issue 4, pp. 273-9, (1995) (PubMed).

    White, Dean, Edwards, Martin: "Type II corticosteroid receptor stimulation increases NPY gene expression in basomedial hypothalamus of rats." dans: The American journal of physiology, Vol. 266, Issue 5 Pt 2, pp. R1523-9, (1994) (PubMed).

    Kaye, Berrettini, Gwirtsman, George: "Altered cerebrospinal fluid neuropeptide Y and peptide YY immunoreactivity in anorexia and bulimia nervosa." dans: Archives of general psychiatry, Vol. 47, Issue 6, pp. 548-56, (1990) (PubMed).

    Allen, Adrian, Allen, Tatemoto, Crow, Bloom, Polak: "Neuropeptide Y distribution in the rat brain." dans: Science (New York, N.Y.), Vol. 221, Issue 4613, pp. 877-9, (1983) (PubMed).

  • Antigène Voir toutes NPY Kits ELISA
    NPY (Neuropeptide Y (NPY))
    Autre désignation
    Neuropeptide Y (NPY Produits)
    Synonymes
    0710005A05Rik Kit ELISA, PYY4 Kit ELISA, NPY02 Kit ELISA, RATNPY Kit ELISA, RATNPY02 Kit ELISA, si:dkey-22m8.5 Kit ELISA, npy Kit ELISA, npyb Kit ELISA, preproNPY Kit ELISA, pyy4 Kit ELISA, xnpy Kit ELISA, npya Kit ELISA, MGC86288 Kit ELISA, prepronpy Kit ELISA, NPY Kit ELISA, NPY1 Kit ELISA, neuropeptide Y Kit ELISA, neuropeptide Y S homeolog Kit ELISA, neuropeptide Y L homeolog Kit ELISA, Npy Kit ELISA, NPY Kit ELISA, npy Kit ELISA, npy.S Kit ELISA, npy.L Kit ELISA, LOC100533423 Kit ELISA
    Sujet
    Neuropeptide Y (NPY) is a 36 amino acid peptide hormone found in the neural system and has an important role in obesity. The main effect of Neuropeptide Y is increased food intake and decreased physical activity. It also increases the proportion of energy stored as fat and blocks nociceptive signals to the brain. In addition to its role in obesity, Neuropeptide Y has been associated with a number of other physiologic processes in the brain, including the regulation of energy balance, memory and learning, and epilepsy. Animal studies strongly demonstrate that the stimulation of Neuropeptide Yergic activity via the administration of certain Neuropeptide Y agonists increases food intake compared to control animals. The effects of Neuropeptide Yergic activity on food intake is also demonstrated by the blockade of certain Neuropeptide Y receptors (Y1 and Y5 receptors), which expectedly inhibited Neuropeptide Yergic activity, thus, decreases food intake. For its role in obesity, an increase in Neuropeptide Y is caused by high levels of glucocorticosteriods through directly activating type II glucocorticosteriods receptors and indirectly, by abolishing the negative feedback of CRF on Neuropeptide Y synthesis and release. Meanwhile, obesity-induced insulin resistance and the mutation of the leptin receptor (ObRb) results in the abolishment of other negative feedback mechanisms to regulate Neuropeptide Yergic activity and ultimately food intake. High levels of Neuropeptide Y were also found in the cerebrospinal fluid of patients with anorexia nervosa.
    ID gène
    4852
    UniProt
    P01303
    Pathways
    Feeding Behaviour
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