Human TIMP-2 Quantikine ELISA Kit

  (18 citations)     
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Assay Procedure
Citations (18)
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  • Assay Type
    Solid Phase Sandwich ELISA
  • Format
    96-well strip plate
  • Assay Length
    4.5 hours
  • Sample Type & Volume Required Per Well
    Cell Culture Supernates (50 uL), Serum (10 uL), EDTA Plasma (10 uL), Heparin Plasma (10 uL), Saliva (10 uL)
  • Sensitivity
    0.064 ng/mL
  • Assay Range
    0.2 - 10 ng/mL (Cell Culture Supernates, Serum, EDTA Plasma, Heparin Plasma, Saliva)
  • Specificity
    Natural and recombinant human TIMP-2. This assay detects approximately 50% of recombinant human TIMP-2 when complexed with recombinant human active MMP-9 in a 1:1 molar ratio.
  • Cross-reactivity
    < 0.5% cross-reactivity observed with available related molecules.Cross-species reactivity not tested.
  • Interference
    No significant interference observed with available related molecules.
Product Summary
The Quantikine Human TIMP-2 Immunoassay is a 4.5 hour solid phase ELISA designed to measure TIMP-2 levels in cell culture supernates, serum, plasma, and saliva. It contains CHO cell-expressed recombinant human TIMP-2, and antibodies raised against the recombinant protein. Natural human TIMP-2 showed dose-response curves that were parallel to the standard curves obtained using the recombinant Quantikine kit standards, indicating that this kit can be used to determine relative levels of natural human TIMP-2.

Precision
Intra-Assay Precision (Precision within an assay) Three samples of known concentration were tested on one plate to assess intra-assay precision
Inter-Assay Precision (Precision between assays) Three samples of known concentration were tested in separate assays to assess inter-assay precision
Cell Culture Supernates, Saliva
Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 40 40 40
Mean 1.01 2.9 5.26 1.01 2.79 5.27
Standard Deviation 0.042 0.173 0.344 0.079 0.188 0.326
CV% 4.2 6 6.5 7.8 6.7 6.2

Serum, EDTA Plasma, Heparin Plasma
Intra-Assay Precision Inter-Assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 40 40 40
Mean 1.23 3.45 6.09 1.26 3.45 6.38
Standard Deviation 0.054 0.116 0.181 0.086 0.197 0.467
CV% 4.4 3.4 3 6.8 5.7 7.3

Recovery

The recovery of TIMP-2 spiked to three different levels throughout the range of the assay was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 95 81-108
Linearity
To assess the linearity of the assay, samples containing or spiked with high concentrations of TIMP-2 were diluted with Calibrator Diluent to produce samples with values within the dynamic range of the assay.
 TIMP-2 [HRP]
Product Datasheets

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Preparation and Storage
  • Storage
    Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Background: TIMP-2
TIMPs-1 through -4 regulate the activity of zinc metalloproteases known as MMPs, ADAMs and ADAMTSs. Structurally, TIMPs contain two domains. The N-terminal domain binds to the active site of mature metalloproteases via a 1:1 non-covalent interaction, blocking access of substrates to the catalytic site. In addition, The C-terminal domain of TIMP-1 and TIMP-2 binds to the hemopexin- like domain of pro-MMP-9 and pro-MMP-2, respectively. The latter binding is essential for the cell surface activation of MMP-2 by MMP-14.
  • Long Name:
    Tissue Inhibitors of Metalloproteinases 2
  • Entrez Gene IDs:
    7077 (Human); 21858 (Mouse); 29543 (Rat)
  • Alternate Names:
    CSC-21Ktissue inhibitor of metalloproteinase 2; metalloproteinase inhibitor 2; TIMP metallopeptidase inhibitor 2; TIMP2; TIMP-2; Tissue inhibitor of metalloproteinases 2
Related Research Areas
Assay Procedure
Refer to the product for complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.
  1.   Prepare all reagents, standard dilutions, and samples as directed in the product insert.
  2.   Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

  3. 100 µL Assay Diluent
  4.   Add 100 µL of Assay Diluent to each well.

  5. 50 µL Standard, Control, or Sample
  6.   Add 50 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours on a horizontal orbital microplate shaker.
  7.   Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.

  8. 200 µL Conjugate
  9.   Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours on the shaker.
  10.   Aspirate and wash 4 times.

  11. 200 µL Substrate Solution
  12.   Add 200 µL Substrate Solution to each well.
  13.   For Cell Culture Supernate & Saliva Samples: Incubate at room temperature for 20 minutes on the benchtop. PROTECT FROM LIGHT.
    For Serum & Plasma Samples: Incubate at room temperature for 30 minutes on the benchtop. PROTECT FROM LIGHT.

  14. 50 µL Stop Solution
  15. Add 50 µL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.
Citations:

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.

18 Citations: Showing 1 - 10
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Species
Sample Type
  1. Human umbilical cord plasma proteins revitalize hippocampal function in aged mice
    Authors: JM Castellano, KI Mosher, RJ Abbey, AA McBride, ML James, D Berdnik, JC Shen, B Zou, XS Xie, M Tingle, IV Hinkson, MS Angst, T Wyss-Coray
    Nature, 2017;544(7651):488-492.
    Species: Human
    Sample Type: Plasma
  2. TIMP1 and MMP9 are predictors of mortality in septic patients in the emergency department and intensive care unit unlike MMP9/TIMP1 ratio: Multivariate model
    Authors: ME Niño, SE Serrano, DC Niño, DM McCosham, ME Cardenas, VP Villareal, M Lopez, A Pazin-Filh, FA Jaimes, F Cunha, R Schulz, D Torres-Due
    PLoS ONE, 2017;12(2):e0171191.
    Species: Human
    Sample Type: Serum
  3. Role of serum VEGFA, TIMP2, MMP2 and MMP9 in monitoring response to adjuvant radiochemotherapy in patients with primary cervical cancer--results of a companion protocol of the randomized NOGGO-AGO phase III clinical trial.
    Authors: Braicu E, Gasimli K, Richter R, Nassir M, Kummel S, Blohmer J, Yalcinkaya I, Chekerov R, Ignat I, Ionescu A, Mentze M, Fotopoulou C, Pop C, Lichtenegger W, Sehouli J
    Anticancer Res, 2014;34(1):385-91.
    Species: Human
    Sample Type: Serum
  4. Role of serum concentration of VEGFR1 and TIMP2 on clinical outcome in primary cervical cancer: results of a companion protocol of the randomized, NOGGO-AGO phase III adjuvant trial of simultaneous cisplatin-based radiochemotherapy vs. carboplatin and paclitaxel containing sequential radiotherapy.
    Authors: Braicu E, Fotopoulou C, Chekerov R, Richter R, Blohmer J, Kummel S, Stamatian F, Yalcinkaya I, Mentze M, Lichtenegger W, Sehouli J
    Cytokine, 2013;61(3):755-8.
    Species: Human
    Sample Type: Serum
  5. Increased levels of leukocyte-derived MMP-9 in patients with stable angina pectoris.
    Authors: Jonasson S, Lundberg A, Kalvegren H, Bergstrom I, Szymanowski A, Jonasson L
    PLoS ONE, 2011;6(4):e19340.
    Species: Human
    Sample Type: Serum
  6. Activation of matrix metalloproteinases following anti-Abeta immunotherapy; implications for microhemorrhage occurrence.
    Authors: Wilcock DM, Morgan D, Gordon MN, Taylor TL, Ridnour LA, Wink DA, Colton CA
    J Neuroinflammation, 2011;8(0):115.
    Species: Mouse
    Sample Type: Tissue Homogenates
  7. Abnormalities in metalloproteinase pathways and IGF-I axis: a link between birth weight, hypertension, and vascular damage in childhood.
    Authors: Sesso R, Franco MC
    Am. J. Hypertens., 2010;23(1):6-11.
    Species: Human
    Sample Type: Plasma
  8. Primary human acute myelogenous leukemia cells release matrix metalloproteases and their inhibitors: release profile and pharmacological modulation.
    Authors: Reikvam H, Hatfield KJ, Oyan AM, Kalland KH, Kittang AO, Bruserud O
    Eur. J. Haematol., 2010;84(3):239-51.
    Species: Human
    Sample Type: Cell Culture Supernates
  9. Salbutamol up-regulates matrix metalloproteinase-9 in the alveolar space in the acute respiratory distress syndrome.
    Authors: O'Kane CM, McKeown SW, Perkins GD, Bassford CR, Gao F, Thickett DR, McAuley DF
    Crit. Care Med., 2009;37(7):2242-9.
    Species: Human
    Sample Type: BALF
  10. MMP expression and abnormal lung permeability are important determinants of outcome in IPF.
    Authors: McKeown S, Richter AG, O'Kane C, O'Kane C, McAuley DF, Thickett DR
    Eur. Respir. J., 2009;33(1):77-84.
    Species: Human
    Sample Type: BALF
  11. Immunomodulation by chronobiologically-based glucocorticoids treatment for multiple sclerosis relapses.
    Authors: Glass-Marmor L, Paperna T, Galboiz Y, Miller A
    J. Neuroimmunol., 2009;210(1):124-7.
    Species: Human
    Sample Type: Serum
  12. Imbalance of tissue inhibitors of metalloproteinases (TIMP) - 1 and - 4 serum levels, in patients with inflammatory bowel disease.
    Authors: Kapsoritakis AN, Kapsoritaki AI, Davidi IP, Lotis VD, Manolakis AC, Mylonis PI, Theodoridou AT, Germenis AE, Potamianos SP
    BMC Gastroenterol, 2008;8(1):55.
    Species: Human
    Sample Type: Serum
  13. Impact of acute exposure to tobacco smoke on gelatinases in the bronchoalveolar space.
    Authors: Glader P, Eldh B, Bozinovski S, Andelid K, Sjostrand M, Malmhall C, Anderson GP, Riise GC, Qvarfordt I, Linden A
    Eur. Respir. J., 2008;32(3):644-50.
    Species: Human
    Sample Type: BALF
  14. Is metalloproteinase-7 specific for idiopathic pulmonary fibrosis?
    Authors: Huh JW, Kim DS, Oh YM, Shim TS, Lim CM, Lee SD, Koh Y, Kim WS, Kim WD, Kim KR
    Chest, 2007;133(5):1101-6.
    Species: Human
    Sample Type: BALF
  15. Actinobacillus actinomycetemcomitans lipopolysaccharide regulates matrix metalloproteinase, tissue inhibitors of matrix metalloproteinase, and plasminogen activator production by human gingival fibroblasts: a potential role in connective tissue destruction.
    Authors: Bodet C, Andrian E, Tanabe S, Grenier D
    J. Cell. Physiol., 2007;212(1):189-94.
    Species: Human
    Sample Type: Cell Culture Supernates
  16. Matrix metalloproteinase-2 dysregulation in type 1 diabetes.
    Authors: Thrailkill KM, Bunn RC, Moreau CS, Cockrell GE, Simpson PM, Coleman HN, Frindik JP, Kemp SF, Fowlkes JL
    Diabetes Care, 2007;30(9):2321-6.
    Species: Human
    Sample Type: Plasma
  17. Plasma MMP-2 and MMP-9 and their inhibitors TIMP-1 and TIMP-2 during human orthotopic liver transplantation. The effect of aprotinin and the relation to ischemia/reperfusion injury.
    Authors: Kuyvenhoven JP, Molenaar IQ, Verspaget HW, Veldman MG, Palareti G, Legnani C, Moolenburgh SE, Terpstra OT, Lamers CB, van Hoek B, Porte RJ
    Thromb. Haemost., 2004;91(3):506-13.
    Species: Human
    Sample Type: Plasma
  18. Matrix metalloproteinases and tissue inhibitors of metalloproteinases in sera and tissue of patients with Dupuytren's disease.
    Authors: Ulrich D, Hrynyschyn K, Pallua N
    Plast. Reconstr. Surg., 2003;112(5):1279-86.
    Species: Human
    Sample Type: Serum
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