The Quantikine Human TIMP-4 Immunoassay is a 4.5 hour solid phase ELISA designed to measure TIMP-4 levels in cell culture supernates, serum, plasma, and human milk. It contains Sf 21-expressed recombinant human TIMP-4 and antibodies raised against the recombinant protein. This kit detects recombinant human TIMP-4 in both free and MMP-bound forms. Natural human TIMP-4 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-4.
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.
Serum, EDTA Plasma, Heparin Plasma, Human Milk
Cell Culture Supernates
The recovery of TIMP-4 spiked to levels throughout the range of the assay in various matrices was evaluated.
Average % Recovery
Cell Culture Supernates (n=4)
EDTA Plasma (n=4)
Heparin Plasma (n=4)
To assess the linearity of the assay, samples containing and/or spiked with high concentrations of TIMP-4 were serially diluted with the appropriate Calibrator Diluent to produce samples with values within the dynamic range of the assay.
Preparation and Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
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.
Tissue Inhibitors of Metalloproteinases 4
Entrez Gene IDs
7079 (Human); 110595 (Mouse);
metalloproteinase inhibitor 4; TIMP metallopeptidase inhibitor 4; TIMP4; TIMP-4; tissue inhibitor of metalloproteinase 4; Tissue inhibitor of metalloproteinases 4;
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.
Prepare all reagents, standard dilutions, and samples as directed in the product insert.
Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.
100 µL Assay Diluent
Add 100 µL of Assay Diluent to each well.
50 µL Standard, Control, or Sample
Add 50 µL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.
Aspirate each well and wash, repeating the process 3 times for a total of 4 washes.
200 µL Conjugate
Add 200 µL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.
Aspirate and wash 4 times.
200 µL Substrate Solution
Add 200 µL Substrate Solution to each well. Incubate at room temperature for 30 minutes. PROTECT FROM LIGHT.
50 µL Stop Solution
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.
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