TIMP-1 (Recombinant)
- Expression system
- HEK293
- Cat. #
- REC-TIMP1
In stock
- SKU
- REC-TIMP1
Target Overview
TIMP-1 (Tissue Inhibitor of Metalloproteinases 1, UniProt P01033) is a secreted 207-amino acid glycoprotein that acts as a stoichiometric inhibitor of matrix metalloproteinases (MMPs). It forms tight 1:1 non-covalent complexes with the catalytic zinc cofactor of target MMPs, effectively and irreversibly neutralising their proteolytic activity. Documented MMP substrates include MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, and MMP16; notably, MMP14 (MT1-MMP) is not inhibited. Beyond its role as a proteinase inhibitor, TIMP-1 functions as a growth factor, engaging the cell-surface receptors CD63 and integrin β1 (ITGB1) to modulate cell differentiation, migration, and apoptotic signalling pathways. This recombinant is produced in HEK293 cells, an expression system that supports the post-translational glycosylation characteristic of the native human secreted form — a relevant consideration for inhibition kinetics assays where glycosylation status can influence MMP-binding affinity. The full 207-residue coding sequence (signal peptide through C-terminus) is expressed, yielding a secreted, processed form comparable to the endogenous protein. In the laboratory, this recombinant is well-suited to MMP inhibition activity assays (e.g., fluorogenic peptide substrate assays with MMP1, MMP2, or MMP9), Ki or IC50 determination for TIMP-1/MMP complexes, and as a reference standard in ELISA or multiplex proteomic workflows measuring extracellular matrix remodelling. Researchers performing Western blot or IHC antibody validation can use this protein as a defined positive control; it pairs directly with the Triple Point Biologics matched antibody (SKU: RP-TIMP1). The recombinant is also suitable as a competitor ligand in CD63 or ITGB1 binding/displacement assays exploring TIMP-1's growth-factor signalling axis.
Background
Applications
- MMP inhibition activity assay using fluorogenic peptide substrates (e.g., MMP1, MMP2, MMP9 cleavage of Mca-PLGL-Dpa-AR-NH2)
- Inhibitor Ki or IC50 determination for TIMP-1/MMP binary complex formation
- Western blot positive control for TIMP-1 detection, paired with TPB antibody SKU RP-TIMP1
- IHC antibody validation: defined mass standard for anti-TIMP-1 antibody titration
- ELISA reference standard in multiplex ECM remodelling panels measuring MMP/TIMP balance
- CD63 or integrin β1 (ITGB1) ligand-binding and receptor displacement assay
- Recombinant competitor in surface plasmon resonance (SPR) or bio-layer interferometry (BLI) affinity measurements with target MMPs
- Spiking standard for quantitative proteomics (e.g., BAL fluid or conditioned medium) to calibrate TIMP-1 abundance measurements
References
- Zhu C et al. Lack of alpha1,2-fucosylation protects mice from bleomycin-induced lung fibrosis: A quantitative proteomic analysis of bronchoalveolar lavage fluid. Carbohydr Res. 2026. doi:10.1016/j.carres.2026.110010. PMID: 42320172.
- Guo X et al. Fupenzic acid ameliorates radiation-induced liver fibrosis by regulating RBMS1 to inhibit the cGAS-STING pathway and restore mitochondrial function. Phytomedicine. 2026. doi:10.1016/j.phymed.2026.158399. PMID: 42314350.
- Perkins RK et al. Aerobic Exercise Training Increases Circulating sRAGE in Adults With Type 2 Diabetes: Associations With Sheddase Regulation. Diabetes Obes Metab. 2026. doi:10.1111/dom.70995. PMID: 42310923.
- Ma W et al. Multi-algorithm machine learning combined with in silico gene knockout reveals the diagnostic value and functional regulatory networks of ferroptosis-related genes in gastric cancer. Transl Cancer Res. 2026. doi:10.21037/tcr-2026-1-0035. PMID: 42305491.
- Zhang A et al. Enhancing the efficacy of ulcerative colitis treatment by inhibiting LCN2-mediated pyroptosis through traditional processing techniques: With the rhizome of Atractylodes macrocephala Koidz. as an example. J Ethnopharmacol. 2026. doi:10.1016/j.jep.2026.122037. PMID: 42302942.
Additional Specifications
| Specific Activity | Functional MMP inhibitor; Ki ≤ nM range against MMP-1/3/9 |
|---|---|
| Storage Buffer | 50 mM Tris-HCl pH 7.5, 150 mM NaCl, 10% glycerol |
| Sequence (aa range) | amino acids 24–207 (mature secreted form, signal peptide removed) |
| Endotoxin Level | <0.1 EU/µg by LAL |
| Purity (%) | >95% by SDS-PAGE |
| Expression System | HEK293 |
| Molecular Weight (kDa) | ~28 kDa (glycosylated) |
| Subcellular Localization | Secreted; Extracellular matrix |
Frequently Asked Questions
What molecular weight band should I expect for recombinant TIMP-1 on SDS-PAGE or Western blot?
The unmodified TIMP-1 polypeptide (UniProt P01033) runs at a predicted ~23 kDa, but the HEK293-expressed recombinant carries native-like N-linked glycosylation, shifting the apparent molecular weight to approximately 28–32 kDa on reducing SDS-PAGE — consistent with the endogenous secreted form seen in conditioned media. Under non-reducing conditions the two conserved disulfide-linked domains remain intact, so mobility may shift slightly. If your Western blot shows a diffuse band in the 28–32 kDa range, that is expected and not indicative of degradation.
Which MMPs does recombinant TIMP-1 inhibit, and does it inhibit MMP-14 (MT1-MMP)?
TIMP-1 forms tight 1:1 non-covalent complexes with the catalytic zinc of MMP-1, -2, -3, -7, -8, -9, -10, -11, -12, -13, and -16, effectively neutralising their proteolytic activity. Notably, MMP-14 (MT1-MMP) is not inhibited by TIMP-1 — a distinction with practical consequences for experiments targeting membrane-type MMPs. If your assay involves MT1-MMP-driven pericellular proteolysis, this recombinant will not serve as a relevant inhibitor control; TIMP-2 or TIMP-3 would be more appropriate choices for that substrate.
What fluorogenic substrate and assay conditions should I use to measure TIMP-1 inhibitory activity against MMP-9?
A standard approach uses the quenched fluorescent substrate Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 (or equivalent MMP-9-preferring peptide) at 10 µM in assay buffer containing 50 mM Tris-HCl pH 7.5, 150 mM NaCl, 5 mM CaCl₂, and 0.05% Brij-35. Pre-incubate activated MMP-9 (1–5 nM) with serial dilutions of TIMP-1 for 30–60 minutes at 37°C before adding substrate. This recombinant is produced in HEK293 cells, preserving glycosylation that influences MMP-binding kinetics, so Ki determinations should reflect physiologically relevant inhibition constants.
What starting concentration of recombinant TIMP-1 should I use for an IC50 experiment with MMP-3?
For IC50 determination against MMP-3, begin with a TIMP-1 concentration range of 0.1–100 nM, titrated in half-log steps. Because TIMP-1 acts stoichiometrically rather than as a classical competitive inhibitor, apparent IC50 values are enzyme-concentration-dependent. Keep MMP-3 at 1–2 nM (well below Ki) to approximate true inhibition constants. Reported Ki values for TIMP-1 against MMP-3 are in the low-nanomolar range (~1–5 nM), so you should observe near-complete inhibition at equimolar or slight molar excess of TIMP-1.
Can I use REC-TIMP1 as a positive control for Western blot with the matched RP-TIMP1 antibody?
Yes — REC-TIMP1 is specifically matched to the RP-TIMP1 rabbit polyclonal antibody and is the recommended positive control for that antibody's Western blot validation. The antibody is validated against the HEK293-expressed form, so band position and signal intensity are well-characterised. Load 20–50 ng of REC-TIMP1 per lane alongside your experimental lysates. Expect a diffuse band at 28–32 kDa under reducing conditions. Full product and validation details for RP-TIMP1 are at /anti-timp1-rabbit-polyclonal-antibody.
How much recombinant TIMP-1 should I load for a Western blot positive control lane?
20–50 ng per lane is sufficient for a clean, strong signal when using the matched RP-TIMP1 antibody. At 20 ng you will confirm specificity without saturating the detection channel; 50 ng is useful when optimising a new blot protocol or lower-sensitivity detection systems. Dilute from your working stock into 1× Laemmli sample buffer (reducing), boil for 5 minutes, and run alongside a molecular weight marker spanning 15–50 kDa to bracket the 28–32 kDa glycosylated band appropriately.
What is the storage buffer for REC-TIMP1 and how should I handle it after thawing?
REC-TIMP1 is supplied in 50 mM Tris-HCl pH 7.5, 150 mM NaCl, 10% glycerol. Store at −20°C in single-use aliquots; repeated freeze-thaw cycles measurably reduce inhibitory activity. After thawing, keep on ice and use within the same working session. If your downstream assay buffer is incompatible with glycerol (e.g., some SPR or ITC formats), dilute at least 1:10 into assay buffer to bring glycerol below 1%. Purity is >95% by SDS-PAGE and endotoxin is <0.1 EU/µg by LAL, so no additional clean-up steps are required for standard cell-based or biochemical assays.
Does TIMP-1 function only as a proteinase inhibitor, or does it have receptor-mediated signalling activity relevant to my cell biology assay?
TIMP-1 has well-documented MMP-independent signalling activity. It engages cell-surface receptors CD63 and integrin β1 (ITGB1) to modulate cell differentiation, migration, and apoptotic pathways. This means that at concentrations used in cell-based assays (typically 50–500 nM), observed phenotypic effects may reflect receptor signalling rather than — or in addition to — MMP inhibition. If you need to isolate the proteinase-inhibitory function from receptor-mediated effects, mutant TIMP-1 variants lacking CD63-binding capacity are described in the literature and should be considered as complementary controls.
Validation imagery coming soon
Western blot validation figures for REC-TIMP1 will be published here as they are produced in-house.
If you would like to see existing validation data for this antibody before publication, request a sample copy.