Anti-MMP-24 Rabbit Polyclonal Antibody

Frequently Asked Questions

What is the predicted molecular weight of MMP-24 on Western blot?

Full-length MMP-24 (MT5-MMP) is a 645-amino acid protein with a predicted molecular weight of approximately 70 kDa. However, MMP-24 undergoes post-translational glycosylation as a membrane-anchored metalloprotease, which often increases its apparent molecular weight to 75-80 kDa on Western blot under reducing conditions. If you observe additional lower molecular weight bands, these may represent processed or shed ectodomain fragments, as membrane-type MMPs are subject to autoproteolytic cleavage and ectodomain shedding. Always include untransfected controls to distinguish specific signal from background, particularly in lysates from neural or immune-enriched tissues where MMP-24 expression is regulated.

What dilution should I start with for Western blot using this MMP-24 antibody?

The recommended starting dilution for Western blot is 1:1000 in blocking buffer (typically 5% non-fat milk or BSA in TBST). Because this is a rabbit polyclonal raised against human MMP-24, sensitivity will vary with your sample type and MMP-24 expression level. Neural stem cells, ependymal tissues, and dorsal root ganglia express MMP-24 at moderate to low endogenous levels, so you may need to optimize between 1:500 and 1:2000. Load 20-40 µg total protein per lane initially, and include a positive control lysate from HEK293 cells transfected with MMP-24 if endogenous signal is weak.

Does this MMP-24 antibody cross-react with other matrix metalloproteinases?

This antibody was raised against human MMP-24 and validated for specificity to MMP-24 in human lysates. The membrane-type MMP family (MT1-MMP through MT6-MMP) shares structural homology, particularly in the catalytic and hemopexin domains, so cross-reactivity with closely related MT-MMPs such as MMP-14 (MT1-MMP) or MMP-15 (MT2-MMP) is theoretically possible but has not been systematically tested. If you are working with samples that co-express multiple MT-MMPs, include siRNA or CRISPR knockdown controls for MMP-24 to confirm band specificity. Reactivity with soluble MMPs such as MMP-2 or MMP-9 is not expected given the structural divergence.

Will this antibody detect MMP-24 in mouse or rat samples?

This antibody is validated for human MMP-24. Cross-reactivity with mouse or rat MMP-24 is predicted but not experimentally validated by Triple Point Biologics. Human and mouse MMP-24 share approximately 80-85% amino acid sequence identity, with the highest conservation in the catalytic domain. If you plan to use this antibody in rodent models of neural stem cell quiescence or thermal nociception, we recommend testing it at 1:500 to 1:1000 on mouse or rat tissue lysates alongside a known human-positive control. Confirm specificity by comparing wild-type and MMP-24 knockout mouse samples if available, as non-specific binding patterns may differ across species.

Can I use this MMP-24 antibody for immunohistochemistry or immunofluorescence?

Triple Point Biologics antibodies are broadly validated for Western blot and immunohistochemistry applications. While the recommended dilution provided is specific to Western blot (1:1000), rabbit polyclonal antibodies against membrane-anchored targets like MMP-24 typically perform well in IHC and immunofluorescence on fixed tissue sections. For IHC, start with a 1:100 to 1:200 dilution following antigen retrieval (citrate buffer, pH 6.0, is a standard starting point for metalloproteases). For immunofluorescence on cultured neural stem cells or DRG neurons, try 1:100 to 1:500 after methanol or paraformaldehyde fixation. Include no-primary-antibody controls to assess background in your tissue type.

What is the best positive control tissue or cell line for MMP-24 expression?

MMP-24 is expressed at physiologically relevant levels in the subependymal zone of the adult brain, where it regulates neural stem cell quiescence by cleaving N-cadherin, and in dorsal root ganglia in the context of thermal nociception. For a readily available positive control, consider lysates from human brain tissue (commercially available) or HEK293 or HeLa cells transiently transfected with full-length human MMP-24 cDNA. Endogenous MMP-24 expression in standard immortalized cell lines is often low or undetectable, so overexpression constructs provide a strong, specific signal for initial antibody validation. Include mock-transfected cells as a negative control to confirm specificity.

How should I store this MMP-24 antibody and what is the expected shelf life?

Store the antibody at -20°C in single-use aliquots to avoid repeated freeze-thaw cycles, which can reduce antibody titer and increase aggregation. The 100 µg pack size is sufficient for approximately 100 Western blots at 1:1000 dilution (assuming 1 mL blocking volume per blot). For long-term storage beyond one year, keep aliquots at -80°C. Once thawed, an aliquot can be stored at 4°C for up to one month if sodium azide (0.02-0.05% final concentration) is added as a preservative. Triple Point Biologics has manufactured proteinase antibodies since 1994, and polyclonal antibodies typically retain activity for several years under proper storage conditions.

Why do I see multiple bands on Western blot with this MMP-24 antibody?

Multiple bands may reflect the complex processing of membrane-anchored MMP-24. The full-length glycosylated form appears at 75-80 kDa, but MMP-24 undergoes proteolytic activation and ectodomain shedding, generating lower molecular weight fragments (typically 50-60 kDa soluble ectodomain and smaller C-terminal membrane-bound fragments). Additionally, differential glycosylation states in different cell types or tissue contexts can produce a smeared or doublet appearance. If you observe high-molecular-weight species above 100 kDa, consider the possibility of MMP-24 homodimers or complexes with tissue inhibitors of metalloproteinases (TIMPs). Run samples under fully reducing conditions (DTT or beta-mercaptoethanol) and boil lysates adequately to minimize aggregation artifacts.