Anti-DPP-6 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight should I expect for DPP-6 on a Western blot?

DPP-6 migrates at approximately 110-130 kDa on Western blot, higher than the predicted 95 kDa for the 865-residue polypeptide due to extensive N-glycosylation. You may observe multiple bands representing different glycosylation states or proteolytic fragments. Some labs report a doublet around 125-130 kDa in brain lysates. Treatment with PNGase F or other deglycosylases will shift the band closer to the predicted molecular weight. Always run a positive control lysate from DPP-6-expressing tissue such as mouse or rat brain to confirm the expected migration pattern.

Does DPP-6 have catalytic activity like other dipeptidyl peptidases?

No. Despite its classification in the S9B serine protease family and sequence homology to DPP4, DPP-6 lacks catalytic activity because critical active-site residues are not conserved. DPP-6 functions exclusively as a non-enzymatic auxiliary subunit that regulates the trafficking and gating kinetics of Kv4 potassium channels, particularly KCND2. This is an important distinction when designing functional assays. If you are studying enzymatic dipeptidyl peptidase activity in your samples, DPP-6 will not contribute to substrate cleavage, and you should target DPP4 or DPP8/9 instead.

What starting dilution should I use for Western blot with this antibody?

Start at 1:1000 dilution for Western blot applications, as recommended for this polyclonal. DPP-6 is abundantly expressed in brain tissue, so you may be able to dilute further to 1:2000 or 1:3000 depending on your sample type and detection system. For low-expressing cell lines or non-neuronal tissues, begin at 1:500 and titrate upward. Load 20-30 µg total protein per lane for tissue lysates. Use a sensitive ECL substrate if working with cardiac samples, where DPP-6 expression is lower than in brain. Always include a positive control lysate to optimize signal-to-noise.

Is this DPP-6 antibody cross-reactive with mouse and rat tissue?

The antibody has validated reactivity with human DPP-6 and predicted cross-reactivity with mouse and rat based on sequence homology in the immunogen region. Mouse and rat DPP-6 share greater than 90 percent identity with human in most epitope-rich domains. We have observed consistent results in rodent brain lysates during internal testing, but we recommend you validate performance in your specific application and tissue type. If you are working with primary neurons or rodent cardiac tissue, run a small pilot experiment to confirm signal specificity before committing to a full study.

What positive control tissue should I use for DPP-6 Western blot?

Adult brain tissue is the optimal positive control for DPP-6, particularly hippocampus, cortex, or cerebellum where expression is highest. Rat or mouse whole-brain lysate is widely available and provides robust signal. Human neuroblastoma or glioblastoma cell lines may express DPP-6 but at variable levels depending on differentiation state. Heart tissue is also positive but yields weaker signal than brain. Avoid using HeLa or HEK293 lysates as negative controls without verifying endogenous expression, since some subclones express low levels. A DPP-6 knockout or knockdown lysate provides the cleanest specificity control.

Can I use this antibody for immunofluorescence on neurons?

Yes, this rabbit polyclonal is suitable for immunofluorescence in addition to Western blot and IHC. For cultured neurons, fix with 4 percent paraformaldehyde for 10-15 minutes and permeabilize with 0.1-0.3 percent Triton X-100. Start at a 1:100 to 1:200 dilution and titrate based on signal. DPP-6 localizes to the plasma membrane and dendritic compartments, often in punctate patterns reflecting its association with Kv4.2 channels. Use a well-characterized secondary antibody with minimal cross-reactivity and always include a no-primary control to assess non-specific binding in your fixation and blocking conditions.

How should I store the 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 titer and increase aggregation. The 100 µg pack size is sufficient for approximately 100 Western blots at 1:1000 dilution. After the first thaw, an aliquot remains stable at 4°C for up to one month if supplemented with 0.02-0.05 percent sodium azide, though we prefer frozen storage for longest activity. Do not store diluted antibody in blocking buffer for more than one week. Typical shelf life is 12-24 months from date of manufacture when stored properly. Check for precipitates before use and clarify by brief centrifugation if needed.

Why do I see multiple bands for DPP-6 in my Western blot?

Multiple bands typically arise from differential glycosylation, alternative splice isoforms, or proteolytic processing. DPP-6 undergoes complex N-glycosylation that produces heterogeneous mobility. At least two major splice variants exist with different N-terminal lengths, which may be expressed in a tissue-specific manner. In some preparations, you may detect a cleaved extracellular fragment around 80-90 kDa resulting from ectodomain shedding by metalloproteinases. To confirm specificity, compare your pattern to published literature for your tissue type, include a knockout or siRNA knockdown control, or treat lysates with deglycosylase to collapse glycoforms into a single band.