Anti-DPP-4 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight should I expect for DPP-4 on Western blot?

DPP-4 migrates as a band around 110 kDa on reducing SDS-PAGE, reflecting the full-length 766-residue type II transmembrane glycoprotein with extensive N-glycosylation. The predicted molecular weight of the polypeptide backbone is approximately 88 kDa, but post-translational glycosylation increases the apparent size. You may also detect a soluble circulating form around 110 kDa in serum or culture supernatants, produced by proteolytic shedding of the ectodomain. Start with the recommended 1:1000 dilution in TBST with 5% non-fat milk and optimize exposure time to capture both membrane-bound and soluble pools if present.

Is this DPP-4 antibody specific for the CD26 T-cell activation antigen?

Yes, DPP-4 and CD26 are the same protein. The antibody recognizes dipeptidyl peptidase 4, which is designated CD26 when functioning as a T-cell costimulatory receptor. The molecule is identical whether it is cleaving incretin hormones in the gut epithelium or binding adenosine deaminase on activated T lymphocytes. The antibody is validated in human samples where DPP-4 is expressed on activated T cells, endothelial cells, and epithelial surfaces. If you are studying T-cell activation, confirm expression levels in your stimulation model, as CD26 upregulation is antigen-dependent and may require 24-72 hours post-activation.

Does this antibody work in rat and mouse samples?

Human reactivity is validated; rat cross-reactivity is predicted based on epitope homology but not experimentally confirmed in-house. Mouse (Mus musculus) is not listed, which typically indicates lower sequence conservation in the immunogen region. Human and rat DPP-4 share approximately 85-90% identity, so cross-reactivity is plausible. We recommend empirical titration starting at 1:500 in rat lysates and running a positive control human lysate in parallel. If you require guaranteed mouse reactivity, request lot-specific validation data or consider a mouse-validated clone. Pan and monkey reactivities are also predicted and should be pilot-tested before committing to large experiments.

What starting dilution should I use for immunofluorescence with this DPP-4 antibody?

Although the recommended Western blot dilution is 1:1000, immunofluorescence typically requires more concentrated antibody. Start at 1:100 to 1:200 for cell monolayers or frozen sections, using PBS with 1-3% BSA and 0.1-0.3% Triton X-100 for permeabilized samples or omitting detergent for surface CD26 staining on intact cells. DPP-4 is a type II membrane protein with a large extracellular domain, so non-permeabilized staining is effective for visualizing surface expression on T cells or endothelium. Include a no-primary-antibody control and optimize incubation time between one hour at room temperature and overnight at four degrees Celsius.

What positive control lysate should I use to validate DPP-4 antibody performance?

Caco-2 human colorectal adenocarcinoma cells express high endogenous DPP-4 and serve as an excellent positive control. Alternatively, use lysates from activated human peripheral blood T cells, Jurkat T lymphocytes, or HUVEC endothelial cells, all of which express CD26. For negative controls, consider DPP-4 knockout cell lines if available, or pre-absorb the antibody with recombinant DPP-4 ectodomain to confirm specificity. Load 20-30 micrograms total protein per lane and expect a prominent 110 kDa band. If studying soluble DPP-4, human serum diluted 1:50 in sample buffer provides a physiological positive control for the circulating protease form.

Can I detect both membrane-bound and soluble DPP-4 with this antibody?

Yes, this polyclonal antibody recognizes epitopes present in both the membrane-anchored and soluble circulating forms of DPP-4. The soluble form is generated by proteolytic cleavage near the membrane anchor, releasing the catalytically active ectodomain into serum and other biological fluids. On Western blot, both forms migrate around 110 kDa and are indistinguishable by size alone. To differentiate them experimentally, fractionate samples into membrane and cytosolic fractions or compare whole-cell lysates with conditioned media or serum. For IHC, membrane staining appears as sharp surface or intercellular boundaries, whereas soluble DPP-4 may produce diffuse extracellular signal depending on fixation.

How should I store this antibody and what is the expected stability?

Store the antibody at minus twenty degrees Celsius in small aliquots to avoid repeated freeze-thaw cycles, which can reduce titre and increase aggregation. The 100 microgram pack size is suitable for dividing into 10-20 microliter aliquots upon receipt. For short-term use over one to two weeks, diluted working stocks can be held at four degrees Celsius with 0.02-0.05% sodium azide as preservative, though neat stock should remain frozen. Rabbit polyclonal antibodies from Triple Point Biologics typically retain activity for two to three years under these conditions. Avoid prolonged exposure to room temperature and ensure the vial is tightly sealed to prevent lyophilization or concentration changes. Centrifuge briefly before each use to pellet any aggregates.

Will this DPP-4 antibody detect the enzyme in its active conformation?

This polyclonal antibody is raised against a recombinant fragment and recognizes linear or denatured epitopes, making it well-suited for Western blot and IHC where protein structure is partially or fully disrupted. It is not conformation-dependent and will detect DPP-4 regardless of enzymatic activity status or inhibitor binding. If you need to correlate antibody signal with catalytic activity, pair immunoblotting with an enzymatic assay using a fluorogenic dipeptide substrate such as Gly-Pro-AMC. The antibody does not distinguish between active enzyme and catalytically inactive mutants or inhibitor-bound complexes. For native immunoprecipitation or flow cytometry, empirical testing is required since epitope accessibility may differ in the folded extracellular domain.