Anti-ADAM-7 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight should I expect for ADAM-7 on a Western blot?

ADAM-7 migrates at approximately 84–90 kDa on reducing SDS-PAGE, consistent with the 754-residue glycoprotein GP-83. The apparent molecular weight may exceed the calculated mass due to post-translational glycosylation. In testicular and epididymal lysates, you may observe additional bands representing proteolytic processing fragments or differentially glycosylated forms. Non-reducing conditions are not typically required, as ADAM-7 is a single-chain transmembrane protein. If you detect multiple bands, consider treating samples with PNGase F to assess the contribution of N-linked glycans to the observed heterogeneity.

Does ADAM-7 have catalytic metalloproteinase activity I need to account for in my experiments?

No. ADAM-7 is classified as a non-catalytic member of the ADAM family and lacks functional metalloproteinase activity, distinguishing it from proteolytically active relatives like ADAM10 or ADAM17. The protein retains the disintegrin and cysteine-rich domains but does not cleave substrates. This is important when designing experiments: ADAM-7 functions primarily through protein-protein interactions and structural roles in the epididymis rather than enzymatic activity. If you are studying proteolytic pathways, ADAM-7 serves a regulatory or scaffolding role rather than acting as a sheddase or ectodomain protease.

What starting dilution should I use for Western blot with this ADAM-7 antibody?

Start at 1:1000 dilution in blocking buffer for overnight incubation at 4°C, which is the validated dilution for this polyclonal. Depending on ADAM-7 expression levels in your tissue or cell line, you may need to optimize between 1:500 and 1:2000. Epididymal tissues and mature spermatozoa express ADAM-7 at higher levels than most somatic tissues, so lysates from testis, caput epididymis, or capacitated sperm typically yield strong signals at 1:1000. For cell lines with lower expression, increase antibody concentration or load more total protein per lane before exceeding 1:500.

Will this antibody detect mouse or rat ADAM-7 in my samples?

Cross-reactivity with mouse and rat ADAM-7 is predicted based on sequence homology but not yet validated in-house. Human ADAM-7 shares approximately 70–75 percent identity with rodent orthologs, suggesting the epitope may be conserved. If you are working with mouse or rat samples, treat this as an optimization experiment: test the antibody at 1:500 and 1:1000 dilutions with epididymal lysates as a positive control. Include a validated human sample in parallel if possible. Document the banding pattern and confirm specificity with a blocking peptide or siRNA knockdown before drawing conclusions.

What positive control tissue should I use for ADAM-7 Western blots?

Human epididymal tissue lysate is the ideal positive control, as ADAM-7 is abundantly expressed in the caput epididymis where it maintains epithelial architecture and supports sperm maturation. If epididymal tissue is unavailable, use lysates from mature human spermatozoa or whole testis homogenate, though signal intensity will be lower in testis. Avoid using general somatic tissues like liver, brain, or kidney, where ADAM-7 expression is minimal or absent. Capacitated sperm preparations can also serve as positive controls, particularly when investigating tyrosine phosphorylation pathways or flagellar protein interactions.

Can I use this ADAM-7 antibody for immunohistochemistry on paraffin sections?

Yes, This antibody is validated for Western blot; IHC application data is in progress. For paraffin-embedded sections, perform antigen retrieval using citrate buffer pH 6.0 with heat-induced epitope retrieval in a pressure cooker or steamer for optimal results. ADAM-7 localizes to the apical membrane of epididymal epithelial cells and the surface of maturing spermatozoa, so expect membrane and cytoplasmic staining patterns in these tissues. Start with a 1:100 to 1:200 dilution and optimize based on background. Include testis or epididymis sections as positive controls and non-reproductive tissues as negatives to confirm specificity.

How should I store this antibody to maintain long-term stability?

Store the antibody at –20°C in single-use aliquots to avoid repeated freeze-thaw cycles, which can reduce titer and increase aggregation. If you use the antibody frequently, a working aliquot can be kept at 4°C with 0.02–0.05 percent sodium azide as preservative for up to one month. Do not store diluted antibody in blocking buffer for more than one week. For long-term archiving beyond six months, –80°C storage is preferable. Always centrifuge thawed aliquots briefly before use to pellet any aggregates that may have formed during storage.

Why am I seeing high background with this polyclonal ADAM-7 antibody?

Polyclonal antibodies can produce background if blocking is insufficient or if non-specific IgG binds to Fc receptors in certain tissues. Increase blocking time to two hours at room temperature using five percent non-fat dry milk or BSA in TBST. For IHC, preincubate sections with serum from the secondary antibody host species. High background in testis or epididymis may reflect abundant immunoglobulin or serum proteins in the interstitial compartment; in that case, include a no-primary-antibody control and consider using an IgG-depleted lysate preparation or affinity-purified secondary antibodies to reduce non-specific signal.