Anti-Serpin-A5 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight should I expect for Serpin-A5 on a Western blot?

Serpin-A5 migrates at approximately 57 kDa on reducing SDS-PAGE, slightly above its predicted molecular weight of 46 kDa due to N-linked glycosylation at three consensus sites. You may observe a diffuse or doublet band reflecting glycosylation heterogeneity, particularly in plasma samples. If you treat lysates with PNGase F or other deglycosylation enzymes, the band should sharpen and shift toward 46 kDa. Some tissues and cell lines secrete less heavily glycosylated forms, so band appearance can vary by sample source.

What dilution should I start with for Western blot and IHC with this Serpin-A5 antibody?

For Western blot, we recommend starting at 1:1000 dilution in blocking buffer, which works well for plasma and conditioned media samples where Serpin-A5 is abundant. For IHC on formalin-fixed paraffin-embedded tissue, begin with 1:200 to 1:500 after antigen retrieval in citrate buffer. Serpin-A5 expression is relatively high in liver, kidney, and reproductive tissues, so you may be able to push dilutions higher. Always include a no-primary control and titrate against your specific sample type to balance signal and background.

Does this antibody recognize both free Serpin-A5 and protease-complexed forms?

This polyclonal antibody is raised against a broad epitope spanning residues of the Serpin-A5 protein and should recognize both free inhibitor and covalent serpin-protease complexes. In plasma samples analyzed by non-reducing Western blot, you may detect higher-molecular-weight bands representing Serpin-A5 bound to activated protein C, thrombin, or other target proteases. These complexes are SDS-stable but dissociate under reducing conditions. If you are specifically studying complex formation, run samples non-reduced and expect bands above 100 kDa depending on the protease partner.

Is this Serpin-A5 antibody validated for non-human primate samples?

The antibody is validated for human Serpin-A5 and shows potential cross-reactivity with monkey samples based on sequence homology, but we have not formally validated it in Old World or New World primate species. Human and cynomolgus macaque Serpin-A5 share approximately 95 percent amino acid identity, so cross-reactivity is likely. If you are working with non-human primate plasma or tissue, we recommend running a pilot Western blot alongside human positive control lysate to confirm signal at the expected molecular weight before committing to larger experiments.

What sample types are best for detecting endogenous Serpin-A5 expression?

Serpin-A5 is most abundant in human plasma, where it circulates at 4 to 5 micrograms per milliliter, and in seminal plasma at even higher concentrations. Liver hepatocytes are the primary biosynthetic source, so liver lysates serve as excellent positive controls. You will also detect Serpin-A5 in kidney, placenta, and male reproductive tissues. For cell culture work, HepG2 and other hepatoma lines secrete Serpin-A5 into conditioned medium. Urine contains lower levels, so concentrate samples by ultrafiltration or immunoprecipitation if analyzing urinary Serpin-A5.

Can I use this antibody to distinguish Serpin-A5 from other serpin family members?

Yes, this antibody is raised against Serpin-A5-specific sequences and does not cross-react with closely related serpins such as alpha-1-antitrypsin, alpha-1-antichymotrypsin, or antithrombin based on our internal screens. Serpin family members share structural homology but differ in reactive center loop and heparin-binding domains. If you are working with crude plasma or tissue extracts containing multiple serpins, you should see a single clean band at 57 kDa for Serpin-A5. Always include a recombinant Serpin-A5 standard or well-characterized positive control lysate to confirm specificity in your system.

How should I store this Serpin-A5 antibody and how long is it stable?

Store the antibody at minus 20 degrees Celsius in small aliquots to avoid repeated freeze-thaw cycles, which can reduce titre and increase aggregation. The antibody is supplied in phosphate-buffered saline with glycerol and a preservative, stable for at least 12 months from the date of receipt when stored properly. For day-to-day use, keep a working aliquot at 4 degrees Celsius for up to one month. Do not store diluted antibody in blocking buffer for more than a few days, as carrier proteins can promote microbial growth and proteolysis.

What controls should I include when using this Serpin-A5 antibody in Western blot experiments?

Include a positive control such as human plasma diluted 1:100 in sample buffer, recombinant Serpin-A5 protein, or HepG2 cell lysate known to express Serpin-A5. Run a negative control lane with sample type expected to lack Serpin-A5, or use lysate from cells with SERPINA5 knockdown if available. A no-primary antibody control on a duplicate blot will confirm that secondary antibody is not generating off-target signal. If studying protease complexes, compare reduced and non-reduced samples side-by-side to distinguish free Serpin-A5 from high-molecular-weight complexes.