Anti-PRSS-16 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight should I expect for PRSS-16 on a Western blot?

PRSS-16 (UniProt Q9NQE7) is a 514-amino acid serine protease with a predicted molecular weight of approximately 57 kDa based on its primary sequence. On reducing SDS-PAGE, you should observe a band near this size in thymic epithelial lysates or transfected cell lines. Keep in mind that post-translational modifications, including glycosylation, may shift the apparent mobility slightly higher. If you see additional lower-mass bands, consider the possibility of proteolytic processing or degradation, particularly given PRSS-16's localization to cytoplasmic vesicles where protease activity is common.

Which tissues or cell lines are best for positive controls when testing PRSS-16 antibody?

PRSS-16 is expressed predominantly in thymic epithelium, making human thymus tissue lysate the most physiologically relevant positive control. If primary tissue is unavailable, consider thymic epithelial cell lines or thymic epithelial tumor samples, where PRSS-16 has been reported as a tissue-restricted marker. For recombinant controls, lysates from HEK293 or similar cells transiently transfected with full-length human PRSS-16 work well. Negative controls should include non-thymic tissues such as liver, skeletal muscle, or peripheral blood lymphocytes, where PRSS-16 expression is minimal or absent.

What is the recommended starting dilution for PRSS-16 antibody in Western blot?

For Western blot, start at 1:1000 dilution in blocking buffer (typically five percent non-fat dry milk or BSA in TBST). This rabbit polyclonal has been validated at this dilution for human samples. If signal is weak in your particular lysate, you can titrate up to 1:500, though higher concentrations may increase background. Conversely, if background is problematic, extend the dilution to 1:2000. Always include a no-primary-antibody control lane to assess non-specific binding of your secondary reagent, and optimize incubation time and blocking conditions for your specific sample type.

Is PRSS-16 antibody likely to work in mouse or rat samples?

This antibody has been validated for human PRSS-16. Cross-reactivity with mouse or rat orthologues is predicted but not experimentally validated by Triple Point Biologics. Human PRSS-16 shares moderate sequence homology with rodent orthologues, so cross-reactivity is plausible, particularly if the epitope region is conserved. If you plan to use mouse or rat thymus lysates, treat the antibody as experimental: run a pilot blot with recombinant human PRSS-16 or human thymus as a positive control on the same membrane, and verify that any rodent bands align with the expected molecular weight and tissue distribution before drawing conclusions.

Can I use this PRSS-16 antibody for immunohistochemistry or immunofluorescence?

Triple Point Biologics antibodies are typically validated for Western blot and immunohistochemistry. While this specific lot has confirmed validation for Western blot at 1:1000, IHC and immunofluorescence applications are feasible but may require optimization. For IHC on formalin-fixed paraffin-embedded thymus sections, start with antigen retrieval using citrate buffer (pH 6.0) and test dilutions between 1:100 and 1:500. For immunofluorescence on fixed cells or cryosections, begin at 1:200. Include appropriate blocking steps and verify specificity with peptide competition or secondary-only controls, especially given PRSS-16's vesicular localization pattern.

Does PRSS-16 have multiple isoforms I should be aware of?

The UniProt entry for human PRSS-16 (Q9NQE7) lists a canonical 514-amino acid isoform as the primary sequence. Alternative splicing has not been extensively characterized for PRSS-16 in the literature, and no well-documented shorter isoforms are commonly reported. If you observe multiple bands on a Western blot, consider post-translational processing rather than splice variants—PRSS-16 is a serine protease localized to vesicles, where autocatalytic cleavage or limited proteolysis may occur. If isoform analysis is critical to your study, confirm band identity by mass spectrometry or siRNA knockdown rather than assuming splice variation.

What are the optimal storage conditions for this antibody?

Store the antibody at -20°C in small aliquots to avoid repeated freeze-thaw cycles, which can reduce titer and increase aggregation. The 100-microgram pack size is convenient for aliquoting upon arrival—consider 10-microliter single-use aliquots if you run blots infrequently. Once thawed, an aliquot can be held at 4°C for up to one month if kept sterile and supplemented with 0.02 percent sodium azide as a preservative, though fresh aliquots generally give the most consistent results. Do not store diluted antibody in blocking buffer for more than a few days, as proteolytic degradation and microbial growth can compromise performance.

What sample preparation steps are critical for detecting PRSS-16 by Western blot?

Because PRSS-16 localizes to cytoplasmic vesicles in thymic epithelial cells, effective lysis is essential. Use a strong detergent buffer such as RIPA with protease inhibitor cocktail to ensure complete solubilization of vesicular compartments. Sonication or multiple freeze-thaw cycles can improve yield from tissue samples. Load 20 to 50 micrograms of total protein per lane—PRSS-16 is relatively tissue-restricted, so lower-abundance samples may require higher loading or enrichment. Run samples under reducing conditions with beta-mercaptoethanol or DTT to fully denature the protein. If using thymic tissue, mechanical homogenization prior to lysis will improve consistency.