Anti-Polyserase-1 Rabbit Polyclonal Antibody

Frequently Asked Questions

What bands should I expect to see on Western blot with the Polyserase-1 antibody?

Polyserase-1 undergoes post-translational processing to generate three separate serine protease domains: Serase-1, Serase-2, and Serase-3. On Western blot, you may observe the full-length 1,059-residue precursor (approximately 116 kDa) along with processed fragments corresponding to individual Serase domains, typically in the 30-40 kDa range depending on glycosylation state. Band patterns vary by cell type and activation status. Using lysates from cells known to express TMPRSS9, such as certain epithelial lines, will help you distinguish specific signal from background. Always include a negative control lysate from a TMPRSS9-null cell line.

Does the Polyserase-1 antibody recognize all three Serase domains or just one?

The epitope mapping for this polyclonal antibody determines which processed forms it detects. Rabbit polyclonals raised against a broad region of Polyserase-1 often recognize epitopes present in one or more of the Serase domains, but cross-reactivity among Serase-1, -2, and -3 is not guaranteed because they share limited sequence identity despite functional similarity. If your experimental goal requires distinguishing individual Serase subunits, confirm epitope location with the datasheet or contact technical support. For general Polyserase-1 expression profiling, detection of any processed form typically suffices to demonstrate pathway activity.

What is the recommended starting dilution for Western blot with this antibody?

The validated dilution for Western blot is 1:1,000 in blocking buffer, typically 5% non-fat milk or BSA in TBST. This ratio has been optimized during validation with human lysates. However, Polyserase-1 expression varies widely by tissue and cell type; epithelial and certain tumor lines show higher expression. If initial signal is weak, titrate down to 1:500 or concentrate your lysate. Conversely, if background is high, extend blocking time or try 1:2,000. Overnight incubation at 4°C often improves sensitivity for low-abundance transmembrane proteases compared to one-hour room-temperature protocols.

Is this Polyserase-1 antibody validated for immunohistochemistry on paraffin sections?

Yes, Triple Point Biologics antibodies are validated for immunohistochemistry, including paraffin-embedded tissue. For Polyserase-1, antigen retrieval is typically required because formalin fixation masks epitopes on transmembrane proteins. Use citrate buffer (pH 6.0) or Tris-EDTA (pH 9.0) retrieval in a pressure cooker or steamer for 20 minutes. Given Polyserase-1 localization at the plasma membrane with extracellular catalytic domains, expect membranous and sometimes cytoplasmic staining in positive cells. Start with a 1:100 to 1:200 dilution and optimize on control tissue known to express TMPRSS9, such as normal colonic or prostate epithelium.

Will this antibody cross-react with rat or monkey Polyserase-1?

Rat and monkey cross-reactivity are listed as potential, meaning sequence homology predicts recognition but formal validation has not been completed. Human and rat TMPRSS9 share approximately 70-75% identity in conserved protease domain regions, so cross-reactivity is plausible but band patterns may differ. Monkey orthologs are typically more similar to human. If working with non-human species, run a pilot Western blot alongside a validated human lysate as positive control. Adjust dilution and exposure time as needed, and consider verifying hits by qPCR or mass spectrometry if this is a critical experiment.

What sample preparation considerations are important for detecting Polyserase-1?

Polyserase-1 is a type II transmembrane protease with active catalytic domains exposed extracellularly. Use lysis buffers containing detergent (1% Triton X-100 or NP-40) to solubilize membrane proteins efficiently. Include protease inhibitors to prevent autocatalytic degradation; Polyserase-1 has trypsin-like activity and may cleave itself or other substrates during prolonged lysis. Work quickly on ice and snap-freeze aliquots to preserve native processing patterns. Because expression is often polarized in epithelial cells, whole-cell lysates are preferable to cytosolic fractions. For surface biotinylation or co-IP experiments, verify that epitope accessibility is maintained under your capture conditions.

How should I store the Polyserase-1 antibody and how long is it stable?

Store the antibody at -20°C in single-use aliquots to avoid freeze-thaw cycles, which can reduce titer and increase aggregation. The 100 µg pack size is convenient for aliquoting 10-20 µL per tube depending on usage. Polyclonal antibodies in glycerol-containing storage buffer typically remain stable for at least two years at -20°C and up to one year at 4°C once thawed, though we recommend -20°C for long-term storage. Avoid prolonged exposure to room temperature. If you observe increased background or loss of signal over time, prepare a fresh aliquot rather than re-using a working stock repeatedly.

What positive control lysate should I use to validate Polyserase-1 antibody performance?

Select a cell line or tissue known to express TMPRSS9 at detectable levels. Published RNA-seq and proteomic databases indicate expression in differentiated epithelial cells of the gastrointestinal tract, prostate, and certain carcinoma lines. HT-29 colon adenocarcinoma or LNCaP prostate cells are reasonable starting points, though expression may be variable. If commercial control lysates are unavailable, prepare your own from confluent cultures grown under differentiation-promoting conditions. Include a TMPRSS9-negative line, such as HEK293T or NIH-3T3, as a specificity control. Comparing induced versus uninduced or siRNA-knockdown lysates provides the most rigorous validation.