Anti-PCSK9 Rabbit Polyclonal Antibody

Frequently Asked Questions

What molecular weight band should I expect for PCSK9 on a Western blot?

PCSK9 migrates as a ~75 kDa band on reducing SDS-PAGE, corresponding to the mature cleaved form. The full-length precursor is 692 amino acids (~75 kDa calculated), but PCSK9 undergoes autocatalytic cleavage in the ER, removing its prosegment while remaining noncovalently associated. You may occasionally see the uncleaved zymogen at a similar molecular weight or degradation products under ~60 kDa depending on sample handling. Use liver lysate or HepG2 cells as positive controls; PCSK9 is highly expressed in hepatocytes and secreted, so serum samples also work well.

Does this PC-9 antibody work for mouse and rat samples or only human?

This antibody is validated for human PCSK9. Mouse and rat cross-reactivity are predicted based on sequence homology—human PCSK9 shares approximately 80 percent identity with mouse and rat orthologs—but we have not formally validated performance in these species. If you are working with rodent samples, start with the recommended 1:1000 dilution and run a positive control (mouse liver lysate) alongside your samples. Titre the antibody if background is high or signal is weak. We recommend confirming specificity with a blocking peptide or siRNA knockdown in your model system.

What is the recommended starting dilution for Western blot with this PC-9 antibody?

Start at 1:1000 dilution in your blocking buffer for Western blot. This polyclonal antibody has been validated at this concentration for detecting endogenous PCSK9 in human liver and hepatoma cell lysates. Incubate overnight at 4°C with gentle agitation. If signal is weak or you are working with low-expression samples, you can increase concentration to 1:500. Conversely, if background is high, dilute further to 1:2000. Use 5 percent non-fat milk or BSA in TBST for blocking; both work well with this antibody in our hands.

Can I use this antibody for immunohistochemistry or immunofluorescence on tissue sections?

Yes, Triple Point Biologics antibodies are typically validated for IHC and immunofluorescence in addition to Western blot. For IHC on paraffin-embedded sections, use antigen retrieval with citrate buffer (pH 6.0) and start with a 1:100 to 1:200 dilution. PCSK9 localizes primarily to hepatocytes in liver tissue and is also expressed in kidney and intestine. For immunofluorescence, begin with 1:100 dilution on fixed cells or cryosections. Validate staining specificity by comparing signal in PCSK9-expressing tissues (liver) versus low-expression controls (skeletal muscle).

What sample types and lysis conditions work best for detecting PCSK9?

PCSK9 is highly expressed in liver and readily detected in hepatocyte-derived cell lines like HepG2 or Huh7. Use RIPA buffer with protease inhibitors for total cell lysates; PCSK9 is both intracellular and secreted, so you can detect it in conditioned media as well. For serum or plasma samples, dilute 1:10 to 1:50 in sample buffer and load 2 to 5 microliters per lane. Avoid repeated freeze-thaw cycles, as PCSK9 stability can be compromised. If detecting secreted PCSK9, concentrate conditioned media by TCA precipitation or ultrafiltration to improve sensitivity.

Does PCSK9 have isoforms or splice variants I should be aware of?

PCSK9 exists primarily as a single functional isoform in humans. The protein undergoes autocatalytic cleavage between Gln152 and Ser153, separating the prosegment from the catalytic domain, but both fragments remain noncovalently associated in the mature secreted form. You will not see distinct isoform bands on a Western blot. However, post-translational modifications and partial degradation can produce minor species. Some commercial PCSK9 preparations contain recombinant furin-cleaved variants, so if you are using purified protein as a standard, verify the form provided by the supplier to match your experimental context.

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

Store the antibody at -20°C as supplied. Avoid repeated freeze-thaw cycles by making single-use aliquots upon receipt; divide the 100 microgram vial into 10 to 20 microliter aliquots in cryovials. Once thawed, an aliquot can be kept at 4°C for up to two weeks for convenience during active experiments. For long-term storage beyond six months, keep at -80°C. Do not add sodium azide if you plan to use the antibody for applications other than Western blot, as it can interfere with HRP-based detection in some protocols. Properly stored, polyclonal antibodies remain stable for several years.

What controls should I include to confirm specificity for PCSK9?

Include a positive control with high endogenous PCSK9 expression, such as HepG2 whole-cell lysate or human liver tissue lysate. As a negative control, use a tissue with low or absent PCSK9 expression like skeletal muscle or pre-treat cells with PCSK9-targeting siRNA to demonstrate signal reduction. A blocking-peptide control is valuable: pre-incubate the antibody with the immunizing peptide (if available from the supplier) and show loss of signal. If working with overexpression systems, compare vector-transfected versus PCSK9-transfected lysates. These controls collectively confirm that the observed band is specific to PCSK9.