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PRKCZ / Protein kinase C zeta type · Western blot design guide
Design a Western Blot for PRKCZ
Source-linked PRKCZ Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-PRKCZ WB antibodies. Everything you need to plan the experiment before you commit precious samples.
Evidence assembled September 2026 · For research use; verify linked source records and product datasheet before use
PRKCZ Western Blot Experimental Design Guide
Expected bands, source-linked protocol options, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~67.7 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Cerebellum (total-target IHC; phospho state unverified) +4 more | |
| Negative control | Adipose tissue (total-target IHC; phospho state unverified) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated | |
| Caveat | Activation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked PRKCZ Western Blot Protocol Options
The P01796 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | HeLa cell treated with Calyculin A lysate (catalog P01796) |
| Gel % | 12–15% (standard starting point) |
| Load | 20–30 µg total protein per lane; optimize for abundance (standard starting point) |
| Transfer | Standard semi-dry transfer; verify efficiency (standard starting point) |
| Membrane | 0.45 µm PVDF (standard starting point) |
| Blocking | 5% BSA in TBST (standard starting point) |
| Primary antibody | P01796; use the WB datasheet starting dilution (standard starting point) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Species-matched HRP conjugate at validated dilution (standard starting point) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL; bracket exposures to avoid saturation (standard starting point) |
Section 2
What Is the Expected PRKCZ Western Blot Band Size?
PRKCZ has a predicted mass of 67.7 kDa; isoforms and phosphorylation are annotated, but their effects on visible migration are not established.
| Band near 67.7 kDa | Consistent with the predicted mass of PRKCZ isoform 1; band identity needs confirmation. |
| Additional bands at other sizes | Isoforms 2 and 3 are possible, but distinct migration is not established. |
| Band with the phospho-T560 antibody | Consistent with PRKCZ phosphorylated at Thr560; confirm specificity with controls. |
| Weak or absent phospho-T560 band | Thr560 phosphorylation may be low or absent even when total PRKCZ is present. |
💡Expected PRKCZ appearancePRKCZ has a predicted mass of 67.7 kDa, but no empirical band size is supplied; confirm any candidate band with total-PRKCZ and band-identity controls, especially when using the phospho-T560 antibody.
| Predicted PRKCZ mass | The 592-residue reference sequence has a calculated mass of 67.7 kDa, without a demonstrated migration position. |
| Isoform 1 | Its individual mass and migration are not supplied. |
| Isoform 2 | Sequence variation could change apparent size, but its direction and magnitude are not supplied. |
| Isoform 3 | Sequence variation could change apparent size, but its direction and magnitude are not supplied. |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | An isoform or unrelated protein could account for the band; no higher migration is established for PRKCZ. | Compare total-PRKCZ detection and PRKCZ depletion controls. |
| Band lower than expected | An isoform or unrelated protein could account for the band; no lower migration is established for PRKCZ. | Check PRKCZ depletion and compare an antibody against another epitope. |
| Multiple bands | PRKCZ has three named isoforms, but their distinct migration is unverified. | Use PRKCZ depletion and isoform-specific controls to assign bands. |
| Weak or no signal | The phospho-T560 antibody recognizes a phosphorylation state that may be scarce in the sample. | Check total PRKCZ and compare appropriately treated and untreated samples. |
| Fragments below expected size | The supplied features do not establish PRKCZ cleavage products. | Check sample integrity and test whether the bands diminish after PRKCZ depletion. |
Experiment evidence
Western blot images showing PRKCZ band
P01796 · PRKCZ
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for PRKCZ Western blot
🧪HPA-IHC candidate guidance (verify in WB): HPA tissue expression does not establish phosphorylation at the selected antibody epitope. Verify activation-state controls for the phospho-specific lead. HPA-IHC candidate guidance (verify in WB): For positive controls for PRKCZ in Western blot, you can use cerebellum lysate, which has high HPA expression.
Positive control: Cerebellum (total-target IHC; phospho state unverified)
Negative control: Adipose tissue (total-target IHC; phospho state unverified)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA reports high expression in cerebellum and no detection in adipose tissue, making tissue controls feasible.
HPA tissue expression evidence for PRKCZ
Comprehensive Human Protein Atlas IHC scoring per tissue. Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Higher expression tissues · candidate positive controls from IHC
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | GLUC cells - cytoplasm/membrane | High | Protein (IHC) | HPA → |
| Kidney | cells in glomeruli | High | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced PRKCZ Western Blot Tips
Deeper troubleshooting and optimisation questions for PRKCZ, answered from its protein features.
Where should the main PRKCZ band appear?
Band shift · The canonical sequence has a predicted mass of 67.7 kDa. No observed band position is supplied, so use that value as a starting point rather than an exact migration target. The listed phosphorylation sites alone do not establish a visible shift.
Could PRKCZ isoforms produce additional bands?
Isoforms · UniProt lists three isoforms. Isoform 2 lacks canonical residues 1–183; isoform 3 replaces canonical residues 1–112 with MLTPRTDE. These sequence differences could affect apparent size and whether an N-terminal antibody detects a band. The supplied evidence does not establish their observed migration.
Which PRKCZ phosphorylation sites matter when choosing an antibody?
PTM · UniProt lists phosphothreonine at Thr410 (by PDPK1 and PI3K), phosphothreonine at Thr560, and phosphoserine at Ser591. These are canonical UniProt coordinates; antibody or paper numbering may differ. Check that the antibody’s stated site and surrounding epitope match the isoform being measured.
UniProt assigns Thr410 phosphorylation to PDPK1 and PI3K. A phospho-Thr410 blot may therefore be relevant when examining that signaling context. The supplied features give no stimulation condition, timing, or predicted change in band intensity.
Measure a site-specific phospho signal alongside total PRKCZ and compare the phospho-to-total ratio within the same samples. Confirm that both antibodies recognize the isoform of interest. UniProt lists Thr410, Thr560, and Ser591 as modified sites but supplies no expected occupancy or response size.
UniProt assigns Thr410 phosphorylation to PDPK1 and PI3K. A phospho-Thr410 blot may therefore be relevant when examining that signaling context. The supplied features give no stimulation condition, timing, or predicted change in band intensity.
Measure a site-specific phospho signal alongside total PRKCZ and compare the phospho-to-total ratio within the same samples. Confirm that both antibodies recognize the isoform of interest. UniProt lists Thr410, Thr560, and Ser591 as modified sites but supplies no expected occupancy or response size.
Does this guide establish induction of PRKCZ?
Induction · No general induction response is established by this guide. A pathway or gene-set association is not evidence of induction in a particular specimen. Verify the relevant treatment and control in a target-specific experiment.
How should transfer be checked for PRKCZ?
Transfer · Standard workflow guidance: verify transfer efficiency for the intended target size before interpreting a weak signal. Use total-protein assessment and optimize transfer for the membrane, gel and apparatus; the labelled catalog values take precedence.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the P01796 datasheet where specified. Otherwise compare 5% milk or 5% BSA in TBST; for a phospho-specific assay start with BSA. Optimize background and specific signal with matched controls.
How should PRKCZ be quantified?
Quantitation · Standard workflow guidance: quantify only a validated target band within the linear exposure range. Use consistent sample preparation and loading, retain biological replicates, and avoid interpreting saturation or loading differences as regulation.
Could sample fractionation affect PRKCZ detection?
Interpretation · PRKCZ is listed in the cytoplasm, endosome, cell junction, and as a peripheral membrane protein. If comparing fractions, keep fraction preparation consistent and interpret changes in one fraction alongside the others; redistribution could change a fraction’s signal without establishing a change in total PRKCZ.
First compare the band with the 67.7 kDa canonical prediction and check whether the antibody epitope is present in isoforms 2 and 3. The listed phosphosites are possible sources of molecular variation, but their presence alone does not explain a mass difference or prove that any particular band is PRKCZ.
First compare the band with the 67.7 kDa canonical prediction and check whether the antibody epitope is present in isoforms 2 and 3. The listed phosphosites are possible sources of molecular variation, but their presence alone does not explain a mass difference or prove that any particular band is PRKCZ.
Boster reagents
PRKCZ Western Blot Antibodies
Catalog antibodies with Western blot application and product-specific WB images. Evaluate suitability with the reported sample, controls and experimental conditions.
Real WB data
Anti-Phospho-PKC zeta (T560) PRKCZ Rabbit Monoclonal Antibody
Cat # P01796
P01796 is a rabbit monoclonal antibody against PRKCZ phosphorylated at T560, with listed human, mouse, and rat reactivity. Its Western blot image shows Calyculin A-treated HeLa cell lysate; the supplied evidence does not show other samples or conditions.
Which to pick: P01796 is the only listed antibody and has a Western blot image in Calyculin A-treated HeLa lysate. Choose it when measuring PRKCZ T560 phosphorylation; listed mouse and rat reactivity is not demonstrated by that image.
Source: BosterBio PRKCZ gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q05513.
- Human Protein Atlas. PRKCZ tissue expression.
- PMC13385426 — target-verified WB comparison