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- Table of Contents
CTSA / Lysosomal protective protein · Western blot design guide
Design a Western Blot for CTSA
Real validated CTSA Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CTSA 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
CTSA Western Blot Experimental Design Guide
Expected bands, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before running
| Expected band | ~54.5 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Adrenal gland (IHC candidate; verify WB) +4 more | |
| Negative control | Suggested KO / knockdown lysate |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Glycosylated + Cleaved | |
| Caveat | Chain-specific detection | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated CTSA Western Blot Protocols
The A02440 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | A549, sp2/0, PC12 (catalog A02440) |
| 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% milk or 5% BSA in TBST (standard starting point) |
| Primary antibody | A02440; 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 CTSA Western Blot Band Size?
CTSA has a predicted 54.5 kDa precursor and disulfide-linked 32 and 20 kDa chains; glycosylation, cleavage, and isoforms could affect migration, but no Western-blot position is demonstrated.
| Band near 54.5 kDa | Candidate full-length CTSA precursor; confirm its identity with antibody controls |
| Bands near 32 and 20 kDa under reducing conditions | Possible separation of the disulfide-linked CTSA chains |
| A larger band under non-reducing conditions | Possible retention of the linked 32 and 20 kDa chains |
| Band below the precursor position | Could reflect signal-peptide removal; this alone does not explain the 32 and 20 kDa chains |
| Band above the predicted precursor position | N-linked glycosylation at Asn145 or Asn333 could contribute, but a visible shift is unproven |
| Multiple bands of different sizes | Isoforms 1 and 2 are possible contributors, though distinct migration is unproven |
💡Expected CTSA appearanceUniProt predicts a 54.5 kDa precursor and identifies disulfide-linked 32 and 20 kDa chains; no empirical Western-blot band size is supplied, so confirm band identity and processing experimentally.
| 54.5 kDa predicted precursor mass | Reference size for the full-length sequence, not a validated blot position |
| N-linked glycosylation at Asn145 | Could increase apparent size; the magnitude and visibility are unknown |
| N-linked glycosylation at Asn333 | Could increase apparent size; the magnitude and visibility are unknown |
| Disulfide-linked 32 and 20 kDa heterodimer | May remain linked without full reduction and separate into chains upon reduction |
| Isoforms 1 and 2 | May differ in size, but distinct band positions are not supplied |
| Signal peptide at residues 1–28 | Removal makes the cleaved product smaller than the full-length precursor |
| Situation | Likely cause | Next action |
|---|---|---|
| Band higher than expected | Incomplete reduction may retain the disulfide-linked chains; N-linked glycosylation may also affect migration | Compare reducing and non-reducing samples, and verify identity with an independent antibody |
| Band lower than expected | CTSA has a cleavable signal peptide and forms 32 and 20 kDa chains | Check which CTSA region the antibody recognizes and compare expected chain positions |
| Broad smear instead of sharp band | Variable N-linked glycosylation is possible but not established by the listed sites | Compare deglycosylated and untreated samples and use a band-identity control |
| Multiple bands | Disulfide-linked chains, precursor processing, or isoforms 1 and 2 may contribute | Compare reducing conditions and confirm bands with an antibody to another CTSA region |
| Weak or no signal | CTSA is lysosomal, so sample preparation or antibody recognition of the detected form may limit signal | Check lysosomal protein recovery, antibody epitope, and a positive-control lysate |
| Fragments below expected size | The reported CTSA complex contains 32 and 20 kDa chains | Test whether the bands track with CTSA using an independent antibody and reducing conditions |
Experiment evidence
Western blot images showing CTSA band
A02440 · CTSA
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CTSA Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CTSA in Western blot, you can use adrenal gland tissue.
Positive control: Adrenal gland (IHC candidate; verify WB)
Negative control: Suggested KO / knockdown lysate
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: No tissue is reported as not detected, so use siRNA knockdown or a KO line for a negative control.
HPA tissue expression evidence for CTSA
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 |
|---|---|---|---|---|
| Adrenal gland | glandular cells | High | Protein (IHC) | HPA → |
| Appendix | glandular cells | High | Protein (IHC) | HPA → |
| Caudate | neuronal cells | High | Protein (IHC) | HPA → |
| Cervix | glandular cells | High | Protein (IHC) | HPA → |
| Colon | glandular cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | adipocytes | Low | Protein (IHC) | HPA → |
| Esophagus | squamous epithelial cells | Low | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Low | Protein (IHC) | HPA → |
| Smooth muscle | smooth muscle cells | Low | Protein (IHC) | HPA → |
| Soft tissue | fibroblasts | Low | Protein (IHC) | HPA → |
Section 3
Advanced CTSA Western Blot Tips
Deeper troubleshooting and optimisation questions for CTSA, answered from its protein features.
How should CTSA band migration be interpreted?
Band shift · Use the separately labelled calculated mass and catalog-observed evidence above. A sequence annotation does not establish an observed migration shift. Verify target identity with orthogonal controls.
Could CTSA isoforms produce different bands?
Isoforms · UniProt lists isoforms 1 and 2. Isoform 2 lacks residues 102–118 relative to the canonical sequence; these are UniProt canonical coordinates. This sequence difference could affect band assignment, but the features do not establish whether the isoforms resolve on your blot.
Which CTSA glycosylation sites matter when interpreting migration?
PTM · UniProt lists N-linked glycosylation at Asn145 and Asn333, using canonical sequence coordinates. Glycosylation may affect migration, but the listed sites alone do not demonstrate a visible shift or explain a measured mass difference.
Does this guide establish induction of CTSA?
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 CTSA?
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 A02440 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 I quantify CTSA if multiple bands appear?
Quantitation · Define which band represents the species your antibody detects before quantifying it. CTSA is reported as a 32 kDa/20 kDa heterodimer and has two isoforms. Quantify the same assigned band across samples; do not assume every CTSA-related band measures the same species.
Why might CTSA bands differ from its predicted 54.5 kDa mass?
Interpretation · The 54.5 kDa prediction describes the full-length sequence. CTSA has a signal peptide at residues 1–28, two N-linked glycosylation sites, and a reported 32 kDa/20 kDa disulfide-linked heterodimer. These features guide band assignment, but none establishes the apparent mass of a particular band.
UniProt describes CTSA as a disulfide-linked heterodimer of a 32 kDa chain and a 20 kDa chain. Check whether the antibody detects either chain before assigning a band; the supplied features do not identify its epitope.
Because the reported 32 kDa and 20 kDa chains are disulfide-linked, compare reducing and nonreducing samples when assigning bands. CTSA also has four listed disulfides. The features do not specify the exact gel pattern under either condition.
Check its position against the full-length 54.5 kDa prediction and the reported 32 kDa and 20 kDa chains. Consider the 1–28 signal peptide, isoform 2 deletion at canonical residues 102–118, N-linked sites at Asn145 and Asn333, and disulfide-linked chains. These features suggest possibilities but cannot identify a band by themselves.
UniProt describes CTSA as a disulfide-linked heterodimer of a 32 kDa chain and a 20 kDa chain. Check whether the antibody detects either chain before assigning a band; the supplied features do not identify its epitope.
Because the reported 32 kDa and 20 kDa chains are disulfide-linked, compare reducing and nonreducing samples when assigning bands. CTSA also has four listed disulfides. The features do not specify the exact gel pattern under either condition.
Check its position against the full-length 54.5 kDa prediction and the reported 32 kDa and 20 kDa chains. Consider the 1–28 signal peptide, isoform 2 deletion at canonical residues 102–118, N-linked sites at Asn145 and Asn333, and disulfide-linked chains. These features suggest possibilities but cannot identify a band by themselves.
Boster reagents
CTSA 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-Lysosomal protective protein CTSA Antibody
Cat # A02440
A02440 is a polyclonal anti-CTSA antibody listed for human, mouse, and rat. Its WB image shows A549, sp2/0, and PC12 whole-cell lysates at 1:500 dilution. These examples document tested samples; no publication evidence is supplied.
Which to pick: A02440 is the only listed CTSA antibody. It has a WB image using A549, sp2/0, and PC12 lysates at 1:500; check whether those sample types and its listed reactivity fit your experiment.
Source: BosterBio CTSA gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P10619.
- Human Protein Atlas. CTSA tissue expression.
- PMC7185279 — target-verified WB comparison
- PMC13185071 — target-verified WB comparison
- PMC12929657 — target-verified WB comparison