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- Table of Contents
SRM / Spermidine synthase · Western blot design guide
Design a Western Blot for SRM
Real validated SRM Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-SRM 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
SRM 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 | ~33.8 kDa | |
| Observed band | ~34 kDa | |
| Gel | 5–20% (catalog A01594-2) | |
| Positive control | Appendix (IHC candidate; verify WB) +4 more | |
| Negative control | Bone marrow (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Acetylated | |
| Caveat | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 1 isoform(s) |
Section 1
Real Curated SRM Western Blot Protocols
The A01594-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human Hela, human HepG2, human PC-3 (catalog A01594-2) |
| Gel % | 5–20% (catalog A01594-2) |
| Load | 30 ug; reducing conditions (catalog A01594-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A01594-2) |
| Membrane | nitrocellulose membrane (catalog A01594-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A01594-2) |
| Primary antibody | A01594-2 · 0.5 μg/mL (catalog A01594-2) |
| Primary incubation | overnight at 4°C (catalog A01594-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A01594-2) |
| Secondary incubation | 1.5 hour at RT (catalog A01594-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A01594-2) |
| Detection | ECL (catalog A01594-2) |
Section 2
What Is the Expected SRM Western Blot Band Size?
SRM is predicted at 33.8 kDa and observed near 34 kDa; the small difference has no established cause in the supplied evidence.
| Band near 34 kDa | Matches the observed SRM band and its 33.8 kDa predicted monomer mass |
| Higher band near twice the monomer size | Could reflect retained SRM homodimers; confirm its identity |
| Higher band near four times the monomer size | Could reflect retained SRM homotetramers; confirm its identity |
| Single sharp band near 34 kDa | Compatible with SRM despite N-acetylmethionine at residue 1, which does not establish a visible shift |
💡Expected SRM appearanceSRM has a predicted monomer mass of 33.8 kDa and an observed band at approximately 34 kDa; confirm band identity with an appropriate control.
| UniProt predicted monomer mass | Places the expected monomer near 33.8 kDa, consistent with the observed 34 kDa band |
| 302-residue SRM chain | Defines the full-length protein underlying the predicted monomer mass |
| SRM homodimer | Could yield a higher band if the complex survives sample preparation |
| SRM homotetramer | Could yield a still higher band if the complex survives sample preparation |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | SRM may be below detection or the assay may have failed | Check loading and detection with a positive whole-cell lysate |
| Band higher than expected | An SRM oligomer may have persisted through sample preparation | Compare denaturing conditions and verify band identity |
| Band lower than expected | Band identity or sample integrity is uncertain | Check a positive control and repeat with fresh sample |
| Multiple bands | Retained oligomers or unrelated antibody binding may contribute | Compare denaturing conditions and use a band-identity control |
| Weak or no signal | Insufficient SRM or ineffective detection may limit signal | Check loading, antibody conditions, and a positive whole-cell lysate |
Experiment evidence
Western blot images showing SRM band
A01594-2 · SRM
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for SRM Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for SRM in Western blot, you can use appendix tissue, which HPA rates as high for SRM.
Positive control: Appendix (IHC candidate; verify WB)
Negative control: Bone marrow (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: HPA identifies both high-expression and not-detected tissues, making tissue controls feasible.
HPA tissue expression evidence for SRM
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 |
|---|---|---|---|---|
| Appendix | glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | neuronal cells | High | Protein (IHC) | HPA → |
| Cerebellum | cells in granular layer | 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 |
|---|---|---|---|---|
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Ovary | ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | smooth muscle cells | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | squamous epithelial cells | Low | Protein (IHC) | HPA → |
| Soft tissue | chondrocytes | Low | Protein (IHC) | HPA → |
Section 3
Advanced SRM Western Blot Tips
Deeper troubleshooting and optimisation questions for SRM, answered from its protein features.
How should SRM 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 SRM isoforms explain multiple bands?
Isoforms · The supplied record lists one isoform and no alternative sequence, so it provides no isoform-based explanation for multiple bands.
Which SRM modification should I consider when interpreting bands?
PTM · UniProt lists N-acetylmethionine at position 1, using UniProt sequence numbering. This modification alone does not establish a visible shift from the predicted 33.8 kDa mass.
Does this guide establish induction of SRM?
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.
What transfer method to use for SRM Western blot?
Transfer · Choose and verify transfer conditions suitable for a protein near 34 kDa. Check the membrane and residual gel to assess transfer; the supplied features do not specify a particular transfer method.
How should blocking be optimized?
Blocking · Standard workflow guidance: follow the A01594-2 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 the SRM band?
Quantitation · Quantify the band near 34 kDa within the linear detection range and normalize for loading. Apply the same band boundary across samples; the supplied features do not identify another isoform band to include.
Why does SRM appear near 34 kDa?
Interpretation · The observed band near 34 kDa closely matches the predicted 33.8 kDa mass of the 302-residue protein. The listed features do not establish a band shift.
SRM is reported as a homodimer or homotetramer, but that does not establish that higher bands on a Western blot are oligomers. Compare the higher bands with the expected ~34 kDa band and verify their identity before assigning them.
SRM is reported as a homodimer or homotetramer, but that does not establish that higher bands on a Western blot are oligomers. Compare the higher bands with the expected ~34 kDa band and verify their identity before assigning them.
Boster reagents
SRM 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-Spermidine synthase/SRM Antibody Picoband®
Cat # A01594-2
The catalog reports one anti-SRM antibody, A01594-2, with human reactivity. Its Western blot image shows an approximately 34 kDa band in HeLa, HepG2, and PC-3 whole-cell lysates. No independent validation evidence is supplied.
Which to pick: A01594-2 is the only listed option. Its Western blot example uses human HeLa, HepG2, and PC-3 lysates and a 0.5 μg/mL primary antibody concentration; suitability for other samples is not established here.
Source: BosterBio SRM gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P19623.
- Human Protein Atlas. SRM tissue expression.
- PMC4873969 — target-verified WB comparison
- PMC12236739 — target-verified WB comparison