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- Table of Contents
SEC61B / Protein transport protein Sec61 subunit beta · Western blot design guide
Design a Western Blot for SEC61B
Source-linked SEC61B Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-SEC61B 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
SEC61B 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 | ~10 kDa | |
| Observed band | ~15 kDa | |
| Gel | 5–20% (catalog A07334-2) | |
| Positive control | Cerebellum (IHC candidate; verify WB) +4 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Acetylated | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB C7 membership | |
| Isoform | 1 isoform(s) |
Section 1
Source-Linked SEC61B Western Blot Protocol Options
The A07334-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human A431, human PC-3, human 293T, human Hela, rat C6, mouse C2C12, mouse RAW264.7, mouse 3T3-L1 (catalog A07334-2) |
| Gel % | 5–20% (catalog A07334-2) |
| Load | 30 ug; reducing conditions (catalog A07334-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A07334-2) |
| Membrane | nitrocellulose membrane (catalog A07334-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A07334-2) |
| Primary antibody | A07334-2 · 0.25 μg/mL (catalog A07334-2) |
| Primary incubation | overnight at 4°C (catalog A07334-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A07334-2) |
| Secondary incubation | 1.5 hour at RT (catalog A07334-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A07334-2) |
| Detection | ECL (catalog A07334-2) |
Section 2
What Is the Expected SEC61B Western Blot Band Size?
SEC61B is predicted at 10 kDa and observed at ~15 kDa in reducing lysates; the cause of the difference is not established.
| Band at ~15 kDa | Empirical SEC61B band in reducing whole-cell lysates; the reason it runs above the predicted mass is not established. |
| Band near 10 kDa | Near the predicted SEC61B mass; confirm its identity with antibody controls. |
| Weak or absent band in whole-cell lysate | SEC61B is an endoplasmic reticulum membrane protein, so recovery may depend on membrane extraction. |
| Nearby doublet | SEC61B has documented phosphorylation sites, but a visible mobility difference is not established; verify band identity. |
💡Expected SEC61B appearanceSEC61B has a predicted mass of 10 kDa and an empirical band at ~15 kDa in reducing whole-cell lysates; the cause of the difference is unestablished, so confirm identity with antibody controls.
| Predicted mass | The 96-residue protein has a calculated mass of 10 kDa. |
| N-acetylproline at residue 2 | This modification is documented, but a visible size effect is not established. |
| Phosphoserine at residue 7 | Phosphorylation is documented, but a visible mobility shift is not established. |
| Phosphothreonine at residue 9 | Phosphorylation is documented, but a visible mobility shift is not established. |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | SEC61B is an endoplasmic reticulum membrane protein and may be poorly recovered. | Check membrane extraction and include a positive lysate control. |
| Band higher than expected | An empirical ~15 kDa band exceeds the predicted 10 kDa; the cause is unestablished. | Compare with the documented reducing-lysate band and verify antibody specificity. |
| Band lower than expected | A lower band has no established SEC61B processing explanation in the supplied features. | Check molecular-weight markers and verify band identity with an independent antibody. |
| Multiple bands | SEC61B has phosphorylation sites, but distinct bands from them are not established. | Confirm which band is SEC61B with an independent antibody or SEC61B depletion. |
| Weak or no signal | Incomplete extraction or transfer of this small membrane protein may limit detection. | Check membrane-protein recovery, transfer, and a positive lysate control. |
Experiment evidence
Western blot images showing SEC61B band
A07334-2 · SEC61B
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for SEC61B Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for SEC61B in Western blot, you can use cerebellum tissue, which shows high expression in HPA.
Positive control: Cerebellum (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside SEC61B.
⚠️Feasibility: As an ER membrane protein, SEC61B requires effective membrane protein extraction for a clear signal.
HPA tissue expression evidence for SEC61B
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 | Purkinje cells | High | Protein (IHC) | HPA → |
| Hippocampus | neuronal cells | High | Protein (IHC) | HPA → |
| Pancreas | exocrine glandular cells | High | Protein (IHC) | HPA → |
| Placenta | trophoblastic cells | High | Protein (IHC) | HPA → |
| Salivary gland | 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 | Not detected | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | follicle cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | smooth muscle cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced SEC61B Western Blot Tips
Deeper troubleshooting and optimisation questions for SEC61B, answered from its protein features.
How should SEC61B 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 a second SEC61B isoform explain another band?
Isoforms · The supplied UniProt record lists one isoform and no alternative sequence. Do not assign another band to a SEC61B isoform based on this record alone; verify its identity independently.
Which listed SEC61B modifications matter when interpreting bands?
PTM · UniProt lists N-acetylproline at position 2, phosphoserine at positions 7, 13, 14 and 17, and phosphothreonine at position 9. These are UniProt coordinates; antibody or paper numbering may differ. The listed sites do not establish that a modification causes a visible band shift.
Does this guide establish induction of SEC61B?
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 SEC61B Western blot?
Transfer · SEC61B is a small, 10 kDa predicted, single-pass membrane protein. Choose transfer conditions that retain small proteins, and check the membrane and post-transfer gel to assess retention and transfer. The supplied features do not specify a particular membrane or transfer setting.
How should blocking conditions be optimised for SEC61B?
Blocking · Compare blocking conditions using the SEC61B antibody and an ER membrane-containing sample. Select the condition that gives a clear band at the empirically observed ~15 kDa position with low background. The UniProt features do not identify a preferred blocker.
How should the SEC61B band be quantified?
Quantitation · Quantify the verified SEC61B band near its empirical ~15 kDa position across exposures within the assay's linear range. Keep sample preparation consistent for this ER membrane protein. The predicted 10 kDa mass alone should not determine which band is measured.
Why might SEC61B appear near 15 kDa instead of its predicted 10 kDa?
Interpretation · The supplied Western blots show an apparent band near 15 kDa, while the predicted mass is 10 kDa. SEC61B is a 96-residue single-pass ER membrane protein, but these features alone do not establish why it migrates differently. Verify band identity before assigning a cause to the difference.
Check bands against the empirical ~15 kDa position and verify their identity. The record lists one isoform, no glycosylation sites and six modified residues, but it does not establish that any of these features produces a particular extra band. SEC61B belongs to a translocon complex; complex membership alone does not identify an unexpected band.
Check bands against the empirical ~15 kDa position and verify their identity. The record lists one isoform, no glycosylation sites and six modified residues, but it does not establish that any of these features produces a particular extra band. SEC61B belongs to a translocon complex; complex membership alone does not identify an unexpected band.
Boster reagents
SEC61B 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-SEC61B Antibody Picoband®
Cat # A07334-2
The catalog reports one anti-SEC61B antibody for Western blot, A07334-2, with reported human, mouse, and rat reactivity. Its product image shows a band near 15 kDa in tested cell lysates, above the expected 10 kDa; no independent validation is supplied.
Which to pick: A07334-2 is the only listed option and has a Western blot image using human A431, PC-3, 293T, and HeLa; rat C6; and mouse C2C12, RAW264.7, and 3T3-L1 lysates. Match your sample to these tested contexts where possible.
Source: BosterBio SEC61B gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P60468.
- Human Protein Atlas. SEC61B tissue expression.
- PMC9629031 — target-verified WB comparison