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- Table of Contents
ERP29 / Endoplasmic reticulum resident protein 29 · Western blot design guide
Design a Western Blot for ERP29
Source-linked ERP29 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-ERP29 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
ERP29 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 | ~29 kDa | |
| Observed band | ~29 kDa | |
| Gel | 5–20% (catalog A03621-2) | |
| Positive control | Appendix (IHC candidate; verify WB) +4 more | |
| Negative control | Ovary (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Cleaved | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 2 isoform(s) |
Section 1
Source-Linked ERP29 Western Blot Protocol Options
The A03621-2 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | human 293T, human HepG2, human Hela, human Jurkat, human K562, human placenta, monkey COS-7 (catalog A03621-2) |
| Gel % | 5–20% (catalog A03621-2) |
| Load | 30 ug; reducing conditions (catalog A03621-2) |
| Transfer | a nitrocellulose membrane at 150 mA for 50-90 minutes (catalog A03621-2) |
| Membrane | nitrocellulose membrane (catalog A03621-2) |
| Blocking | 5% non-fat milk/TBS for 1.5 hour at RT (catalog A03621-2) |
| Primary antibody | A03621-2 · 0.5 μg/mL (catalog A03621-2) |
| Primary incubation | overnight at 4°C (catalog A03621-2) |
| Secondary antibody | goat anti-rabbit IgG-HRP, 1:5000 (catalog A03621-2) |
| Secondary incubation | 1.5 hour at RT (catalog A03621-2) |
| Wash | TBS-0.1%Tween 3 times with 5 minutes each (catalog A03621-2) |
| Detection | ECL (catalog A03621-2) |
Section 2
What Is the Expected ERP29 Western Blot Band Size?
ERP29 is predicted at 29 kDa and observed near 29 kDa; the evidence does not establish a cause for any migration difference.
| Band near 29 kDa | Empirical ERP29 band in reducing whole-cell lysates, matching the predicted precursor mass |
| Band below 29 kDa | Could reflect removal of the 1–32 signal peptide; band identity requires confirmation |
| Band near twice the monomer size under non-reducing conditions | Could reflect retained ERP29 homodimers; no inter-chain disulfide is listed |
| Additional bands at different positions | Could reflect isoforms 1 and 2 if their migration differs; their masses are not supplied |
💡Expected ERP29 appearanceERP29 has a predicted precursor mass of 29 kDa and an empirical band near 29 kDa in reducing whole-cell lysates; confirm band identity with appropriate controls because its features do not establish distinct migration shifts.
| Predicted ERP29 precursor mass | 29 kDa predicted, matching the empirical band near 29 kDa |
| Signal peptide at residues 1–32 | Cleavage can make mature ERP29 smaller than its full-length precursor |
| ERP29 homodimer | Could appear near twice the monomer size if the dimer survives sample preparation |
| Isoforms 1 and 2 | May differ in size, but their masses and separation on a blot are not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | ERP29 in the ER lumen may be poorly recovered during extraction | Check extraction of an ER-luminal control and compare with the reported whole-cell lysate band near 29 kDa |
| Band higher than expected | ERP29 homodimers may persist during sample preparation | Compare reducing and non-reducing preparations and verify band identity |
| Band lower than expected | Signal peptide cleavage can reduce the precursor size | Check whether the antibody detects mature ERP29 and verify the lower band with an identity control |
| Multiple bands | Isoforms 1 and 2 could migrate differently | Verify each band's identity; do not assign an isoform by size alone |
| Weak or no signal | Recovery of ER-luminal ERP29 may be insufficient | Check ER protein recovery, loading, and antibody performance against a positive lysate |
Experiment evidence
Western blot images showing ERP29 band
A03621-2 · ERP29
M03621-1 · ERP29
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for ERP29 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for ERP29 in Western blot, you can use appendix lysate, which has high HPA expression.
Positive control: Appendix (IHC candidate; verify WB)
Negative control: Ovary (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside.
⚠️Feasibility: Ovary is a feasible negative candidate because HPA reports ERP29 as not detected there.
HPA tissue expression evidence for ERP29
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 → |
| Bone marrow | hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | glial cells | High | Protein (IHC) | HPA → |
Lower expression tissues · IHC evidence, not confirmed WB-negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Ovary | ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | myocytes | Low | Protein (IHC) | HPA → |
| Adipose tissue | adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | glandular cells | Medium | Protein (IHC) | HPA → |
| Colon | glandular cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced ERP29 Western Blot Tips
Deeper troubleshooting and optimisation questions for ERP29, answered from its protein features.
How should ERP29 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 ERP29 isoform 2 produce a different band?
Isoforms · Yes. UniProt isoform 2 replaces residues 49..53 (VIPKS to IMVTS) and lacks residues 54..261, so it is much shorter than isoform 1. An antibody targeting the missing region would not detect isoform 2.
Can ERP29 phosphorylation explain a band shift?
PTM · UniProt lists phosphotyrosine at residues 64 and 66, both by PKDCC. These are UniProt coordinates; antibody or paper numbering may differ. Their presence does not establish that phosphorylation causes a visible shift. Both sites are absent from isoform 2.
Track phosphotyrosine signal separately from total ERP29, using the same samples for comparison. UniProt assigns PKDCC-dependent phosphorylation to Tyr64 and Tyr66; isoform 2 lacks both sites, so a phosphosite readout cannot represent both isoforms.
Track phosphotyrosine signal separately from total ERP29, using the same samples for comparison. UniProt assigns PKDCC-dependent phosphorylation to Tyr64 and Tyr66; isoform 2 lacks both sites, so a phosphosite readout cannot represent both isoforms.
Does this guide establish induction of ERP29?
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 ERP29?
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 A03621-2 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 ERP29 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.
Why is the observed ERP29 band near 29 kDa?
Interpretation · Canonical ERP29 is predicted at 29 kDa, matching the reported apparent band. Its signal peptide spans UniProt residues 1..32, but that feature alone does not establish the mature protein's apparent mobility or explain a mass difference.
ERP29 is listed in the endoplasmic reticulum lumen and was identified in melanosome fractions from stages I to IV. Compare like fractions and account for their different protein composition when interpreting band intensity.
Consider the much shorter isoform 2 and ERP29's reported homodimer when evaluating additional bands. UniProt also lists a large chaperone complex containing small amounts of ERP29. These features suggest possibilities but do not identify an unexpected band; its identity requires experimental verification.
ERP29 is listed in the endoplasmic reticulum lumen and was identified in melanosome fractions from stages I to IV. Compare like fractions and account for their different protein composition when interpreting band intensity.
Consider the much shorter isoform 2 and ERP29's reported homodimer when evaluating additional bands. UniProt also lists a large chaperone complex containing small amounts of ERP29. These features suggest possibilities but do not identify an unexpected band; its identity requires experimental verification.
Boster reagents
ERP29 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-ERP29 Antibody Picoband®
Cat # A03621-2
Real WB data
Anti-ERp29 Rabbit Monoclonal Antibody
Cat # M03621-1
Both listed anti-ERP29 antibodies have Western blot images. A03621-2 shows an approximately 29 kDa band in human and monkey lysates; M03621-1 shows HepG2 lysate. The supplied images do not show mouse or rat samples.
Which to pick: For human samples, either antibody has a WB image. Choose A03621-2 for the documented monkey COS-7 example or its broader set of human specimens. Both list mouse and rat reactivity, but neither supplied caption shows those samples.
Source: BosterBio ERP29 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P30040.
- Human Protein Atlas. ERP29 tissue expression.
- PMC8526686 — target-verified WB comparison
- PMC5585903 — target-verified WB comparison
- PMC4588584 — target-verified WB comparison