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- Table of Contents
POLD3 / DNA polymerase delta subunit 3 · Western blot design guide
Design a Western Blot for POLD3
Source-linked POLD3 Western blot protocol options, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-POLD3 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
POLD3 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 | ~51.4 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Appendix (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 | Modification-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 3 isoform(s) |
Section 1
Source-Linked POLD3 Western Blot Protocol Options
The A30646 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | hela cells (catalog A30646) |
| 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 | A30646 · 1:500 (catalog A30646) |
| 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 POLD3 Western Blot Band Size?
POLD3 has a predicted mass of 51.4 kDa; listed isoforms could affect band patterns, but their migration has not been demonstrated here.
| Band near 51.4 kDa | consistent with the predicted POLD3 mass; confirm its identity with controls |
| Several bands at different positions | could include isoforms 1, 2, and 3; their migration is not established |
| Band in a nuclear fraction | consistent with the reported nuclear location of POLD3 |
| Band in a cytoplasmic fraction | consistent with the reported cytoplasmic location of POLD3 |
💡Expected POLD3 appearanceUniProt predicts 51.4 kDa for POLD3; no empirical band size is supplied, and isoform migration is unknown, so confirm a candidate band with antibody specificity and fractionation controls.
| UniProt predicted mass | 51.4 kDa provides the reference for a candidate band |
| Isoform 1 | its individual apparent size is not supplied |
| Isoform 2 | may differ in size from other isoforms, but its migration is not supplied |
| Isoform 3 | may differ in size from other isoforms, but its migration is not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | POLD3 may be missed if its nuclear fraction is poorly recovered | check nuclear extraction and a nuclear fraction control |
| Band higher than expected | identity or migration of the band is unconfirmed | compare with a POLD3 depletion control and molecular weight markers |
| Band lower than expected | an isoform is possible, but isoform sizes are unknown | verify identity with POLD3 depletion or an antibody to another epitope |
| Multiple bands | three isoforms are listed, but distinct bands are not established | test which bands respond to POLD3 depletion |
| Weak or no signal | POLD3 recovery or antibody recognition may be inadequate | check extraction, loading, and a positive control lysate |
Experiment evidence
Western blot images showing POLD3 band
A30646 · POLD3
A30646 · POLD3
A05111-2 · POLD3
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for POLD3 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for POLD3 in Western blot, you can use appendix tissue, which HPA scores as High.
Positive control: Appendix (IHC candidate; verify WB)
Negative control: Adipose tissue (IHC candidate; verify WB)
Loading controls: Run GAPDH, β-actin, and a total-protein stain alongside the samples.
⚠️Feasibility: An intracellular positive is feasible, but validate HPA Not-detected adipose tissue as a Western-blot negative.
HPA tissue expression evidence for POLD3
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 → |
| Bronchus | respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Colon | glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | 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 → |
| Adrenal gland | glandular cells | Low | Protein (IHC) | HPA → |
| Caudate | glial cells | Low | Protein (IHC) | HPA → |
| Cerebral cortex | neuronal cells | Low | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Low | Protein (IHC) | HPA → |
Section 3
Advanced POLD3 Western Blot Tips
Deeper troubleshooting and optimisation questions for POLD3, answered from its protein features.
How should POLD3 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 POLD3 isoforms produce smaller bands?
Isoforms · Yes. Isoform 2 lacks residues 1–39, and isoform 3 lacks residues 1–106 relative to the 466-residue canonical sequence. They may produce smaller bands, but their apparent positions are not supplied. Check whether the antibody recognizes a region retained in each isoform.
Which POLD3 modifications matter when interpreting bands?
PTM · UniProt lists N-acetylalanine at position 2; phosphothreonine at 277 and 411; and phosphoserine at 307, 407, 409, 413, and 458. These are UniProt coordinates, which may differ from paper or antibody numbering. Their presence alone does not establish a visible band shift.
Phosphorylation is listed at UniProt positions 277, 307, 407, 409, 411, 413, and 458. These sites make phosphorylation a possibility to investigate, but the feature list supplies no observed shift and cannot establish that phosphorylation caused one.
Phosphorylation is listed at UniProt positions 277, 307, 407, 409, 411, 413, and 458. These sites make phosphorylation a possibility to investigate, but the feature list supplies no observed shift and cannot establish that phosphorylation caused one.
Does this guide establish induction of POLD3?
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 POLD3?
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 A30646 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 POLD3 bands be quantified across samples?
Quantitation · Quantify the same resolved band or defined set of bands across samples. POLD3 has three isoforms, and its location feature places it in the nucleus and cytoplasm, with partial colocalization at S phase replication sites. Keep the sampled fraction consistent when comparing signals.
Should POLD3 run at its predicted 51.4 kDa?
Interpretation · 51.4 kDa is the predicted mass. No observed band position is supplied, so an apparent mass difference cannot be established or assigned to a modification from these features alone.
The supplied location feature reports recruitment to DNA damage sites within 2 hours after UV irradiation. It does not establish increased total POLD3 abundance. If comparing UV-treated samples, distinguish changes in localization from changes in total signal.
Isoform 2 lacks the first 39 residues and isoform 3 the first 106 residues of the canonical sequence, so either is a candidate. The supplied features do not identify an observed band or establish its identity. Check whether the antibody's recognition region is present in the proposed isoform.
The supplied location feature reports recruitment to DNA damage sites within 2 hours after UV irradiation. It does not establish increased total POLD3 abundance. If comparing UV-treated samples, distinguish changes in localization from changes in total signal.
Isoform 2 lacks the first 39 residues and isoform 3 the first 106 residues of the canonical sequence, so either is a candidate. The supplied features do not identify an observed band or establish its identity. Check whether the antibody's recognition region is present in the proposed isoform.
Boster reagents
POLD3 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-POLD3 Antibody
Cat # A30646
Real WB data
Anti-POLD3 Antibody Picoband®
Cat # A05111-2
Both the supplier anti-POLD3 antibodies list human, mouse, and rat reactivity and have WB images. A30646 shows HeLa cells; A05111-2 shows human MCF-7 and HeLa cells plus rat and mouse testis lysates. A05111-2 reports a ~70 kDa band versus an expected 51 kDa.
Which to pick: Choose A05111-2 for its documented WB conditions and examples across the listed species, while assessing the band-size discrepancy. A30646 provides a HeLa WB example at 1:500, with fewer conditions reported in its caption.
Source: BosterBio POLD3 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry Q15054.
- Human Protein Atlas. POLD3 tissue expression.
- PMC6283425 — target-verified WB comparison
- PMC11252662 — target-verified WB comparison
- PMC5384111 — target-verified WB comparison