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CCND2 / G1/S-specific cyclin-D2 · Western blot design guide
Design a Western Blot for CCND2
Real validated CCND2 Western blot protocols, expected-band and isoform facts, troubleshooting for weak or shifted signal, and recommended anti-CCND2 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
CCND2 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 | ~33.1 kDa | |
| Gel | 12–15% (standard starting point) | |
| Positive control | Epididymis (IHC candidate; verify WB) +3 more | |
| Negative control | Adipose tissue (IHC candidate; verify WB) |
Important caveats
Reasons your observed band may differ from the expected size.
| PTM | Phosphorylated + Ubl conjugation | |
| Caveat | Phosphorylation-state controls | |
| Gene-set association | MSigDB Hallmark membership | |
| Isoform | 2 isoform(s) |
Section 1
Real Curated CCND2 Western Blot Protocols
The A01079 protocol combines labelled catalog values with standard starting conditions. Published comparisons retain their own sample, reagent and detection scope.
| Sample / lysate | RD, (catalog A01079) |
| 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 | Blocking buffer: 3% nonfat dry milk in TBST (catalog A01079) |
| Primary antibody | A01079 · 1:500 (catalog A01079) |
| Primary incubation | Overnight at 4 °C (standard starting point) |
| Secondary antibody | Goat Anti-Rabbit IgG, 1:10000 (catalog A01079) |
| Secondary incubation | 1 h at room temperature (standard starting point) |
| Wash | 3 × 5 min in TBST (standard starting point) |
| Detection | ECL (catalog A01079) |
Section 2
What Is the Expected CCND2 Western Blot Band Size?
CCND2 is predicted at 33.1 kDa; splice isoforms and phosphorylation are documented, but their effects on Western-blot migration are not demonstrated.
| Band near 33.1 kDa | Consistent with the predicted mass of CCND2; confirm identity with an appropriate control |
| Additional bands near the main band | Could include splice isoforms 1 and 2, but their migration is unknown |
| Slightly slower migrating band | Could reflect phosphorylation; the listed sites do not establish a visible shift |
| Unequal signal in nuclear and cytoplasmic fractions | Consistent with CCND2 localization in both compartments |
💡Expected CCND2 appearanceCCND2 has a predicted mass of 33.1 kDa, but no empirical band size is supplied; confirm any band near that size with an appropriate identity control.
| Predicted CCND2 mass | Places the full-length protein near 33.1 kDa before any unproven migration effects |
| Phosphoserine 271 | Could affect migration, but no visible size shift is established |
| Phosphothreonine 280 | Could affect migration, but no visible size shift is established |
| Splice isoforms 1 and 2 | Could differ in apparent size; their masses and separation are not supplied |
| Situation | Likely cause | Next action |
|---|---|---|
| No band in lysate | CCND2 may be missed if extraction poorly recovers its nuclear or cytoplasmic pool | Check fraction recovery and test an expression-positive lysate |
| Band higher than expected | Phosphorylation is documented, but its effect on migration is unproven | Compare phosphatase-treated and untreated samples and confirm band identity |
| Band lower than expected | An alternate splice product is possible, but no isoform mass is supplied | Check isoform expression and confirm the band with CCND2 depletion |
| Multiple bands | Isoforms 1 and 2 or phosphorylation could contribute; neither is shown to resolve as separate bands | Use CCND2 depletion and compare phosphatase-treated samples |
| Weak or no signal | The tested sample may have little recoverable CCND2 | Check loading and extraction, then test an expression-positive control |
Experiment evidence
Western blot images showing CCND2 band
A01079 · CCND2
← swipe / scroll for more images →
Real WB data behind the band-size expectations in this section.
Sample controls for CCND2 Western blot
🧪HPA-IHC candidate guidance (verify in WB): For positive controls for CCND2 in Western blot, you can use epididymis tissue, which HPA lists as a positive candidate with medium expression.
Positive control: Epididymis (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: Positive tissue expression is medium, so the CCND2 signal may be modest.
HPA tissue expression evidence for CCND2
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 |
|---|---|---|---|---|
| Epididymis | glandular cells | Medium | Protein (IHC) | HPA → |
| Heart muscle | cardiomyocytes | Medium | Protein (IHC) | HPA → |
| Rectum | glandular cells | Medium | Protein (IHC) | HPA → |
| Thyroid gland | glandular cells | Medium | 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 → |
| Bone marrow | hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
| Caudate | glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CCND2 Western Blot Tips
Deeper troubleshooting and optimisation questions for CCND2, answered from its protein features.
How should CCND2 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.
How might the two CCND2 isoforms affect band interpretation?
Isoforms · Isoform 2 replaces canonical residues 138–209 with a different sequence and lacks residues 210–289, making it shorter than isoform 1. The features do not establish where either isoform runs on a blot. Check which isoform the antibody recognizes before assigning bands.
Check the antibody epitope against the isoform sequences. An epitope within canonical residues 210–289 is absent from isoform 2; residues 138–209 are replaced in isoform 2. An antibody targeting either region may detect the isoforms differently.
Check the antibody epitope against the isoform sequences. An epitope within canonical residues 210–289 is absent from isoform 2; residues 138–209 are replaced in isoform 2. An antibody targeting either region may detect the isoforms differently.
Could phosphorylation account for a shifted CCND2 band?
PTM · UniProt lists phosphoserine 271 and phosphothreonine 280 in the canonical sequence. Isoform 2 lacks that region. These coordinates use UniProt canonical numbering, which may differ from antibody or paper numbering. Phosphorylation is a possibility to investigate, but these annotations do not demonstrate a visible shift.
No glycosylation sites, signal peptide or propeptide are listed. The supplied features therefore provide no basis to assign an unexpected band to glycosylation or signal-peptide or propeptide cleavage. Assess antibody specificity and isoform recognition before assigning its identity.
No glycosylation sites, signal peptide or propeptide are listed. The supplied features therefore provide no basis to assign an unexpected band to glycosylation or signal-peptide or propeptide cleavage. Assess antibody specificity and isoform recognition before assigning its identity.
Does this guide establish induction of CCND2?
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 CCND2?
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 A01079 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 CCND2 be quantified across cell fractions?
Quantitation · CCND2 is reported in the nucleus, cytoplasm and nuclear membrane. Compare equivalent fractions and normalize within each fraction; a change in one fraction may reflect redistribution. Cyclin D–CDK4 complexes accumulate at the nuclear membrane and translocate into the nucleus through interaction with KIP/CIP family members.
Should CCND2 run exactly at its predicted 33.1 kDa?
Interpretation · 33.1 kDa is the predicted mass of the 289-residue canonical protein. Apparent migration can differ, and no observed band size is supplied. The listed features alone do not establish a visible shift or explain a mass difference.
Boster reagents
CCND2 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-Cyclin D2 CCND2 Antibody
Cat # A01079
The catalog reports A01079, an anti-CCND2 antibody with stated human, mouse, and rat reactivity. Its Western blot image shows RD lysate at 25 µg per lane with a 1:500 primary dilution; the supplied evidence shows no other tested specimens.
Which to pick: A01079 is the only listed option and has a Western blot image using RD lysate. Consider the stated reactivity for your samples, while recognizing that the supplied image documents only the RD test context.
Source: BosterBio CCND2 gene-info card — filtered to Western-blot-capable antibodies; each card shows that product's actual WB validation figure.
References
- UniProt Consortium. UniProt entry P30279.
- Human Protein Atlas. CCND2 tissue expression.
- PMC7318366 — target-verified WB comparison
- PMC13260899 — target-verified WB comparison
- PMC13365504 — target-verified WB comparison