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CDCA2 / Cell division cycle-associated protein 2 · IHC design guide
Design Immunohistochemistry for CDCA2
Plan chromogenic CDCA2 IHC around nuclear staining in most tissues and strong staining of pachytene spermatocytes in testis (HPA tissue IHC). Score staining by cell type and account for redistribution onto chromatin during anaphase (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CDCA2 Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Predominantly nuclear in tissue IHC (HPA tissue IHC) | |
| Staining pattern | Nuclear staining in most tissues; strongest in testis (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A07293-2) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A07293-2) | |
| Caveat | Mitotic redistribution can alter the expected nuclear pattern (UniProt) | |
| Regulation | Cell-cycle-dependent chromatin association (UniProt) | |
| Isoform / epitope | Two isoforms reported; check epitope coverage (UniProt) |
Section 1
Recommended CDCA2 IHC & IF Protocols
The catalog antibody protocol uses Tris-EDTA retrieval at pH 9.0 (datasheet A07293-2). Two published CDCA2 IHC protocols provide tissue preparation and staining details (PMC7039090; PMC7248370).
| Sample | Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A07293-2) |
| Fixation | Image fixative and duration unreported (datasheet A07293-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Tris-EDTA pH 9.0 (datasheet A07293-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-CDCA2, 1:100-1:300 (datasheet A07293-2) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CDCA2-positive staining in endocrine cells of appendix (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in a majority of tissues, most abundant in testis. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated Tris-EDTA retrieval at pH 9.0 for the catalog antibody (datasheet A07293-2); the published excerpts do not specify retrieval conditions (PMC7039090; PMC7248370).
Section 2
What Is the Expected CDCA2 Staining Pattern?
CDCA2 is a nuclear, nonmembrane protein that occupies the nucleoplasm during interphase and redistributes during mitosis (UniProt Q69YH5: localization and topology). In paraffin-section IHC, expect nuclear staining in a majority of tissues, with especially abundant staining in testis (HPA: tissue IHC profile). HPA rates the tissue profile Enhanced, while describing medium consistency between antibody staining and RNA expression (HPA: tissue IHC reliability).
| Distinct nuclear staining in testis pachytene spermatocytes, with little staining in nearby negative cells. | This matches a reported high-signal cell population and CDCA2’s usual nuclear location (HPA: High in testis pachytene spermatocytes; UniProt Q69YH5: nucleus). Score the named cell population and compartment together; a dark area elsewhere in the section does not establish the same pattern. |
| A few dividing cells show less sharply nuclear staining, while interphase cells retain nuclear staining. | Cell-cycle stage can change the appearance: CDCA2 is nucleoplasmic during G1, S and G2, becomes diffuse as the nuclear membrane breaks down, then accumulates on chromatin by anaphase (UniProt Q69YH5: cell-cycle localization). Interpret these cells alongside the section’s overall nuclear pattern. |
| Strong, widespread staining is confined to cell surfaces or cytoplasm, without a convincing nuclear pattern. | That distribution conflicts with the expected IHC profile and the absence of a transmembrane segment (HPA: nuclear expression in a majority of tissues; UniProt Q69YH5: localization and topology). Reassess specificity and detection before calling it CDCA2; isolated mitotic redistribution does not explain a section-wide surface pattern. |
| Adipocytes stain strongly, or staining follows tissue edges and other unexpected cell populations. | Adipocytes are reported as not detected, so strong staining there raises concern for cross-reactivity or endogenous detection activity (HPA: adipose adipocytes not detected). Compare staining with a control lacking primary antibody, and judge any unexpected population against its own HPA entry rather than treating an entire tissue as uniformly negative. |
| Color is diffuse across the section, or a known-positive population has no detectable signal. | Diffuse color without cell-restricted nuclear staining is difficult to score as CDCA2 (HPA: nuclear tissue IHC profile). Conversely, absence in testis pachytene spermatocytes conflicts with their reported high staining (HPA: High in testis pachytene spermatocytes). Check slide quality and controls before interpreting either result biologically. |
💡Expected CDCA2 appearanceCall the IHC result positive when staining is clearly nuclear and strongest in a documented high-staining population such as testis pachytene spermatocytes (HPA: High; UniProt Q69YH5: nucleus); diffuse, cell-surface or broadly unexpected staining should prompt a specificity check (HPA: tissue IHC profile; UniProt Q69YH5: topology).
| Cell-cycle position (UniProt Q69YH5: localization) | Interphase CDCA2 occupies nucleoplasm and is excluded from nucleoli; mitotic cells can appear diffuse before chromatin accumulation at anaphase (UniProt Q69YH5: cell-cycle localization). A rare dividing cell should therefore be interpreted in context, rather than used to redefine the expected whole-section staining pattern. |
| Cell population and tissue (HPA: tissue IHC profile) | HPA reports high staining in testis pachytene spermatocytes and intestinal endocrine cells, medium staining in bone-marrow hematopoietic cells and breast glandular cells, and no detection in listed populations including adipocytes (HPA: tissue IHC). Select and score the documented cell population; neighboring cells need not share its staining level. |
| Antibody evidence (HPA: antibody validation) | HPA lists Enhanced IHC validation for HPA030049, meaning its pattern was reproduced by independent antibodies or orthogonal data; HPA026293 has no listed IHC status (HPA: antibodies). This supports using the validated pattern as a reference, while a different antibody’s result still needs its own controls. |
| IF/ICC: where is CDCA2 seen? (HPA: subcellular ICC-IF) | The supported main location is nucleoplasm; HPA also lists cytosol as an uncertain additional location (HPA: subcellular ICC-IF). Treat an IF/ICC cytosolic signal cautiously and consult the separate IF/ICC guide for that application. The ICC-IF observation does not by itself change the expected nuclear IHC interpretation. |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in testis pachytene spermatocytes (HPA: High in testis pachytene spermatocytes). | The run may have failed, the relevant cells may be absent from the examined field, or the antibody may not work under the chosen IHC conditions; the supplied sources do not identify a CDCA2-specific fixation effect. | First locate the reported cell population, then inspect the run controls. If controls indicate a technical failure, revisit the catalog antibody’s validated IHC-P workflow and repeat the run before assigning a biological negative. |
| Staining is only cytoplasmic or at cell borders across many cells (HPA: nuclear tissue IHC profile). | A broad nonnuclear pattern is inconsistent with CDCA2’s usual localization and lack of a transmembrane segment (UniProt Q69YH5: localization and topology); nonspecific staining or detection background is plausible. | Compare with a control lacking primary antibody and inspect whether any clear nuclear signal remains. Review antibody specificity and detection conditions before scoring the nonnuclear color as CDCA2. |
| Adipocytes or another HPA-negative population stain strongly (HPA: adipocytes not detected). | Cross-reactivity or endogenous detection activity can mimic a positive chromogenic result. An HPA-negative designation applies to the named cell population, not automatically to every cell in that tissue (HPA: tissue IHC entries). | Confirm the stained cell identity, compare a control lacking primary antibody, and assess the detection system’s background. Report unexpected staining separately from staining in documented positive populations. |
| Diffuse brown haze obscures nuclei across the section. | High general background can prevent assessment of the cell-restricted nuclear pattern reported by HPA (HPA: tissue IHC profile). This appearance alone cannot establish a CDCA2-specific cause. | Review a control lacking primary antibody, washing, blocking and detection steps as general IHC quality checks. Optimize the run until nuclei and cell boundaries can be assessed, then rescore the same tissue. |
| Only scattered dividing cells look diffuse, while other positive cells look nuclear. | CDCA2 redistributes when the nuclear membrane breaks down and later accumulates on anaphase chromatin (UniProt Q69YH5: cell-cycle localization). These stage-dependent appearances can differ from interphase nucleoplasmic staining. | Assess nuclear staining in interphase cells and note the morphology of the scattered dividing cells. Avoid classifying a whole slide as having cytoplasmic CDCA2 on the basis of those cells alone. |
| A newly selected antibody gives a different tissue pattern from the reference. | HPA reports Enhanced IHC validation for HPA030049 but no listed IHC status for HPA026293; that evidence does not validate every antibody equally (HPA: antibodies). | Check the new antibody’s documented IHC suitability, run controls and the named HPA-positive cell populations. If the disagreement persists, describe it as an antibody-dependent result rather than changing the expected pattern. |
Experiment evidence
IHC & IF images showing CDCA2 staining
A07293-2 · IHCImmunohistochemical analysis of paraffin-embedded human tonsil. 1, Antibody was diluted at 1:200 (4° overnight). 2, Tris-EDTA, pH9.0 was use
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000184661-CDCA2/tissue/cerebral+cortex#imid_20759392
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000184661-CDCA2/tissue/colon#imid_20759302
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000184661-CDCA2/tissue/liver#imid_20759319
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000184661-CDCA2/tissue/kidney#imid_20759312
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA026293; green = CDCA2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA026293; green = CDCA2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CDCA2 IHC & IF
🧪Run colon first; its endocrine cells should stain for CDCA2 (HPA: High in colon endocrine cells). Use heart muscle as the negative tissue because cardiomyocytes are not detected (HPA: Not detected in heart muscle cardiomyocytes); on the colon slide, neighboring non-endocrine cells should show background-level staining to serve as an internal reference, but their CDCA2 status requires verification (UniProt Q69YH5: ubiquitously expressed).
Positive control tissue: Appendix (Endocrine cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CDCA2 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and immunoglobulin class, with clonality considered when selecting a comparable control (standard IHC practice). A CDCA2 knockout sample, if available, provides a biological negative; block endogenous peroxidase for chromogenic detection in the colon section (standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported in the supplied evidence, and the selected A07293-2 paraffin-section caption does not state a fixative (selected tissue-IHC caption: fixative unreported). That caption uses Tris-EDTA at pH 9.0 for retrieval, but does not establish that retrieval is required (selected tissue-IHC caption). No supplied comparison shows whether frozen sections or IF are easier; for colon IHC, distinguish staining in endocrine cells from background in adjacent cells (HPA: High in colon endocrine cells; standard IHC practice).
HPA tissue IHC evidence for CDCA2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Appendix | Endocrine cells | High | Protein (IHC) | HPA → |
| Colon | Endocrine cells | High | Protein (IHC) | HPA → |
| Duodenum | Endocrine cells | High | Protein (IHC) | HPA → |
| Rectum | Endocrine cells | High | Protein (IHC) | HPA → |
| Small intestine | Endocrine cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CDCA2 IHC Tips
Troubleshoot CDCA2 staining by checking nuclear localisation, cell identity and section controls before comparing signal across specimens (UniProt Q69YH5; HPA tissue IHC).
How should I adjust retrieval when CDCA2 nuclear staining is weak?
Start with heat-mediated Tris-EDTA retrieval at pH 9.0 for paraffin sections (datasheet A07293-2). The documented human tonsil image used that buffer, a 1:200 primary antibody dilution overnight at 4°C, and a 1:200 secondary antibody for 45 minutes at room temperature (datasheet A07293-2). If nuclei remain weak, compare retrieval duration on adjacent sections while holding antibody dilution and chromogen development constant (standard IHC practice). Check that any gain appears in plausible nucleoplasm rather than section edges or diffuse background; CDCA2 is mainly nucleoplasmic during interphase (UniProt Q69YH5; HPA subcellular).
Could fixation explain weak or uneven CDCA2 staining?
Target-specific sensitivity to fixation is unknown from the supplied evidence; the documented CDCA2 image identifies a paraffin-embedded human tonsil section but does not state its fixative (datasheet A07293-2). Record the actual fixative and fixation duration for each specimen, then compare similarly processed sections before changing the antibody dilution (standard IHC practice). Use matched positive tissue and a no-primary control to separate loss of nuclear signal from nonspecific chromogen deposition (HPA tissue IHC; standard IHC practice). Do not infer a CDCA2 fixation effect from its nuclear localisation, phosphorylation sites or tissue staining pattern (UniProt Q69YH5; HPA tissue IHC).
Where should convincing CDCA2 staining appear in tissue sections?
In interphase cells, expect predominantly nucleoplasmic staining with nucleolar exclusion (UniProt Q69YH5); HPA reports nuclear staining across a majority of tissues (HPA tissue IHC). A mitotic cell can look different: CDCA2 becomes diffuse after nuclear envelope breakdown and accumulates on chromatin by anaphase (UniProt Q69YH5). Review cell morphology and a nuclear counterstain before calling an isolated cytoplasmic profile positive; HPA supports nucleoplasm as the main location but labels additional cytosol localisation uncertain (HPA subcellular). Score nuclear and cytoplasmic patterns separately, and verify unusual patterns against an adjacent section and controls (standard IHC practice).
Could isoforms or epitope accessibility alter the staining pattern?
CDCA2 has 2 annotated isoforms and a PP1-binding region at residues 389–424 (UniProt Q69YH5). Its 19 listed modified residues include phosphoserines, so an epitope-specific explanation requires the antibody's mapped immunogen or epitope; those details are absent here (UniProt Q69YH5; supplied product caption). Keep retrieval, primary dilution and detection constant when comparing sections, and record whether discrepant staining is nuclear or diffuse (standard IHC practice; UniProt Q69YH5). Do not assign an isoform or phosphorylation state from chromogenic intensity alone; use an independently validated reagent or orthogonal assay if that distinction matters (standard IHC practice).
How can IF help assess an ambiguous CDCA2 IHC pattern?
Use IF/ICC as a separate localisation check, with a nuclear stain and a validated cell-type marker for the population being examined; HPA reports high CDCA2 staining in pachytene spermatocytes (HPA tissue IHC). Choose a fluorophore channel with low measured tissue autofluorescence, and include single-channel and no-primary controls when multiplexing (standard IF practice). CDCA2 has no transmembrane segment and is mainly nucleoplasmic, so intracellular access calls for permeabilisation appropriate to the antibody's mapped epitope (UniProt Q69YH5; standard IF practice). Compare the resulting nuclear pattern with the chromogenic section, while treating HPA's additional cytosol localisation as uncertain (HPA subcellular).
What should I check when CDCA2 staining is diffuse or widespread?
Check a no-primary section for detection-system signal, and confirm that the peroxidase block and chromogen development were controlled consistently (standard chromogenic IHC practice). Compare the suspect section with the documented 1:200 primary dilution and Tris-EDTA pH 9.0 retrieval, changing only one condition at a time (datasheet A07293-2; standard IHC practice). Strong diffuse cytoplasmic staining warrants scrutiny because CDCA2 is mainly nucleoplasmic during interphase, although it disperses during mitosis (UniProt Q69YH5). Inspect section edges, folds and damaged areas separately from intact cells before attributing their staining to CDCA2 (standard IHC practice).
How should I quantify CDCA2 across sections with different cellularity? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue area before scoring, then report the percentage of CDCA2-positive nuclei and nuclear intensity using a consistent threshold (standard IHC practice; HPA tissue IHC). An H-score can combine intensity and the fraction of cells at each intensity; alternatively, report positive nuclei per mm² of evaluable tissue (standard IHC practice). Normalise counts to all eligible nuclei in the same compartment, and exclude folds, necrosis and edge artefacts by a prespecified rule (standard IHC practice). Record mitotic cells separately where feasible because CDCA2 redistributes during mitosis (UniProt Q69YH5).
How can I distinguish true CDCA2 positivity from staining artefacts?
Give greatest weight to staining within intact nuclei of the expected cells: HPA reports high staining in testicular pachytene spermatocytes and medium staining in bone marrow hematopoietic cells (HPA tissue IHC). Unexpected cytoplasmic signal needs morphological review because interphase CDCA2 is nucleoplasmic, while mitotic CDCA2 can become diffuse (UniProt Q69YH5). Compare section edges and necrotic areas with intact tissue, and check a no-primary control for endogenous enzyme or detection background (standard IHC practice). Interpret intensity cautiously: HPA rates its tissue evidence Enhanced but reports only medium consistency between antibody staining and RNA expression (HPA tissue IHC).
Boster reagents
Best CDCA2 / Cell division cycle-associated protein 2 IHC Antibodies
A07293-2 covers human IHC and IF (catalog: applications and reactivity), with an IHC image of paraffin-embedded human tonsil (catalog: IHC caption).
Real IHC data
Anti-Repo-Man CDCA2 Antibody
Cat # A07293-2
A07293-2 is listed for human IHC and IF (catalog: applications and reactivity). Its IHC image shows paraffin-embedded human tonsil; no IF image is supplied (catalog: IHC caption; IF image list empty).
Which to pick: Choose A07293-2 for paraffin-section IHC: its own caption shows human tonsil stained at 1:200 after Tris-EDTA retrieval at pH 9.0 (catalog: IHC caption); the fixative is unreported. For IF/ICC, A07293-2 lists IF at 1:50, but supplies no IF image (catalog: dilution and IF image list). No cross-species option is documented, and clonality is unreported (catalog: Human reactivity; clone unspecified).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q69YH5 (CDCA2_HUMAN, Cell division cycle-associated protein 2).
- Human Protein Atlas. CDCA2 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CDCA2 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol..
- Human Protein Atlas. CDCA2 antibody validation summary (2 antibodies).
- CDCA2 acts as an oncogene and induces proliferation of clear cell renal cell carcinoma cells. Oncology letters 2020 — PMC7039090.
- CDCA2 promotes the proliferation of colorectal cancer cells by activating the AKT/CCND1 pathway in vitro and in vivo. BMC cancer 2019 — PMC6567669.
- CDCA2 Inhibits Apoptosis and Promotes Cell Proliferation in Prostate Cancer and Is Directly Regulated by HIF-1α Pathway. Frontiers in oncology 2020 — PMC7248370.
- Identification of CDCA2 as a Diagnostic and Prognostic Marker for Hepatocellular Carcinoma. Frontiers in oncology 2021 — PMC8517522.
- PubMed PMID:17974005 — UniProt-cited evidence.
- PubMed PMID:16421571 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CDCA2 (Cell division cycle-associated protein 2). Expected localisation Predominantly nuclear in tissue IHC (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, heat-mediated (datasheet A07293-2). Validated IHC antibody: A07293-2 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Endocrine cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human tonsil tissue; fixative not specified (datasheet A07293-2); 2. Fixation: Image fixative and duration unreported (datasheet A07293-2); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: Tris-EDTA pH 9.0 (datasheet A07293-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-CDCA2, 1:100-1:300 (datasheet A07293-2); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A07293-2); Caveat: Mitotic redistribution can alter the expected nuclear pattern (UniProt); Isoform / epitope: Two isoforms reported; check epitope coverage (UniProt); Regulation: Cell-cycle-dependent chromatin association (UniProt). Evidence: UniProt Q69YH5, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A07293-2, PMC7039090, PMC7248370. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CDCA2/immunohistochemistry-ihc-guide.