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CDC45 / Cell division control protein 45 homolog · IHC design guide
Design Immunohistochemistry for CDC45
Plan chromogenic CDC45 IHC around the nuclear pattern observed in most tissues (HPA tissue IHC). This guide highlights high staining in testis pachytene spermatocytes and the need to score staining by cell population (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CDC45 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 | Nuclear in most tissues (HPA tissue IHC) | |
| Staining pattern | Nuclear staining, highest in testicular seminiferous ducts (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet M01367-2) | |
| Positive control | Testis+3 more · see all | |
| Negative control | Adrenal gland+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining differs among cell populations within a tissue (HPA tissue IHC) | |
| Regulation | Cell-cycle associated (UniProt) | |
| Isoform / epitope | 3 isoforms; confirm epitope coverage (UniProt) |
Section 1
Recommended CDC45 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet: M01367-2) with three published CDC45 IHC protocols (PMC4714687; PMC7919231; PMC10956518).
| Sample | Paraffin-embedded human ovarian cancer tissue; fixative not specified (datasheet M01367-2) |
| Fixation | Image fixative and duration unreported (datasheet M01367-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet M01367-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet M01367-2) |
| Primary antibody | Mouse monoclonal (clone 6H6) anti-CDC45, 2μg/ml (datasheet M01367-2) |
| Primary incubation | Overnight at 4 °C (datasheet M01367-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet M01367-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CDC45-positive staining in pachytene spermatocytes of testis (HPA tissue IHC: High). HPA tissue profile: Nuclear expression in most tissues, most abundant in testicular seminiferous duct. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval (datasheet: M01367-2); citrate pH 6.0 is a published alternative for liver tissue microarrays (PMC7919231).
Section 2
What Is the Expected CDC45 Staining Pattern?
CDC45 is nuclear and associates with chromatin (UniProt O75419). In paraffin-section IHC, expect nuclear staining across many tissues, strongest in pachytene spermatocytes within testicular seminiferous ducts; lymphoid populations can also stain (HPA tissue IHC). CDC45 has no transmembrane segment, so a membrane pattern is unexpected (UniProt O75419 topology). HPA rates its tissue profile Enhanced while reporting medium consistency between antibody staining and RNA expression (HPA tissue IHC).
| Strong nuclear staining in testicular pachytene spermatocytes, with weaker staining in nearby cells (HPA tissue IHC). | This matches HPA's high staining level for pachytene spermatocytes and its designation of the seminiferous duct as the most abundant site. Score the identified cell population and its nuclei, rather than assigning one intensity to the entire testis section (HPA tissue IHC; general IHC practice). |
| Predominantly membranous or diffuse cytoplasmic chromogen, with little nuclear signal. | Question the IHC result: UniProt places CDC45 in the nucleus and on chromosomes and reports no transmembrane segment (UniProt O75419). HPA does report additional cytosolic localization by ICC-IF, so an isolated cytoplasmic IF signal needs context; it does not establish a predominantly cytoplasmic tissue-IHC pattern (HPA ICC-IF; HPA tissue IHC). |
| Strong staining in adrenal glandular cells or colon glandular cells (HPA tissue IHC). | HPA reports CDC45 as not detected in those specific cell populations; investigate cross-reactivity or endogenous chromogenic activity before calling them positive (HPA tissue IHC; general IHC practice). This is a cell-type comparison, not a claim that every cell in either tissue must be negative (HPA tissue IHC). |
| Haze across nuclei, cytoplasm, and tissue-free areas. | A pattern extending beyond cells cannot be assigned to nuclear CDC45 (UniProt O75419; general IHC practice). Check staining without primary antibody and review blocking, detection, and washes for background; a diffuse deposit alone does not identify which step caused it (general IHC practice). |
| No convincing nuclear staining in pachytene spermatocytes of a suitable testis section. | This conflicts with HPA's high signal in that population (HPA tissue IHC). Confirm the cells are present, then review the antibody's IHC-P instructions and run a positive control with the same detection workflow; absence of signal alone cannot distinguish specimen, antibody, retrieval, and detection problems (general IHC practice). |
💡Expected CDC45 appearanceCall CDC45 positive when chromogen is concentrated in pachytene spermatocyte nuclei at high intensity, with possible nuclear signal in other cell populations; isolated membrane staining or uniform diffuse deposit is suspect (HPA tissue IHC; UniProt O75419 topology; general IHC practice).
| Cell population and tissue | HPA records high staining in testicular pachytene spermatocytes, medium staining in appendix lymphoid tissue and lymph-node germinal-center cells, and low staining in several other specified populations. Compare matched cells, not whole-section averages (HPA tissue IHC; general IHC practice). |
| Subcellular context | Nuclear and chromatin association supports nuclear IHC scoring (UniProt O75419). In ICC-IF, HPA supports nucleoplasmic and cytosolic localization and approves an additional centrosome location; those observations need their own imaging context (HPA ICC-IF). |
| Evidence and antibody validation | HPA marks the tissue profile Enhanced but describes only medium consistency with RNA data (HPA tissue IHC). It also lists two rabbit polyclonal antibodies, HPA000614 and CAB009459, as IHC Enhanced; agreement strengthens pattern assessment without making every stained cell specific (HPA antibodies; general IHC practice). |
| Isoforms and epitope coverage | UniProt lists three CDC45 isoforms but supplies no antibody epitope here (UniProt O75419). Do not infer that an IHC-validated antibody recognizes every isoform; establish coverage from its own documentation if isoform-specific interpretation matters (general IHC practice). |
| IF/ICC Q: What localization should be checked? | A: Check nucleoplasmic signal first; HPA also reports cytosol and centrosome localization in ICC-IF (HPA ICC-IF). Use the separate IF/ICC guide for imaging conditions; this tissue-IHC pattern does not specify an IF protocol (HPA tissue IHC; HPA ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| Testis control is blank in pachytene spermatocytes. | The expected high-staining population may be absent from the section, or an IHC workflow step may have failed (HPA tissue IHC; general IHC practice). | Verify cell identity and tissue integrity; review the IHC-validated antibody's IHC-P instructions, including its stated retrieval and dilution, then check detection with a positive control (general IHC practice). |
| Chromogen appears mainly at cell borders. | A membrane-dominant pattern conflicts with CDC45's nuclear/chromatin annotation and lack of a transmembrane segment (UniProt O75419). | Recheck morphology and nuclear counterstain; compare with a known-positive testis section and a no-primary control before scoring the border signal as CDC45 (HPA tissue IHC; general IHC practice). |
| Adrenal or colon glandular cells stain strongly. | Those particular populations are listed as not detected; cross-reactivity or endogenous detection activity may contribute (HPA tissue IHC; general IHC practice). | Compare cell identity and staining compartment, run a no-primary control, and assess whether a second IHC-validated antibody reproduces the pattern (HPA tissue IHC; HPA antibodies; general IHC practice). |
| Background obscures nuclear boundaries. | Diffuse staining may reflect nonspecific antibody binding or detection background; appearance alone cannot separate them (general IHC practice). | Inspect the no-primary control, blocking and wash steps, then reassess whether chromogen is confined to identifiable nuclei in an expected population (general IHC practice; HPA tissue IHC). |
| Lymphoid staining seems weaker than testis. | HPA grades appendix lymphoid tissue and lymph-node germinal-center cells medium, versus high for testicular pachytene spermatocytes (HPA tissue IHC). | Identify the lymphoid population before scoring and compare nuclear staining within that population; do not require it to match testis intensity (HPA tissue IHC; general IHC practice). |
| A low-level signal is treated as proof that the assay worked. | HPA records low staining in several specified populations, so a faint result offers a less decisive positive reference than its high-staining testis population (HPA tissue IHC; general IHC practice). | Use identifiable pachytene spermatocytes as the positive tissue reference when available, and judge nuclear localization alongside intensity and control results (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing CDC45 staining
M01367-2 · IHCIHC analysis of CDC45L using anti-CDC45L antibody (M01367-2). CDC45L was detected in paraffin-embedded section of human ovarian cancer tissu
M01367-2 · IHCIHC analysis of CDC45L using anti-CDC45L antibody (M01367-2). CDC45L was detected in paraffin-embedded section of human breast cancer tissue
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000093009-CDC45/tissue/cerebral+cortex#imid_223202
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000093009-CDC45/tissue/colon#imid_223120
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000093009-CDC45/tissue/liver#imid_223121
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000093009-CDC45/tissue/pancreas#imid_2834770
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000614; green = CDC45, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000614; green = CDC45, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CDC45 IHC & IF
🧪Run testis first and expect staining in pachytene spermatocytes (HPA: High). Use adrenal gland glandular cells as a negative tissue (HPA: Not detected); on the testis slide, adjacent cells without staining should provide a background reference, but their CDC45 status must be verified rather than assumed (HPA: High in pachytene spermatocytes).
Positive control tissue: Testis (Pachytene spermatocytes, HPA High)
Negative control tissue: Adrenal gland (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CDC45 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, a concentration-matched mouse IgG isotype control (hero caption: mouse anti-CDC45L), and a biological specificity control such as CDC45 knockout material or peptide competition if the immunizing peptide is available. For the testis DAB assay, quench endogenous peroxidase and check for endogenous biotin background (hero caption: SABC/DAB detection).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected paraffin-section caption does not state the fixative (hero caption: fixative not stated). The demonstrated tissue-IHC workflow uses heat retrieval in EDTA at pH 8.0; retrieval dependence has not been established by a comparison without retrieval (hero caption: EDTA pH 8.0 heat retrieval). Frozen sections have no supplied validation, while ICC-IF images exist for A-431, U-251MG and U2OS; neither is established as easier for this antibody (HPA: ICC-IF image cell lines). In testis, endogenous peroxidase or biotin can complicate SABC/DAB background assessment (hero caption: SABC/DAB detection).
HPA tissue IHC evidence for CDC45
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 |
|---|---|---|---|---|
| Testis | Pachytene spermatocytes | High | Protein (IHC) | HPA → |
| Appendix | Lymphoid tissue | Medium | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Medium | Protein (IHC) | HPA → |
| Tonsil | Non-germinal center cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cerebral cortex | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CDC45 IHC Tips
Troubleshoot CDC45 staining in paraffin sections by checking nuclear localisation, tissue controls and the documented retrieval and detection conditions.
What retrieval conditions should I start with for CDC45 in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 (datasheet M01367-2). The documented paraffin-section example used that retrieval before 10% goat-serum blocking and 2 µg/mL primary antibody overnight at 4°C (datasheet M01367-2). If nuclear staining is weak, first check that sections were fully deparaffinised and that heating, cooling and buffer pH were consistent across the run (standard IHC practice). Compare any retrieval adjustment on adjacent sections using the same detection conditions and a positive tissue control, such as testis with pachytene spermatocytes (HPA: High in pachytene spermatocytes). Score nuclear signal separately from diffuse background (UniProt O75419: nucleus and chromatin association).
How should I troubleshoot fixation-related loss of CDC45 staining?
Target-specific CDC45 sensitivity to fixation is unknown: the selected paraffin-section caption does not state a fixative (datasheet M01367-2). Record the actual fixative, fixation duration and tissue thickness for each block, then compare sections with matched retrieval and detection conditions (standard IHC practice). Use the documented EDTA pH 8.0 retrieval and 2 µg/mL overnight primary incubation as a controlled starting point (datasheet M01367-2). If one block stains poorly, assess tissue preservation and include a separately processed positive control before changing antibody concentration (standard IHC practice). Do not infer a CDC45 fixation effect from its nuclear location, phosphorylation sites or tissue distribution (UniProt O75419: localisation and modified residues; HPA: tissue IHC profile).
Where should convincing CDC45 chromogenic staining appear?
Prioritise nuclear staining because CDC45 localises to the nucleus and associates with chromatin (UniProt O75419: subcellular location). The tissue IHC profile reports nuclear expression in most tissues and its highest abundance in the testicular seminiferous duct (HPA: tissue IHC profile). Use a nuclear counterstain to check whether chromogen overlaps intact nuclei, and score nuclear and extranuclear signal separately (standard IHC practice). If staining appears mainly cytoplasmic, inspect the negative control and detection reagents before calling it CDC45; cytosol is an additional location in cell-based imaging (HPA: subcellular profile). Compare the same compartment across adjacent sections under the documented EDTA pH 8.0 retrieval condition (datasheet M01367-2).
Could an isoform or modified epitope explain uneven CDC45 staining?
CDC45 has 3 annotated isoforms, but the supplied antibody evidence does not identify which sequence its epitope recognises (UniProt O75419: isoforms; datasheet M01367-2: selected caption). Therefore, do not interpret variable chromogenic intensity as isoform-specific staining without an epitope map and an independent validation method (standard IHC practice). Phosphorylation is annotated at residues 130, 144 and 148, but epitope overlap or a staining effect has not been established here (UniProt O75419: modified residues). CDC45 has no annotated transmembrane segment or glycosylation sites, so evaluate the observed signal against its nuclear and chromatin-associated location (UniProt O75419: topology, glycosylation and localisation). Keep retrieval at EDTA pH 8.0 while comparing sections (datasheet M01367-2).
How can IF help check an ambiguous CDC45 IHC pattern?
Use IF as a separate localisation check alongside the chromogenic paraffin-section result; the selected antibody caption documents IHC conditions, not an IF protocol (datasheet M01367-2). Multiplex CDC45 with a marker that identifies the expected cell population, such as pachytene spermatocytes in testis, and confirm overlap with a nuclear counterstain (HPA: High in pachytene spermatocytes; UniProt O75419: nuclear localisation). Choose a fluorophore in a low-autofluorescence channel after examining an unstained section, and include single-label controls (standard IF practice). Because CDC45 has no transmembrane segment and is mainly nuclear, optimise permeabilisation for access to an intracellular nuclear epitope rather than assuming a cell-surface assay (UniProt O75419: topology and localisation). Compare any cytosolic or centrosomal signal with appropriate controls (HPA: subcellular profile).
How do I distinguish CDC45 signal from chromogenic background?
Run a no-primary control through the same secondary, detection and chromogen steps to reveal background unrelated to primary-antibody binding (standard IHC practice). The documented paraffin-section example used a biotinylated goat anti-mouse secondary for 30 minutes at 37°C, followed by an avidin-biotin detection complex and DAB (datasheet M01367-2). Check endogenous peroxidase and biotin contributions when that detection system produces widespread colour; peroxidase blocking is a general chromogenic IHC step (standard IHC practice). Compare background in nuclei, stroma and section edges with the no-primary control before adjusting blocking or washes (standard IHC practice). Retain CDC45 calls for convincing nuclear staining in intact cells (UniProt O75419: nuclear localisation).
How should I quantify CDC45 staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and nuclear scoring rule before comparing sections, because the expected CDC45 pattern is nuclear (HPA: tissue IHC profile; UniProt O75419: localisation). Report the percentage of positive nuclei and, when intensity matters, an H-score from 0–300 using the same intensity thresholds across slides (standard IHC scoring practice). For a spatial question, report positive nuclei per mm² of evaluable tissue and document excluded necrotic or damaged areas (standard IHC practice). Normalise counts to the relevant intact cell population or measured tissue area, and keep retrieval, imaging and threshold settings consistent (standard IHC practice). Record tissue compartment separately because HPA reports different levels across cell populations (HPA: tissue IHC profile).
Which findings make a CDC45-positive IHC result credible?
A credible result places chromogen in intact nuclei, consistent with CDC45's nuclear and chromatin-associated localisation (UniProt O75419: subcellular location). Compare an expected positive population, such as testicular pachytene spermatocytes, with an appropriate low-signal population under the same staining run (HPA: High in pachytene spermatocytes; HPA: selected Not detected populations). Inspect section edges, necrotic areas and poorly preserved cells before scoring, since these can produce misleading staining patterns (standard IHC practice). Check a no-primary control for colour from endogenous enzyme activity or the detection system, especially when using DAB (standard IHC practice). Treat diffuse extranuclear colour cautiously even though additional cytosolic and centrosomal locations are reported in cell-based imaging (HPA: subcellular profile).
Boster reagents
Best CDC45 / Cell division control protein 45 homolog IHC Antibodies
The catalog includes human, mouse, and rat reactive CDC45 antibodies (catalog: reactivity); real IHC images show human paraffin sections with M01367-2 (M01367-2 image captions). No IF image is supplied (catalog: IF images).
Real IHC data
Anti-CDC45L ® Antibody (monoclonal, 6H6)
Cat # M01367-2
M01367-2 is the only SKU with a rendered card; its IHC captions show human ovarian, breast, esophageal squamous, and pancreatic cancer paraffin sections (M01367-2 image captions). M01367 is listed for IHC and IF/ICC in human, mouse, and rat, but has no IHC or IF image in the payload (catalog: M01367 applications, reactivity, and image lists).
Which to pick: For tissue IHC, choose mouse monoclonal M01367-2: its own captions document human paraffin sections, and the fixative is unreported (catalog: M01367-2 host and clone; M01367-2 image captions). For IF/ICC, choose rabbit monoclonal M01367 because those applications are listed, although no IF image is supplied (catalog: M01367 host, clone, applications, and IF images). Both SKUs list human, mouse, and rat reactivity for cross-species planning; the supplied tissue images document human samples only (catalog: reactivity; M01367-2 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O75419 (CDC45_HUMAN, Cell division control protein 45 homolog).
- Human Protein Atlas. CDC45 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. CDC45 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol and centrosome..
- Human Protein Atlas. CDC45 antibody validation summary (2 antibodies).
- Elevated CDC45 Expression Predicts Poorer Overall Survival Prognoses and Worse Immune Responses for Kidney Renal Clear Cell Carcinoma via Single-Cell and Bulk RNA-Sequencing. Biochemical genetics 2024 — PMC11186877.
- Significant role of Psf3 expression in non-small-cell lung cancer. Cancer science 2015 — PMC4714687.
- Expression of Cell Division Cycle Protein 45 in Tissue Microarrays and the CDC45 Gene by Bioinformatics Analysis in Human Hepatocellular Carcinoma and Patient Outcomes. Medical science monitor : international medical journal of experimental and clinical research 2021 — PMC7919231.
- Validation of CDC45 as a novel biomarker for diagnosis and prognosis of gastric cancer. PeerJ 2024 — PMC10956518.
- PubMed PMID:9755170 — UniProt-cited evidence.
- PubMed PMID:9660782 — UniProt-cited evidence.
- PubMed PMID:9724329 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CDC45 (Cell division control protein 45 homolog). Expected localisation Nuclear in most tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet M01367-2). Validated IHC antibody: M01367-2 at 2μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Testis (Pachytene spermatocytes); negative Adrenal gland. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human ovarian cancer tissue; fixative not specified (datasheet M01367-2); 2. Fixation: Image fixative and duration unreported (datasheet M01367-2); 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: EDTA pH 8.0 (datasheet M01367-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet M01367-2); 8. Primary antibody: Mouse monoclonal (clone 6H6) anti-CDC45, 2μg/ml (datasheet M01367-2); 9. Primary incubation: Overnight at 4 °C (datasheet M01367-2). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Staining differs among cell populations within a tissue (HPA tissue IHC); Isoform / epitope: 3 isoforms; confirm epitope coverage (UniProt); Regulation: Cell-cycle associated (UniProt). Evidence: UniProt O75419, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet M01367-2, PMC4714687, PMC7919231. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CDC45/immunohistochemistry-ihc-guide.