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CDK5RAP3 / CDK5 regulatory subunit-associated protein 3 · IHC design guide
Design Immunohistochemistry for CDK5RAP3
Plan CDK5RAP3 IHC in paraffin sections with an IHC-validated antibody diluted 1:100–1:300 (datasheet A05770-1). Use the observed cytoplasmic pattern and cell-specific staining levels to choose controls and interpret results (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CDK5RAP3 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 | General cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Strong cytoplasmic staining in bronchial respiratory epithelium (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 8.0 HIER, heat-mediated (datasheet A05770-1) | |
| Positive control | Bronchus+4 more · see all | |
| Negative control | Adipose tissue+2 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific) | |
| Caveat | IHC signal varies by cell type despite low RNA tissue specificity (HPA tissue IHC) | |
| Regulation | Ubiquitously expressed; no inducer specified (UniProt) | |
| Isoform / epitope | 4 isoforms; isoform epitope coverage is unknown (UniProt) |
Section 1
Recommended CDK5RAP3 IHC & IF Protocols
The catalog antibody protocol (datasheet A05770-1) is followed by 3 published CDK5RAP3 IHC protocols (PMC6141336; PMC6839262; PMC6466961).
| Sample | Paraffin-embedded Human brain tissue; fixative not specified (datasheet A05770-1) |
| Fixation | Image fixative and duration unreported (datasheet A05770-1); 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 8.0 (datasheet A05770-1); 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-CDK5RAP3, 1:100-1:300 (datasheet A05770-1) |
| 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 | CDK5RAP3-positive staining in respiratory epithelial cells of bronchus (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated Tris-EDTA, pH 8.0 (datasheet A05770-1). Citrate, pH 6.0, is a published alternative (PMC6141336; PMC6839262).
Section 2
What Is the Expected CDK5RAP3 Staining Pattern?
CDK5RAP3 should show mainly cytoplasmic staining in paraffin sections, with intensity judged by cell type rather than assumed uniform across a tissue (HPA: general cytoplasmic expression; Supported IHC reliability, with medium consistency against RNA). High staining is reported in respiratory epithelium, type II alveolar cells, and selected endocrine and glandular cells (HPA: tissue IHC). Its ER association does not imply a transmembrane staining pattern (UniProt Q96JB5: ER tethering; no transmembrane segment).
| Cytoplasmic staining in bronchial respiratory epithelium or lung type II alveolar cells. | This matches reported High staining in those cell populations (HPA: bronchus and lung tissue IHC). Judge the identified cells against their neighbors; widespread expression does not require every cell to stain equally (UniProt Q96JB5: ubiquitous expression; HPA: cell-specific levels). |
| Predominantly nuclear, surface-rim, or extracellular staining with little cytoplasmic signal. | Review as a compartment mismatch against the general tissue IHC pattern (HPA: general cytoplasmic expression). A nuclear component alone is possible (UniProt Q96JB5: nucleus); persistent rim or extracellular signal calls for controls because no transmembrane segment is annotated (UniProt Q96JB5: topology; general IHC practice). |
| Strong staining in adipocytes or skeletal-muscle myocytes while expected positive cells are faint. | Consider nonspecific binding or endogenous detection activity and check controls (HPA: adipocytes and myocytes not detected by tissue IHC; general IHC practice). This is a comparison of cell types, not proof that an entire tissue lacks CDK5RAP3 (UniProt Q96JB5: broad tissue expression). |
| Colour spreads across stroma, empty spaces, or most cells without clear cytoplasmic boundaries. | Treat this as diffuse background until controlled (general IHC practice). It obscures the cell-specific pattern needed to interpret high respiratory, endocrine, or glandular staining (HPA: tissue IHC); apparent intensity alone does not identify the antigen (general IHC practice). |
| No visible signal in a bronchus or lung section containing the relevant cells. | An absent signal conflicts with reported High staining in bronchial respiratory epithelium or type II alveolar cells (HPA: tissue IHC). Check that those cells are present, then assess the staining run and antibody conditions using a positive-control section (general IHC practice). |
💡Expected CDK5RAP3 appearanceCall a positive result when identifiable respiratory epithelial or type II alveolar cells show clear, relatively strong cytoplasmic reaction product (HPA: High; general cytoplasmic IHC); diffuse extracellular colour or dominant cell-surface outlines are suspect (HPA: tissue profile; UniProt Q96JB5: no transmembrane segment).
| Cell identity and tissue context | HPA reports High staining in bronchial respiratory epithelium, colon and duodenum endocrine cells, and epididymal and gallbladder glandular cells, but low staining in hepatocytes (HPA: tissue IHC). Score the named cell population, since a tissue-wide average can hide these differences (general IHC practice). |
| Subcellular interpretation | The tissue IHC profile is generally cytoplasmic (HPA: tissue IHC). ER tethering, cytoplasm, centrosome, cytoskeleton, and nucleus are annotated (UniProt Q96JB5: subcellular locations); section-level IHC should not be expected to resolve every annotated structure (general IHC practice). |
| Antibody evidence | HPA022141, HPA022882, and HPA027883 each have Supported IHC status (HPA: antibody validation). The tissue profile itself has Supported reliability with medium staining–RNA consistency (HPA: tissue IHC), so unexpected patterns still need local controls (general IHC practice). |
| Isoforms and epitope scope | Four isoforms are annotated, and isoform 3 is reported in kidney, liver, skeletal muscle, and placenta (UniProt Q96JB5: isoforms and tissue specificity). The supplied evidence gives no antibody epitope or isoform coverage; an IHC result cannot assign an isoform from location or intensity alone (UniProt Q96JB5: isoforms; general IHC practice). |
| IF/ICC Q&A: what localization is expected? | Mainly cytosolic signal, with possible nucleolar and vesicular signal (HPA: supported cytosol; approved additional locations). This is the separate ICC-IF observation; use the cytoplasmic tissue IHC profile when judging paraffin-section chromogenic staining (HPA: subcellular ICC-IF and tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in the intended positive control. | The relevant cell population may be absent, or the IHC run may have failed (HPA: High bronchial respiratory and type II alveolar staining; general IHC practice). | Confirm cell identity on the section; run a known-positive section alongside the test and review primary-antibody, retrieval, and detection steps as general IHC checks (general IHC practice). |
| Weak signal where HPA reports high staining. | Signal may be below the detection threshold, or comparison may include the wrong cells (HPA: cell-specific High levels; general IHC practice). | Compare matched cell populations and review the catalog antibody's validated IHC-P conditions; adjust retrieval or detection only within an independently controlled IHC workflow (general IHC practice). |
| Strong signal appears in adipocytes or skeletal-muscle myocytes. | Those cells are reported as not detected by tissue IHC, so nonspecific staining or endogenous detection activity is plausible (HPA: tissue IHC; general IHC practice). | Compare a section without primary antibody and check blocking and chromogen controls before assigning the signal to CDK5RAP3 (general IHC practice). |
| The slide shows mainly surface-rim or extracellular colour. | This conflicts with general cytoplasmic tissue staining and lacks support from a transmembrane annotation (HPA: tissue IHC; UniProt Q96JB5: topology). | Inspect tissue edges and control sections for deposited chromogen or nonspecific signal; confirm that true positive cells retain cytoplasmic staining (general IHC practice; HPA: tissue IHC). |
| Nuclear colour dominates with little cytoplasmic staining. | Nuclear localization is annotated, but it does not match the predominant tissue IHC profile on its own (UniProt Q96JB5: nucleus; HPA: general cytoplasmic expression). | Compare a positive tissue control and a section without primary antibody; interpret nuclear signal cautiously if the expected cytoplasmic pattern is absent (general IHC practice; HPA: tissue IHC). |
| Diffuse colour prevents cell-level scoring. | Background from antibody binding or detection chemistry can obscure the reported cell-specific distribution (general IHC practice; HPA: tissue IHC). | Check a section without primary antibody, blocking, washes, and detection conditions; score only sections where cellular cytoplasm can be distinguished from surrounding background (general IHC practice). |
Experiment evidence
IHC & IF images showing CDK5RAP3 staining
A05770-1 · IHCImmunohistochemical analysis of paraffin-embedded Human brain. Antibody was diluted at 1:100 (4° overnight). High-pressure and temperature T
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000108465-CDK5RAP3/tissue/cerebral+cortex#imid_5983528
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000108465-CDK5RAP3/tissue/colon#imid_5983454
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000108465-CDK5RAP3/tissue/liver#imid_5983455
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000108465-CDK5RAP3/tissue/kidney#imid_5983448
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA022141; green = CDK5RAP3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA022141; green = CDK5RAP3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CDK5RAP3 IHC & IF
🧪Run bronchus first; respiratory epithelial cells should stain strongly (HPA: High in bronchus respiratory epithelial cells). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the bronchus slide, assess non-epithelial cells for background staining rather than assuming they are established target-negative cells.
Positive control tissue: Bronchus (Respiratory epithelial cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CDK5RAP3 in A-431, U-251MG, U2OS, A-549, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and concentration-matched isotype controls matched to the primary antibody’s host species and clonality, plus a knockout sample or immunogen-peptide block; the selected antibody caption documents peptide preabsorption as its negative control (caption: A05770-1). For bronchus IHC, quench endogenous peroxidase and check inflammatory cells for residual chromogen signal (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported, and the selected paraffin-section caption does not state a fixative (caption: A05770-1). The caption uses high-pressure, high-temperature Tris-EDTA retrieval at pH 8.0 and a 1:100 antibody dilution overnight at 4°C, but provides no comparison establishing retrieval dependence (caption: A05770-1). Relative ease of frozen sections or IF is unreported; bronchial inflammatory cells can produce endogenous-peroxidase background in chromogenic IHC (standard IHC practice).
HPA tissue IHC evidence for CDK5RAP3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — 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 |
|---|---|---|---|---|
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Colon | Endocrine cells | High | Protein (IHC) | HPA → |
| Duodenum | Endocrine cells | High | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
Section 3
Advanced CDK5RAP3 IHC Tips
Troubleshoot CDK5RAP3 chromogenic IHC by checking retrieval, compartment, cell type and controls before comparing staining across sections.
What should I change when CDK5RAP3 staining is weak after antigen retrieval?
Start with heat-mediated retrieval in Tris-EDTA at pH 8.0 for this catalog antibody (datasheet A05770-1). Its paraffin-section image used high pressure and temperature with that buffer, followed by a 1:100 primary dilution overnight at 4 °C; the caption does not specify retrieval duration (datasheet A05770-1). Keep section thickness, heating equipment and detection conditions consistent while comparing a short and longer retrieval cycle on adjacent sections (general IHC practice). If staining remains weak, test an alternative retrieval buffer as a fallback alongside the stated method, with a known positive control in each run (general IHC practice). Judge improvement by cytoplasmic signal in expected cells and stable background, rather than by stronger staining alone (HPA tissue IHC; general IHC practice).
Could fixation explain variable CDK5RAP3 staining between paraffin sections?
Target-specific fixation sensitivity is unknown: the catalog image identifies paraffin-embedded human brain but does not state its fixative or fixation duration (datasheet A05770-1). Record fixative, time to fixation, fixation duration and section processing for each specimen before attributing a staining difference to CDK5RAP3 biology (general IHC practice). Compare adjacent sections processed together, using the same Tris-EDTA pH 8.0 retrieval and matched detection settings (datasheet A05770-1; general IHC practice). Include a section with established staining in every run to separate specimen variation from run variation (general IHC practice). Do not infer a fixation effect from tissue-expression patterns or the protein’s annotated locations (HPA tissue IHC; UniProt Q96JB5 subcellular location).
How should I assess nuclear or punctate CDK5RAP3 staining in IHC?
Assess diffuse cytoplasmic staining first, because the tissue profile describes general cytoplasmic expression (HPA tissue IHC). Nuclear staining can be biologically plausible because CDK5RAP3 is annotated in the nucleus, while its association with the endoplasmic reticulum and cytoskeleton can complicate interpretation of puncta in chromogenic sections (UniProt Q96JB5 subcellular location). Record cytoplasmic, nuclear and punctate signal separately instead of collapsing them into one positive score (general IHC practice). Compare each pattern with matched negative controls and the same cell type across adjacent sections (general IHC practice). Resolve unexpected nuclear dominance with an independent antibody or orthogonal evidence before assigning a biological explanation (general IHC practice).
Can this IHC antibody distinguish CDK5RAP3 isoforms or a modified epitope?
Do not assign an IHC signal to one splice form without an epitope map: CDK5RAP3 has 4 annotated isoforms, and the supplied catalog caption does not identify the recognized sequence (UniProt Q96JB5 isoforms; datasheet A05770-1). Isoform 3 is reported in kidney, liver, skeletal muscle and placenta, but tissue distribution alone cannot establish which isoform a stained cell contains (UniProt Q96JB5 tissue specificity; general IHC practice). The record lists no annotated glycosylation sites or modified residues, which does not validate modification-specific staining (UniProt Q96JB5 features). Obtain the immunogen sequence and compare it with isoform sequences before claiming specificity (general IHC practice). Treat peptide pre-absorption as evidence of immunogen-dependent staining, not proof of isoform specificity (datasheet A05770-1; general IHC practice).
How can I investigate CDK5RAP3 localisation by IF alongside this IHC guide?
Use IF as a separate assay to examine the cytosolic pattern and possible nucleolar or vesicular signal reported in cell images (HPA subcellular). For tissue multiplexing, pair CDK5RAP3 with a validated marker for an expected positive cell type, such as alveolar type II cells, and confirm each channel with single-stain controls (HPA tissue IHC; general IF practice). Choose a fluorophore channel with low tissue autofluorescence and inspect an unstained section before interpreting weak puncta (general IF practice). CDK5RAP3 has no annotated transmembrane segment but associates with the endoplasmic reticulum; the antibody’s epitope orientation is unspecified (UniProt Q96JB5 topology and subcellular location; datasheet A05770-1). Compare mild and stronger permeabilisation to establish access to the recognized epitope while checking structural preservation (general IF practice).
What should I check when the CDK5RAP3 DAB stain looks diffuse or nonspecific?
Run a no-primary control and inspect it for endogenous enzyme signal before changing the CDK5RAP3 antibody concentration (general IHC practice). Apply a peroxidase block before DAB detection, and check whether background follows tissue edges, folds or damaged regions (general IHC practice). The catalog image reports a 1:100 primary dilution overnight at 4 °C and a peptide-preabsorbed negative control; those details provide a comparison point, not a universal optimum (datasheet A05770-1). If the no-primary control is clean, titrate the primary antibody and adjust blocking and washes while keeping retrieval constant (general IHC practice). Evaluate whether the remaining signal follows the reported general cytoplasmic pattern and expected cells (HPA tissue IHC).
How should I quantify CDK5RAP3 staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because the reported tissue pattern is generally cytoplasmic and expression varies by cell type (HPA tissue IHC). For each annotated cell population, report the percentage of positive cells or a cytoplasmic H-score from 0–300, using consistent intensity thresholds across batches (general IHC practice). If counting stained cells per area, express density per mm² of viable tissue and exclude folds, necrosis and empty space by a fixed rule (general IHC practice). Normalise cell counts to the number of eligible cells, or area-based counts to evaluated viable area, rather than comparing raw DAB totals (general IHC practice). Retain separate nuclear scores when that compartment is a stated research endpoint (UniProt Q96JB5 subcellular location; general IHC practice).
How can I distinguish genuine CDK5RAP3 signal from staining artefact?
A credible result has reproducible signal in the expected compartment and cell population: tissue staining is generally cytoplasmic, with high signal reported in bronchial respiratory epithelial cells and lung alveolar type II cells (HPA tissue IHC). Treat strong staining restricted to tissue edges, folds or necrosis as suspect until it persists in well-preserved regions (general IHC practice). Check no-primary and peroxidase-block controls when diffuse DAB signal could reflect detection background (general IHC practice). Nuclear signal merits separate review because nuclear localisation is annotated, although it is not the general tissue pattern (UniProt Q96JB5 subcellular location; HPA tissue IHC). The peptide-preabsorbed image supports immunogen-dependent staining in the illustrated paraffin section, but does not establish specificity in every tissue or compartment (datasheet A05770-1; general IHC practice).
Boster reagents
Best CDK5RAP3 / CDK5 regulatory subunit-associated protein 3 IHC Antibodies
The catalog antibody A05770-1 has IHC data from paraffin-embedded human brain (catalog image caption) and lists IF as an application, with human, mouse, and rat reactivity (catalog applications/reactivity).
Real IHC data
Anti-HSF-27 CDK5RAP3 Antibody
Cat # A05770-1
A05770-1 will render with its human brain paraffin-section IHC figure, acquired at 1:100 with Tris-EDTA retrieval (catalog image caption). IF is listed for A05770-1, but no IF image is supplied (catalog applications/IF image alts).
Which to pick: For tissue IHC, choose A05770-1: its own caption documents paraffin-embedded human brain, Tris-EDTA retrieval, and an immunogen peptide pre-absorption control; the fixative is unreported (catalog image caption). For IF, A05770-1 lists IF and a 1:50 dilution, but ICC use and an IF specimen are unreported (catalog applications/dilution/IF image alts). For cross-species work, A05770-1 is a rabbit polyclonal with listed human, mouse, and rat reactivity; its IHC figure documents human tissue only (catalog host/dilution/reactivity/image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q96JB5 (CK5P3_HUMAN, CDK5 regulatory subunit-associated protein 3).
- Human Protein Atlas. CDK5RAP3 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. CDK5RAP3 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoli and vesicles..
- Human Protein Atlas. CDK5RAP3 antibody validation summary (3 antibodies).
- Low expression of CDK5RAP3 and DDRGK1 indicates a poor prognosis in patients with gastric cancer. World journal of gastroenterology 2018 — PMC6141336.
- CDK5RAP3 inhibits angiogenesis in gastric neuroendocrine carcinoma by modulating AKT/HIF-1α/VEGFA signaling. Cancer cell international 2019 — PMC6839262.
- CDK5RAP3 Participates in Autophagy Regulation and Is Downregulated in Renal Cancer. Disease markers 2019 — PMC6466961.
- Endoplasmic reticulum stress-related genes as prognostic and immunogenic biomarkers in prostate cancer. European journal of medical research 2024 — PMC11031923.
- PubMed PMID:12054757 — UniProt-cited evidence.
- PubMed PMID:12737517 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CDK5RAP3 (CDK5 regulatory subunit-associated protein 3). Expected localisation General cytoplasmic staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 8.0 HIER, heat-mediated (datasheet A05770-1). Validated IHC antibody: A05770-1 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Bronchus (Respiratory epithelial cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded Human brain tissue; fixative not specified (datasheet A05770-1); 2. Fixation: Image fixative and duration unreported (datasheet A05770-1); 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 8.0 (datasheet A05770-1); 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-CDK5RAP3, 1:100-1:300 (datasheet A05770-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific); Caveat: IHC signal varies by cell type despite low RNA tissue specificity (HPA tissue IHC); Isoform / epitope: 4 isoforms; isoform epitope coverage is unknown (UniProt); Regulation: Ubiquitously expressed; no inducer specified (UniProt). Evidence: UniProt Q96JB5, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the…, Datasheet A05770-1, PMC6141336, PMC6839262. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CDK5RAP3/immunohistochemistry-ihc-guide.