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KPNA5 / Importin subunit alpha-6 · IHC design guide
Design Immunohistochemistry for KPNA5
Plan chromogenic IHC-P for KPNA5 using human testis as a positive control and the catalog antibody at 5 μg/mL (datasheet: IHC-P; HPA tissue IHC). Assess nuclear and cytoplasmic staining in late spermatids alongside the broader cytoplasmic tissue pattern (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
KPNA5 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 | Cytoplasmic in several tissues; nuclear and cytoplasmic in testis (HPA tissue IHC) | |
| Staining pattern | High nuclear and cytoplasmic staining in late spermatids (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Testis+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Only a subset of testis cells shows high staining (HPA tissue IHC) | |
| Regulation | Testis-enhanced RNA expression (HPA tissue RNA) | |
| Isoform / epitope | One 1–539 chain; no annotated isoforms or processing (UniProt) |
Section 1
Recommended KPNA5 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol with a published KPNA5 protocol for FFPE sections (PMC9961296).
| Sample | Tissue sections; selected-image fixative not specified (standard IHC workflow) |
| Fixation | Image fixative and duration unreported (datasheet A09441); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| 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-KPNA5, 5 μg/mL (datasheet A09441) |
| 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 | KPNA5-positive staining in elongated or late spermatids of testis (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several tissues, high nuclear and cytoplasmic abundance in a subset of cells in testis. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); the published protocol also uses citrate pH 6.0 (PMC9961296).
Section 2
What Is the Expected KPNA5 Staining Pattern?
KPNA5 staining in paraffin sections is expected mainly in the cytoplasm across several tissues; a subset of testis cells can show strong nuclear and cytoplasmic staining (HPA tissue IHC). Elongated or late spermatids are the clearest high-staining reference cells (HPA: High in testis). UniProt places KPNA5 in the cytoplasm and reports no transmembrane segment (UniProt O15131). HPA rates the tissue pattern Enhanced, with medium staining–RNA consistency and external verification pending (HPA tissue IHC).
| Strong nuclear and cytoplasmic staining in elongated or late spermatids, with cytoplasmic staining elsewhere. | This matches the reported testis subset and broader tissue profile (HPA tissue IHC). Score the spermatid population separately from adjacent cells: a high result in that population does not imply uniformly high staining throughout testis (HPA: High in elongated or late spermatids). |
| Predominantly nuclear staining outside the reported testis subset, or staining confined to cell membranes. | Treat this as a compartment mismatch requiring investigation. Cytoplasmic staining is the reported general tissue pattern, while high nuclear and cytoplasmic staining is reported in a subset of testis cells (HPA tissue IHC); UniProt reports cytoplasmic localisation and no transmembrane segment (UniProt O15131). |
| Strong staining in adipocytes or cells in endometrial stroma. | These cell populations are reported as not detected (HPA tissue IHC). Consider nonspecific antibody binding or endogenous detection activity before calling the signal KPNA5; compare its distribution with a negative reagent control and an established positive section (standard IHC practice). |
| Diffuse chromogen across cells and surrounding tissue, without a discernible cellular pattern. | This is background rather than the expected cytoplasmic pattern or the reported testis subset pattern (HPA tissue IHC). Check background with appropriate negative controls, then review blocking, antibody dilution, washes and detection reagents (standard IHC practice). |
| No interpretable signal in elongated or late spermatids on a testis section. | The expected high-staining reference population is missing (HPA: High in elongated or late spermatids). First confirm those cells are present and identifiable; then check the antibody and detection run with controls (standard IHC practice). A failed stain alone cannot establish absent KPNA5. |
💡Expected KPNA5 appearanceCall the result positive when elongated or late spermatids show strong nuclear and cytoplasmic staining, with cytoplasmic staining in other reported positive cells; isolated membrane staining or strong staining in reported not-detected cells warrants review (HPA tissue IHC; UniProt O15131 topology).
| Cell population and tissue selection | Testis offers a high-staining reference in elongated or late spermatids (HPA tissue IHC). Glandular cells in appendix and distal tubules in kidney are reported at Medium, while adipocytes are Not detected (HPA tissue IHC). Choose and score the named cells; a tissue-wide average can obscure the reported pattern. |
| Compartment used for scoring | Use cytoplasmic staining as the general tissue expectation, allowing nuclear and cytoplasmic abundance in a subset of testis cells (HPA tissue IHC). UniProt lists KPNA5 as cytoplasmic and reports no transmembrane segment (UniProt O15131). A nuclear signal needs its cell context before interpretation. |
| Strength of pattern evidence | Both listed antibodies, HPA054037 and CAB026464, have Enhanced IHC validation (HPA antibodies). The tissue profile is rated Enhanced, but staining and RNA show medium consistency and external verification is pending (HPA tissue IHC). Interpret an unexpected cell or compartment pattern with controls rather than treating validation as proof for that individual section. |
| IF/ICC comparison | HPA reports supported cytosolic localisation by ICC-IF; both listed antibodies have Supported ICC status (HPA subcellular; HPA antibodies). This supports a cytosolic comparison, while tissue IHC separately reports high nuclear and cytoplasmic abundance in a testis subset (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| The testis positive reference shows no signal. | The section may lack identifiable elongated or late spermatids, or the staining run may have failed (HPA: High in elongated or late spermatids; standard IHC practice). | Identify the relevant cells on the section, then review positive-run controls, primary antibody use and chromogenic detection (standard IHC practice). Repeat before interpreting the result as biological absence. |
| A section shows nuclear-only or membrane-confined staining. | Its distribution conflicts with the general cytoplasmic tissue profile and, for membrane confinement, the reported topology (HPA tissue IHC; UniProt O15131). | Check cell identity and compare a testis reference and negative control. Record nuclear and cytoplasmic staining separately, since the reported testis subset can show both (HPA tissue IHC; standard IHC practice). |
| Adipocytes stain strongly. | Adipocytes are reported as Not detected; nonspecific binding or endogenous detection activity is possible (HPA tissue IHC; standard IHC practice). | Compare the same cell population with a negative reagent control. Review blocking and detection background before scoring it as KPNA5 (standard IHC practice). |
| Diffuse colour obscures cell boundaries. | Excess background can prevent assessment of the expected cellular distribution (HPA tissue IHC; standard IHC practice). | Inspect negative controls and review blocking, primary antibody dilution, washing and chromogen development (standard IHC practice). Reassess localisation only when cells remain distinguishable. |
| A reported Medium population appears weak or patchy. | HPA reports Medium staining for specified cells, not uniform staining of every cell in the tissue; its tissue profile also has medium staining–RNA consistency (HPA tissue IHC). | Score the named population, such as appendix glandular cells or kidney distal tubules, and compare it with the testis high-staining reference within the same staining workflow (HPA tissue IHC; standard IHC practice). |
| What localisation should an IF/ICC image show? | HPA's ICC-IF summary reports supported cytosolic localisation; tissue IHC also describes a nuclear and cytoplasmic testis subset (HPA subcellular; HPA tissue IHC). | Interpret an IF/ICC image primarily against the supported cytosolic pattern (HPA subcellular). Keep the tissue-specific testis observation separate when comparing results across applications (HPA tissue IHC). |
Experiment evidence
IHC & IF images showing KPNA5 staining
A09441 · IHCImmunohistochemistry of KPNA5 in human testis tissue with KPNA5 antibody at 5 μg/mL.
A09441-1 · IFImmunocytochemistry of KPNA5 in EL4 cells with KPNA5 antibody at 5 μg/mL.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000196911-KPNA5/tissue/cerebral+cortex#imid_6678776
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000196911-KPNA5/tissue/colon#imid_15291272
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000196911-KPNA5/tissue/liver#imid_15291273
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000196911-KPNA5/tissue/kidney#imid_15291266
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB026464; green = KPNA5, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB026464; green = KPNA5, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for KPNA5 IHC & IF
🧪Run testis first and expect staining in elongated or late spermatids (HPA: High in elongated or late spermatids). Use adipose tissue as the negative tissue and assess adipocytes (HPA: Not detected in adipocytes); on the testis slide, treat neighboring cells without visible signal as internal background references, without assuming a named cell type is KPNA5-negative.
Positive control tissue: Testis (Elongated or late spermatids, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show KPNA5 in A-431, U-251MG, U2OS, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, an isotype control matched to the primary antibody’s host species and isotype, and a KPNA5-knockout specimen or peptide-block control where available. For chromogenic testis IHC, quench endogenous peroxidase and check the control slide for residual background.
⚠️Feasibility: A target-specific fixation window and retrieval dependency are unreported in the supplied evidence; the selected A09441 testis IHC caption reports 5 μg/mL but does not state a fixative (selected-SKU tissue-IHC caption). The evidence does not establish whether frozen sections or IF would be easier; ICC-IF places KPNA5 in the cytosol (HPA: Cytosol, supported). In testis, use morphology and counterstain to score the closely grouped spermatids identified as the positive population (HPA: High in elongated or late spermatids).
HPA tissue IHC evidence for KPNA5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Pending external verification.). 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 | Elongated or late spermatids | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | Medium | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Endometrium | Cells in endometrial stroma | Not detected | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Fallopian tube | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced KPNA5 IHC Tips
Troubleshoot KPNA5 staining in paraffin sections by checking retrieval, controls, cell type and compartment before interpreting chromogenic signal.
How should I retrieve KPNA5 in paraffin sections with weak staining?
Start with citrate buffer at pH 6.0 for heat-induced epitope retrieval at 95–98 °C for 20 min (page retrieval rule: cytoplasmic antigen). If staining remains weak, compare a small retrieval time series on adjacent sections while keeping antibody concentration and detection constant; include human testis as a positive control (selected A09441 tissue-IHC caption: testis staining at 5 μg/mL). Excess retrieval can damage section morphology or increase background, so assess both signal and tissue preservation (standard IHC practice). Score elongated or late spermatids separately from surrounding cells because their reported expression is high (HPA: testis, elongated or late spermatids high).
Could fixation explain weak or uneven KPNA5 staining?
KPNA5-specific sensitivity to fixation is unknown: the selected human testis IHC caption does not state a fixative (selected A09441 tissue-IHC caption: fixative not stated). For paraffin-section troubleshooting, record the actual fixative, fixation duration and processing history for each specimen, then compare similarly processed sections in the same run (standard IHC practice). Check morphology and staining across the section before changing retrieval, since uneven processing can complicate interpretation (standard IHC practice). Use the documented testis staining at 5 μg/mL as evidence of tissue IHC staining, without treating it as a validated fixation condition (selected A09441 tissue-IHC caption).
Should nuclear KPNA5 staining in testis be considered unexpected?
Assess both compartments in testis: UniProt lists KPNA5 as cytoplasmic, while HPA reports high nuclear and cytoplasmic abundance in a subset of testis cells (UniProt O15131: subcellular location; HPA: tissue profile). Outside that context, compare nuclear signal with cytoplasmic signal in the same identifiable cell type and check whether the pattern repeats across sections (standard IHC practice). HPA subcellular imaging supports cytosolic localisation, so isolated widespread nuclear staining deserves additional controls before interpretation (HPA: subcellular localisation, cytosol supported). KPNA5 participates in nuclear protein import, but that function alone does not establish the compartment responsible for a particular chromogenic signal (UniProt O15131: function; standard IHC interpretation).
How can epitope choice affect KPNA5 IHC interpretation?
Check the antibody’s stated immunogen or mapped epitope before attributing a staining difference to KPNA5; the supplied record does not locate the catalog antibody’s epitope (supplied antibody evidence: epitope unspecified). KPNA5 has a 1–60 IBB domain and a single listed 1–539 chain, with 0 isoforms in this record (UniProt O15131: domain, processing and isoforms). The record lists 0 glycosylation sites and 0 modified residues, which does not demonstrate that fixation or retrieval preserves every antibody epitope (UniProt O15131: PTM annotations; standard IHC practice). If an alternate antibody gives a different pattern, compare its documented epitope and staining controls before invoking an isoform explanation (standard IHC practice).
How should I check a KPNA5 IHC pattern by immunofluorescence?
Treat IF as a separate optimisation and pair KPNA5 with a marker that identifies the expected elongated or late spermatid population in testis (HPA: testis, elongated or late spermatids high; standard IF practice). Select fluorophores after inspecting unstained tissue in each imaging channel, favouring channels with less tissue autofluorescence, and include single-stain controls for spectral spillover (standard IF practice). KPNA5 has no transmembrane segment and is supported as cytosolic, so test mild permeabilisation for access to intracellular epitopes while preserving morphology (UniProt O15131: topology; HPA: cytosol supported; standard IF practice). The catalog caption documents tissue IHC at 5 μg/mL, not an IF dilution or fixation condition (selected A09441 tissue-IHC caption).
What should I check when KPNA5 chromogenic staining is diffuse?
Run a no-primary control alongside testis and inspect whether diffuse colour persists after the usual peroxidase block and DAB detection steps (standard chromogenic IHC practice; HPA: testis enrichment). If colour appears without primary antibody, examine endogenous enzyme activity, detection reagents and washing before assigning it to KPNA5 (standard chromogenic IHC practice). If only the primary-treated section is diffuse, titrate antibody concentration and check blocking, retrieval intensity and incubation conditions against a same-run control (standard IHC practice). Compare the final pattern with cytoplasmic expression across several tissues and the reported high staining in elongated or late spermatids, while accounting for the testis subset with nuclear staining (HPA: tissue profile and testis).
How should I quantify KPNA5 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring: report elongated or late spermatids separately in testis and record nuclear and cytoplasmic signal independently where visible (HPA: testis staining; standard IHC scoring practice). For intensity, use an H-score with 0–3 intensity grades and a 0–300 range; alternatively report the percentage of positive cells or positive-cell density per mm² (standard IHC scoring practice). Normalise cell counts to the number of evaluable cells in that population, or area measurements to evaluable tissue area, excluding damaged regions by a preset rule (standard IHC scoring practice). Keep staining run, imaging settings and positivity threshold consistent, and report the threshold used (standard IHC scoring practice).
How can I distinguish genuine KPNA5 staining from artefact?
Give most weight to reproducible staining in the expected cell and compartment: elongated or late spermatids stain highly, and a testis subset can show both nuclear and cytoplasmic signal (HPA: testis tissue IHC profile). Check whether apparent positives instead cluster at section edges or necrotic areas, or persist in a no-primary control after chromogenic detection (standard IHC artefact checks). Strong staining confined to an unexpected compartment or cell population needs corroboration with another control or method before attribution to KPNA5 (UniProt O15131: cytoplasm; HPA: cytosol supported; standard IHC practice). HPA rates tissue staining as Enhanced but reports medium RNA–protein consistency and pending external verification, so retain that uncertainty when drawing conclusions (HPA: tissue reliability).
Boster reagents
Best KPNA5 / Importin subunit alpha-6 IHC Antibodies
Two anti-KPNA5 antibodies have IHC data from human testis tissue and IF/ICC data from EL4 cells, respectively (catalog: A09441 and A09441-1 image captions).
Real IHC data
Anti-Importin subunit alpha-6 KPNA5 Antibody
Cat # A09441
Real IF data
Anti-Importin subunit alpha-6 KPNA5 Antibody
Cat # A09441-1
A09441 is listed for human IHC-P, with an IHC image of human testis tissue at 5 μg/mL (catalog: A09441 applications, reactivity, image caption). A09441-1 is listed for human and mouse IF/ICC, with an ICC image of EL4 cells at 5 μg/mL (catalog: A09441-1 applications, reactivity, image caption).
Which to pick: Choose A09441, listed as polyclonal, for human paraffin-section IHC; its image shows human testis IHC at 5 μg/mL, and the fixative is unreported (catalog: A09441 dilution_raw, applications, reactivity, image caption). Choose A09441-1 for IF/ICC or work involving human and mouse reactivity; it is listed for both species and those applications, while its clonality is unreported (catalog: A09441-1 reactivity, applications, clone). Its EL4 image documents ICC at 5 μg/mL, and IHC-P is absent from its application list (catalog: A09441-1 image caption, applications).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O15131 (IMA6_HUMAN, Importin subunit alpha-6).
- Human Protein Atlas. KPNA5 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. KPNA5 subcellular location (ICC-IF): Localized to the cytosol..
- Human Protein Atlas. KPNA5 antibody validation summary (2 antibodies).
- LncRNA SNHG6 Upregulates KPNA5 to Overcome Gemcitabine Resistance in Pancreatic Cancer via Sponging miR-944. Pharmaceuticals (Basel, Switzerland) 2023 — PMC9961296.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:14574404 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for KPNA5 (Importin subunit alpha-6). Expected localisation Cytoplasmic in several tissues; nuclear and cytoplasmic in testis (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A09441 at 5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Testis (Elongated or late spermatids); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Tissue sections; selected-image fixative not specified (standard IHC workflow); 2. Fixation: Image fixative and duration unreported (datasheet A09441); 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-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 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-KPNA5, 5 μg/mL (datasheet A09441); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Only a subset of testis cells shows high staining (HPA tissue IHC); Isoform / epitope: One 1–539 chain; no annotated isoforms or processing (UniProt); Regulation: Testis-enhanced RNA expression (HPA tissue RNA). Evidence: UniProt O15131, HPA tissue IHC · Enhanced, HPA ICC-IF · cytosol., Datasheet A09441, PMC9961296. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/KPNA5/immunohistochemistry-ihc-guide.