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ARPC5 / Actin-related protein 2/3 complex subunit 5 · IHC design guide
Design Immunohistochemistry for ARPC5
This guide helps plan chromogenic ARPC5 IHC in paraffin sections using the reported cytoplasmic tissue pattern (HPA tissue IHC). Testis elongated or late spermatids provide a high-staining reference, but interpret antibody specificity cautiously because the HPA antibody may target more than one gene (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ARPC5 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 tissue staining (HPA tissue IHC); nuclear location annotated (UniProt) | |
| Staining pattern | Several tissue cell types show general cytoplasmic staining (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear 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 consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Staining may include a target from another gene (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended ARPC5 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is supplemented by two published ARPC5 protocols using paraffin sections (PMC9485570; PMC3584917).
| Sample | Paraffin-embedded human colon tissue; fixative not specified (datasheet M02096) |
| Fixation | Image fixative and duration unreported (datasheet M02096); 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 Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear 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 monoclonal (clone CCD-1) anti-ARPC5, 1:50 (datasheet M02096) |
| 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 | ARPC5-positive staining in elongated or late spermatids of testis (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression in several different tissue types. No signal in the no-primary control. |
💡Decision noteStart with this page’s Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min; the rat cortex study provides a citrate pH 6.0 comparison (PMC3584917).
Section 2
What Is the Expected ARPC5 Staining Pattern?
ARPC5 is associated with the cytoplasmic cytoskeleton, cell projections and nucleus, and has no transmembrane segment (UniProt O15511). In paraffin section IHC, expect predominantly cytoplasmic staining across several tissues, with high staining in testicular elongated or late spermatids and medium staining in selected glandular, respiratory epithelial and neuronal cells (HPA tissue IHC). Interpret specificity cautiously: HPA rates its tissue staining Approved but reports low agreement with RNA data and cautions that the antibody targets protein from more than one gene (HPA tissue IHC).
| Cytoplasmic chromogen in elongated or late spermatids; weaker staining in other documented positive cells. | This fits the strongest listed testis signal (High) and the broader cytoplasmic profile. Adrenal, appendix and breast glandular cells, bronchial respiratory epithelium, and listed neuronal cells are Medium comparators (HPA tissue IHC). |
| Predominantly crisp nuclear staining with little cytoplasmic signal, or a uniform outline of every cell. | Reassess specificity and detection: HPA describes general cytoplasmic tissue staining (HPA tissue IHC). UniProt also lists the nucleus, so nuclear signal alone cannot be dismissed as impossible; ARPC5 has no transmembrane segment, although ICC-IF reports membrane and junction staining (UniProt O15511; HPA ICC-IF). |
| Strong signal in a listed undetected cell type, such as adipocytes or lymph node germinal center cells. | That result conflicts with those cell-specific HPA observations (HPA tissue IHC). Consider antibody cross-reactivity or endogenous detection activity, then compare an omission control and tissue morphology (standard IHC practice). HPA's multi-gene targeting caution makes a positive stain insufficient for ARPC5-specific attribution (HPA tissue IHC). |
| Diffuse color across cells, stroma and empty areas, with poorly defined cell boundaries. | Treat this as background until controls support cellular staining; excess primary or detection reagent, incomplete blocking or washing, and chromogen deposits are general IHC possibilities (standard IHC practice). HPA's cell-specific intensity categories do not validate a diffuse haze (HPA tissue IHC). |
| No signal in elongated or late spermatids while the section and counterstain remain interpretable. | An absent signal in this High reference cell population warrants a run check before calling another tissue negative (HPA tissue IHC; standard IHC practice). Check the antibody's documented IHC-P conditions, detection reagents and control section; no ARPC5-specific retrieval or fixation effect is supplied (standard IHC practice). |
💡Expected ARPC5 appearanceA convincing positive is predominantly cytoplasmic staining in the expected cells, strongest in elongated or late spermatids (High) and moderate in listed Medium populations; uniform haze or strong staining in listed undetected cell types needs control review (HPA tissue IHC; standard IHC practice).
| Tissue and cell selection | Compare like cell populations: HPA records High elongated or late spermatids, Medium adrenal, appendix and breast glandular cells, and undetected adipocytes, among other entries. A negative call applies to the specified cell type, not every cell in its tissue (HPA tissue IHC). |
| IHC evidence and specificity | The tissue profile is Approved, while HPA reports low staining–RNA consistency and possible recognition of protein from more than one gene. HPA022013 is Approved for IHC; that status supports use of its observations but does not prove every stained structure contains ARPC5 (HPA tissue IHC; HPA antibodies). |
| Compartment and topology | UniProt lists cytoplasm, cytoskeleton, cell projection and nucleus, with no transmembrane segment (UniProt O15511). HPA's tissue IHC summary is generally cytoplasmic; use that section-level pattern as the primary visual comparator (HPA tissue IHC). |
| Isoforms and processing | UniProt lists 2 isoforms, no signal peptide or propeptide, and a chain beginning at residue 2 (UniProt O15511). Epitope coverage across isoforms is not supplied; do not infer an isoform-specific stain or a shed extracellular pool from this record (UniProt O15511). |
| IF/ICC: what pattern should I expect? | HPA ICC-IF reports vesicles, plasma membrane, cell junctions and cytosol, with images in A-431, U-251MG and U2OS cells (HPA ICC-IF). This is a separate imaging context: those locations do not establish an IHC-P membrane pattern, and HPA031972 has Uncertain ICC status (HPA antibodies). |
| Situation | Likely cause | Next action |
|---|---|---|
| All tissues, including testis, appear negative. | The positive reference may have failed, or the IHC detection run may have failed (HPA tissue IHC; standard IHC practice). | Inspect elongated or late spermatids on the control section, then check the documented IHC-P antibody conditions and detection reagents. Optimize retrieval against a positive control only as a general IHC step; ARPC5-specific retrieval sensitivity is unreported (HPA tissue IHC; standard IHC practice). |
| Weak signal in a tissue expected to be medium. | HPA's Medium designation is cell-specific, and weak contrast can also reflect a general staining-run problem (HPA tissue IHC; standard IHC practice). | Identify the relevant glandular, respiratory epithelial or neuronal population before scoring; compare a same-run testis control and review antibody concentration, detection and counterstain using documented IHC-P conditions (HPA tissue IHC; standard IHC practice). |
| Strong color appears in listed undetected cells. | Non-specific binding or endogenous detection activity may contribute; HPA also cautions about multi-gene targeting (standard IHC practice; HPA tissue IHC). | Verify the cell identity, review primary-omission and detection controls, and apply appropriate endogenous activity blocking for the detection chemistry. Do not assign the stain to ARPC5 solely from color intensity (standard IHC practice; HPA tissue IHC). |
| Brown haze or deposits obscure cellular boundaries. | Background binding, inadequate washing or chromogen precipitation are general IHC causes (standard IHC practice). | Compare the primary-omission control; review blocking, wash steps, reagent concentration and chromogen development. Score only localized cellular signal after the background is resolved (standard IHC practice). |
| Signal is mainly nuclear or outlines cell borders. | A mismatch with HPA's general cytoplasmic IHC profile needs review; UniProt permits nuclear localization, and HPA ICC-IF reports membrane and junction locations in cells (HPA tissue IHC; UniProt O15511; HPA ICC-IF). | Confirm morphology and compartment against controls and the cytoplasmic positive reference. Treat the ICC-IF pattern as context, not proof of the same paraffin-section distribution (standard IHC practice; HPA tissue IHC; HPA ICC-IF). |
| A weak or absent result is being called an ARPC5-negative tissue. | HPA reports low tissue RNA specificity and lists staining by specific cell population; its IHC reliability note also limits certainty (HPA tissue IHC). | Record the tissue, cell type, compartment and intensity, then compare a same-run positive reference. Report an undetected stain as an assay observation rather than proof that the tissue lacks ARPC5 (HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing ARPC5 staining
M02096 · IHCImmunohistochemical analysis of paraffin-embedded human colon, using p16 ARC Antibody.
M02096-1 · IFImmunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized U-2 OS (human osteosarcoma cell line) cells labelin
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000162704-ARPC5/tissue/cerebral+cortex#imid_5785972
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000162704-ARPC5/tissue/colon#imid_5785890
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000162704-ARPC5/tissue/liver#imid_5785899
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000162704-ARPC5/tissue/kidney#imid_5785892
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA031972; green = ARPC5, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA031972; green = ARPC5, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ARPC5 IHC & IF
🧪Run testis first and require staining in elongated or late spermatids (HPA: High). Use adipose tissue as the negative tissue and assess its adipocytes (HPA: Not detected); on the testis slide, count neighboring cells as internal negatives only if they remain at background staining, since their ARPC5 status is not specified by the supplied HPA rows.
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 ARPC5 in A-431, U-251MG, U2OS, with annotated localisation: Vesicles (approved), Plasma membrane (approved), Cell Junctions (approved), Cytosol (approved) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, an isotype control matched to the primary antibody’s host species and clonality, and ARPC5-knockout material as a biological negative (standard IHC practice). For chromogenic testis IHC, quench endogenous peroxidase and check background on the no-primary slide (standard IHC practice).
⚠️Feasibility: No target-specific fixation window, fixation effect, or antigen-retrieval dependency is reported in the supplied evidence; the selected M02096 paraffin-section caption does not state a fixative (selected-SKU caption). Whether frozen sections or IF are easier is unreported; HPA provides ICC-IF images in A-431, U-251MG, and U2OS but no comparison with tissue IHC (HPA subcellular record). In testis, identify the elongated or late spermatids by morphology before scoring their signal against adjacent background (HPA: High in elongated or late spermatids; standard IHC practice).
HPA tissue IHC evidence for ARPC5
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low consistency between antibody staining and RNA expression data. Caution, targets protein from more than one gene.). 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 → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
| Oral mucosa | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ARPC5 IHC Tips
Troubleshoot ARPC5 staining in paraffin section chromogenic IHC by checking retrieval, compartment, specificity and cell level scoring before interpreting intensity.
Which retrieval condition should I start with for ARPC5 paraffin section IHC?
Start with Tris-EDTA at pH 9.0, heated to 95–98 °C for 20 min (page retrieval rule). Let slides cool in the retrieval buffer, then compare staining with a matched section processed identically except for retrieval; keep antibody concentration and detection time fixed (standard IHC practice). The selected antibody has a paraffin section image from human colon, but its caption gives no retrieval method or fixative, so that image does not establish the optimum condition (catalog antibody M02096 caption). If signal remains weak, test a citrate buffer at pH 6.0 as a fallback, recording both signal and tissue preservation (standard IHC practice).
How can I assess whether fixation is masking ARPC5 staining?
Target-specific fixation sensitivity is unknown: the selected human colon caption identifies a paraffin section but does not report its fixative (catalog antibody M02096 caption). Record the actual fixative and fixation duration for each specimen, and compare sections with similar processing history before changing retrieval or antibody concentration (standard IHC practice). If matched material is available, compare a short and a longer fixation condition while holding section thickness, pH 9.0 retrieval and detection constant (standard IHC practice; page retrieval rule). Treat any difference as a result for those specimens and conditions, since neither the reported tissue pattern nor protein features establish an ARPC5-specific fixation effect (HPA tissue IHC; UniProt O15511).
Which staining compartments are plausible for ARPC5, and which should prompt review?
Expect a substantial cytoplasmic signal in tissue sections: ARPC5 is assigned to the cytoplasm and cytoskeleton, cell projections and nucleus, while tissue IHC reports general cytoplasmic expression (UniProt O15511 subcellular; HPA tissue IHC). Plasma membrane, junctional, vesicular and cytosolic signals are also reported by cell imaging, so inspect cell boundaries and cytoplasm at high power before assigning a compartment (HPA subcellular). Nuclear staining can be biologically plausible, but require a crisp cellular pattern and a clean no-primary control before scoring it (UniProt O15511 subcellular; standard IHC practice). Diffuse extracellular deposits or staining confined to torn edges warrant a processing or detection check (standard IHC practice).
Could isoforms or epitope placement explain discordant ARPC5 staining?
ARPC5 has 2 annotated isoforms, but the supplied antibody caption does not specify its epitope or isoform coverage (UniProt O15511 isoforms; catalog antibody M02096 caption). Check the antibody's documented immunogen against both isoform sequences before interpreting a negative section as absence of all ARPC5 protein (standard IHC practice; UniProt O15511 isoforms). The annotated mature chain spans residues 2–151, with N-terminal acetylserine at residue 2 and no annotated transmembrane segment or glycosylation sites; these facts alone do not predict epitope accessibility (UniProt O15511 processing, modified residues, topology and glycosylation). Compare an independent epitope antibody or an orthogonal assay on matched material when isoform specificity matters (standard IHC practice).
How should I check ARPC5 localisation by multiplex IF?
Use IF as a separate validation experiment, since the selected antibody's supplied image documents paraffin section IHC in human colon, without an IF condition (catalog antibody M02096 caption). Pair ARPC5 with a validated marker identifying the cell population under study, and inspect whether cytosolic, junctional or vesicular signal belongs to those marked cells (HPA subcellular; standard IF practice). Choose a far-red fluorophore when the specimen shows strong shorter-wavelength autofluorescence, and include single-stain controls for spectral bleed-through (standard IF practice). For an intracellular epitope, test mild detergent permeabilisation against an unpermeabilised control; for a surface-facing epitope, start without detergent, since the antibody epitope is unspecified and ARPC5 has no transmembrane segment (standard IF practice; UniProt O15511 topology).
What controls distinguish ARPC5 signal from chromogenic background?
Run a no-primary control through the same secondary reagent, peroxidase block and DAB development to reveal detection background in the section (standard IHC practice). Compare staining in the expected cellular compartments with matched tissue morphology, rather than judging a brown field by intensity alone (UniProt O15511 subcellular; standard IHC practice). HPA describes its tissue antibody as approved but cautions that staining and RNA expression have low consistency and that the antibody targets protein from more than one gene; treat unexpected widespread staining as a specificity question (HPA tissue IHC). If background persists, titrate the primary antibody and DAB development separately, then seek an independent antibody or orthogonal readout (standard IHC practice).
How should I score heterogeneous ARPC5 IHC across a section? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because cytoplasmic tissue staining is reported and ARPC5 also has other annotated locations (HPA tissue IHC; UniProt O15511 subcellular). For cytoplasmic staining, record the percentage of positive cells and intensity on a prespecified 0–3 scale; an H-score sums each intensity's percentage contribution and ranges from 0–300 (standard IHC practice). Normalise positive counts to the number of eligible cells, or positive cell density to the measured viable tissue area in mm², using the same rule across specimens (standard IHC practice). Exclude folds, torn edges and necrotic areas by a documented rule before comparing groups (standard IHC practice).
When is an apparent ARPC5-positive cell convincing?
A convincing cell shows reproducible staining in a plausible compartment, with preserved morphology and a clean no-primary control; cytoplasmic staining is the main tissue IHC pattern reported for ARPC5 (HPA tissue IHC; standard IHC practice). Check cell identity: HPA reports high staining in elongated or late spermatids, medium staining in several glandular and neuronal populations, and no detection in listed adipocytes and fibroblasts, so avoid treating every cell type as interchangeable (HPA tissue IHC). Review isolated edge staining, necrotic debris and residual endogenous peroxidase signal before calling a positive result (standard IHC practice). Because HPA cautions about cross-gene antibody reactivity and low staining-to-RNA consistency, confirm consequential findings with independent evidence (HPA tissue IHC; standard IHC practice).
Boster reagents
Best ARPC5 / Actin-related protein 2/3 complex subunit 5 IHC Antibodies
The catalog includes ARPC5 antibodies for paraffin-section IHC and cellular IF, with images from human colon and U-2 OS cells, respectively (M02096 IHC caption; M02096-1 IF caption).
Real IHC data
Anti-p16 ARC ARPC5 Rabbit Monoclonal Antibody
Cat # M02096
Real IF data
Anti-ARPC5 Antibody (Center)
Cat # M02096-1
M02096 has an IHC image from paraffin-embedded human colon; its listed reactivity is human, mouse and rat (M02096 IHC caption; catalog applications/reactivity). M02096-1 has an IF image from paraformaldehyde-fixed, permeabilized human U-2 OS cells; its listed reactivity is human, mouse and rat (M02096-1 IF caption; catalog applications/reactivity).
Which to pick: Choose monoclonal M02096 for tissue IHC: its image documents human paraffin sections, but does not report the fixative (M02096 IHC caption; catalog clone CCD-1). Choose M02096-1 for cellular IF because its image documents that application; its catalog lists IF, but not ICC (M02096-1 IF caption; catalog applications). For work across human, mouse and rat, M02096 lists IHC and IF and all three species, although its IHC image documents human tissue only (M02096 catalog applications/reactivity; M02096 IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O15511 (ARPC5_HUMAN, Actin-related protein 2/3 complex subunit 5).
- Human Protein Atlas. ARPC5 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ARPC5 subcellular location (ICC-IF): Localized to the plasma membrane, cell junctions, vesicles and cytosol..
- Human Protein Atlas. ARPC5 antibody validation summary (2 antibodies).
- A comprehensively prognostic and immunological analysis of actin-related protein 2/3 complex subunit 5 in pan-cancer and identification in hepatocellular carcinoma. Frontiers in immunology 2022 — PMC9485570.
- Effects of hypothyroidism on expression of CRMP2B and ARPC5 during development of the rat frontal cortex. International journal of biological sciences 2013 — PMC3584917.
- ARPC5 acts as a potential prognostic biomarker that is associated with cell proliferation, migration and immune infiltrate in gliomas. BMC cancer 2023 — PMC10548738.
- Identification of Arp2/3 Complex Subunits as Prognostic Biomarkers for Hepatocellular Carcinoma. Frontiers in molecular biosciences 2021 — PMC8299467.
- PubMed PMID:9230079 — UniProt-cited evidence.
- PubMed PMID:9359840 — UniProt-cited evidence.
- PubMed PMID:16710414 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ARPC5 (Actin-related protein 2/3 complex subunit 5). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC); nuclear location annotated (UniProt). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: M02096 at 1:50 (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: Paraffin-embedded human colon tissue; fixative not specified (datasheet M02096); 2. Fixation: Image fixative and duration unreported (datasheet M02096); 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 Tris-EDTA buffer, pH 9.0 , 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 monoclonal (clone CCD-1) anti-ARPC5, 1:50 (datasheet M02096); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Staining may include a target from another gene (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unspecified (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt O15511, HPA tissue IHC · Approved, HPA ICC-IF · plasma membrane, cell junctions, vesicles and cytosol., Datasheet M02096, PMC9485570, PMC3584917. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ARPC5/immunohistochemistry-ihc-guide.