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USP24 / Ubiquitin carboxyl-terminal hydrolase 24 · IHC design guide
Design Immunohistochemistry for USP24
Plan chromogenic paraffin IHC for USP24 around the cytoplasmic staining reported in most cell types (HPA tissue IHC). Compare high staining in hepatocytes with undetected staining in adipocytes (HPA tissue IHC), and titrate the IHC-validated antibody across 1:100–1:300 (datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
USP24 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 staining in most cell types (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic in most cell types; texture unspecified (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep formalin fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes can be unstained despite the broad profile (HPA tissue IHC) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | No annotated isoforms, signal peptide or processed chain (UniProt) |
Section 1
Recommended USP24 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by three published USP24 IHC protocols for paraffin sections (PMC10394155; PMC13433775; PMC11461744).
| Sample | Paraffin-embedded human breast carcinoma tissue; fixative not specified (datasheet A08241) |
| Fixation | Image fixative and duration unreported (datasheet A08241); 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-USP24, 1:100 - 1:300 (datasheet A08241) |
| 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 | USP24-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most cell types. No signal in the no-primary control. |
💡Decision noteStart with this page’s citrate pH 6.0 heat retrieval at 95–98 °C for 20 min; two published protocols also specify citrate-based retrieval (PMC13433775; PMC11461744).
Section 2
What Is the Expected USP24 Staining Pattern?
USP24 is reported as cytoplasmic in most cell types by tissue IHC, with high staining in several glandular populations, hepatocytes and trophoblastic cells (HPA tissue IHC). HPA rates this tissue profile Approved, with medium agreement between staining and RNA data and external verification pending (HPA tissue IHC). UniProt lists no transmembrane segment or annotated subcellular location, so the IHC pattern should be judged against the observed HPA tissue images (UniProt Q9UPU5 topology; HPA tissue IHC).
| Clear cytoplasmic staining in colon glandular cells or liver hepatocytes, with readable cell boundaries and little surrounding deposit. | This matches cell populations scored High by HPA tissue IHC. Compare staining within the relevant tissue architecture; HPA's Approved rating still carries medium RNA agreement and pending external verification (HPA tissue IHC). |
| Predominantly sharp membranous or extracellular staining, while expected cellular cytoplasm remains pale. | This differs from the reported cytoplasmic tissue IHC profile (HPA tissue IHC). Treat it as a possible staining artefact and review controls and morphology. A nuclear component alone needs separate assessment because HPA ICC-IF supports nucleoplasmic localization (HPA subcellular ICC-IF). |
| Strong staining in adipocytes, skeletal myocytes or soft-tissue fibroblasts, especially beside unstained expected-positive cells. | Those populations are listed as Not detected in HPA tissue IHC. Unexpected signal raises possible cross-reactivity or endogenous detection activity (general IHC practice); it does not prove either cause, and HPA negatives are reference observations, not universal exclusions. |
| A uniform brown haze covers cells and stroma, or tissue edges are darker than intact central regions. | Diffuse deposit obscures the cell-specific pattern needed for interpretation (general IHC practice). Review background and detection controls before assigning USP24 positivity; a broad haze cannot establish the cytoplasmic pattern reported by HPA tissue IHC. |
| No convincing cellular staining appears in colon glandular cells, liver hepatocytes or another HPA high-expression population. | An absent result in a reference-positive population warrants a run-level check before a biological negative call (HPA tissue IHC; general IHC practice). HPA reports high staining in these cells, but its medium-consistency rating limits certainty for any single specimen (HPA tissue IHC). |
💡Expected USP24 appearanceCall an IHC result positive when distinct cytoplasmic staining is visible in the relevant glandular cells, hepatocytes or trophoblastic cells at an intensity comparable with HPA High reference populations; diffuse extracellular deposit or isolated signal in HPA Not detected populations calls for control review (HPA tissue IHC; general IHC practice).
| Tissue and cell population | HPA tissue IHC scores appendix, colon, duodenum and gallbladder glandular cells, liver hepatocytes, pancreatic exocrine cells, nasopharyngeal respiratory epithelial cells and placental trophoblastic cells High. Its Not detected calls apply to specified cells, including adipocytes and skeletal myocytes, rather than every cell in those tissues (HPA tissue IHC). |
| Assay-specific compartment evidence | Tissue IHC describes predominantly cytoplasmic expression, whereas ICC-IF supports nucleoplasm as the main location and cytosol as an additional location (HPA tissue IHC; HPA subcellular ICC-IF). Interpret each pattern in its assay context; the ICC-IF finding does not by itself validate a nuclear chromogenic IHC result. |
| Antibody evidence | HPA lists HPA026723 and HPA028428 as Approved for IHC and ICC; HPA067124 is Approved for ICC, with no IHC status supplied (HPA antibodies). The overall tissue IHC profile is Approved but has medium staining–RNA consistency and awaits external verification (HPA tissue IHC). |
| Topology and processing | UniProt reports no transmembrane segment, signal peptide or propeptide, and annotates one chain spanning residues 1–2620 (UniProt Q9UPU5). These annotations give no basis to expect a membrane-restricted or secreted USP24 pattern; they do not establish fixation sensitivity or an antibody epitope. |
| IF/ICC Q&A: What pattern should I expect? | HPA ICC-IF supports mainly nucleoplasmic USP24 with additional cytosolic localization and lists images from A-431, U-251MG, U2OS and SK-MEL-30 (HPA subcellular ICC-IF). That is an assay-specific observation; this IHC section supplies no IF/ICC protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| A reference-positive colon or liver section is blank. | The expected cells stain High in HPA tissue IHC, so a blank run may reflect a workflow or detection problem; the payload does not identify a USP24-specific failure mechanism (HPA tissue IHC; general IHC practice). | Confirm that the reference cells are present, then check primary-antibody application, detection reagents and the run's positive control (general IHC practice). Repeat before scoring the specimen negative. |
| Cytoplasmic signal is faint in a HPA high-expression population. | Weak intensity may arise from the staining workflow or specimen variation; HPA's medium staining–RNA consistency does not guarantee strong staining in every section (HPA tissue IHC; general IHC practice). | Compare with a concurrently processed reference-positive section. Review the documented IHC dilution and retrieval conditions for the antibody being used; no target-specific settings are supplied here (general IHC practice). |
| Brown precipitate or diffuse haze makes cell boundaries unreadable. | Background from detection reagents or nonspecific staining can obscure a cellular chromogenic signal (general IHC practice). HPA's cytoplasmic profile cannot be confirmed from diffuse deposit (HPA tissue IHC). | Inspect a no-primary control and check blocking, washes and detection conditions. Score USP24 only where cellular morphology remains interpretable (general IHC practice). |
| Unexpected staining concentrates in skeletal myocytes, adipocytes or soft-tissue fibroblasts. | HPA lists these specified populations as Not detected; cross-reactivity or endogenous detection activity is possible, but the staining pattern alone cannot distinguish them (HPA tissue IHC; general IHC practice). | Compare the unexpected cells with adjacent morphology and a no-primary control; review endogenous enzyme blocking if an enzyme-based detector is used (general IHC practice). |
| Signal outlines cell membranes or appears outside cells instead of in cytoplasm. | This distribution conflicts with HPA's tissue IHC profile; UniProt reports no transmembrane segment, but neither source identifies the cause of the deposit (HPA tissue IHC; UniProt Q9UPU5 topology). | Check whether the pattern persists in a no-primary control and whether extracellular staining follows damaged tissue or precipitate. Reassess against reference-positive cellular staining (general IHC practice; HPA tissue IHC). |
| A nuclear IHC signal seems to conflict with the cytoplasmic IHC reference. | HPA reports cytoplasmic tissue IHC and supported nucleoplasmic ICC-IF localization. These assay observations differ, so nuclear IHC signal cannot be accepted or rejected from compartment alone (HPA tissue IHC; HPA subcellular ICC-IF). | Examine cytoplasmic staining in reference-positive cells and compare control sections. Record nuclear signal separately, with its assay and antibody, pending independent validation (HPA tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing USP24 staining
A08241 · IHCImmunohistochemistry analysis of paraffin-embedded human breast carcinoma tissue, using USP24 Antibody. The picture on the right is blocked
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000162402-USP24/tissue/cerebral+cortex#imid_7170328
HPA · StomachHuman Protein Atlas IHC, Stomach. Source: https://www.proteinatlas.org/ENSG00000162402-USP24/tissue/stomach+1#imid_7170228
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000162402-USP24/tissue/colon#imid_7170254
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000162402-USP24/tissue/liver#imid_7170255
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA026723; green = USP24, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA026723; green = USP24, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for USP24 IHC & IF
🧪Run colon first and expect staining in glandular cells (HPA: High in colon glandular cells). Use skeletal muscle myocytes as the biological negative (HPA: Not detected in skeletal muscle myocytes); on the colon slide, compare glandular staining with adjacent nonglandular tissue as a background reference, without assuming those cells lack USP24.
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show USP24 in A-431, U-251MG, U2OS, SK-MEL-30, with annotated localisation: Nucleoplasm (supported) (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 a knockout sample or peptide-block control; the selected product caption shows peptide blocking in a paraffin-section image (A08241 tissue-IHC caption). Quench endogenous peroxidase for chromogenic IHC and check any residual signal in the colon controls (standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported in the supplied evidence, and the selected SKU’s paraffin-section caption does not state a fixative (A08241 tissue-IHC caption). An antigen-retrieval requirement is unreported; optimise retrieval empirically for the IHC-validated antibody (supplied application evidence). HPA has ICC-IF images for USP24, but the evidence does not establish whether frozen sections or IF are easier than paraffin IHC (HPA: ICC-IF images); inspect colonic luminal material for nonspecific chromogen deposits (standard IHC practice).
HPA tissue IHC evidence for USP24
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Liver | Hepatocytes | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced USP24 IHC Tips
Troubleshoot USP24 staining in paraffin sections by checking retrieval, cell compartment, controls and scoring before interpreting chromogenic signal.
What retrieval should I try when USP24 staining is weak?
Use citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval specification). Let sections cool in buffer, then compare staining with a concurrently processed positive control, such as hepatocytes, which show high tissue staining (standard IHC practice; HPA: high in hepatocytes). If signal remains weak, test a small parallel section with EDTA at pH 9.0, treating that condition as an unvalidated fallback (standard IHC practice). Keep detection and imaging conditions matched so retrieval is the variable under test (standard IHC practice). Record whether stronger staining retains the expected cytoplasmic pattern rather than increasing diffuse background (HPA: cytoplasmic tissue expression; standard IHC practice).
How should I handle variable fixation across USP24 paraffin specimens?
Target-specific fixation sensitivity for USP24 is unknown, and the A08241 paraffin-section caption does not report a fixative (A08241 tissue-IHC caption). Record fixative, time before fixation and fixation duration for every specimen before comparing stain intensity (standard IHC practice). For prospective material, use a consistent routine fixation workflow and compare matched sections if fixation duration varies (standard IHC practice). Process a positive control in the same batch; glandular cells in colon show high USP24 tissue staining (standard IHC practice; HPA: high in colon glandular cells). Interpret differences between fixation groups cautiously until matched controls show that staining remains comparable under those conditions (standard IHC practice).
Should nuclear USP24 staining in tissue sections be considered genuine?
Expect predominantly cytoplasmic staining in tissue IHC, because that is the reported pattern across most cell types (HPA: tissue IHC profile). Supported ICC/IF observations place USP24 mainly in the nucleoplasm and additionally in the cytosol, so nuclear signal deserves investigation (HPA: subcellular profile). UniProt supplies no subcellular annotation and reports no transmembrane segment; neither fact establishes a tissue-section compartment (UniProt Q9UPU5 record). Score nuclear and cytoplasmic staining separately in morphologically intact cells, using the same detection settings across sections (standard IHC practice). Check nuclear signal against controls and the corresponding cytoplasmic pattern before calling it USP24-specific (standard IHC practice; HPA: tissue IHC profile).
Could epitope masking explain inconsistent USP24 staining?
The supplied record lists 0 annotated isoforms, a UBA domain at residues 3–44, and a USP domain at 1689–2042 (UniProt Q9UPU5 record). It also lists modified residues, including phosphoserines and phosphotyrosine, but does not locate this antibody’s epitope (UniProt Q9UPU5 record; supplied A08241 caption). Consequently, do not assign a staining difference to an isoform or modification without epitope information and direct validation (standard IHC interpretation). The A08241 tissue image includes a synthesized-peptide blocking comparison, which can support epitope-dependent staining in that specimen (A08241 tissue-IHC caption). Compare retrieval conditions and independent specificity controls before interpreting regional signal loss as USP24 biology (standard IHC practice).
How can IF help resolve ambiguous USP24 staining in tissue?
Use IF as a separate validation experiment when chromogenic IHC cannot resolve cellular or nuclear boundaries (standard IF practice). Pair USP24 with a marker of the expected cell type, such as a glandular epithelial marker when examining high-staining colon glands (HPA: high in colon glandular cells; standard IF practice). Choose a far-red fluorophore where shorter-wavelength tissue autofluorescence obscures signal, and include unstained and single-label controls (standard IF practice). For intracellular access, trial gentle permeabilisation, for example 0.1% Triton X-100 for 5 min; USP24 has no transmembrane segment (standard IF practice; UniProt Q9UPU5 topology). Assess nucleoplasmic and cytosolic signal separately, since both have supported ICC/IF localisation evidence (HPA: subcellular profile).
What should I check when USP24 DAB staining is diffuse?
Run a no-primary control to reveal signal from the detection system, and inspect whether diffuse stain tracks tissue edges or damaged areas (standard IHC practice). Block endogenous peroxidase before DAB development; 3% hydrogen peroxide for 10 min is a starting condition to validate locally (standard IHC practice). Optimise protein blocking, primary-antibody concentration and wash stringency on matched sections while keeping development time constant (standard IHC practice). Compare with a high-staining control, such as liver hepatocytes, and a reported undetected cell type, such as adipocytes (HPA: high in hepatocytes; HPA: not detected in adipocytes). Retain only discrete cellular staining that survives these controls for interpretation (standard IHC practice).
How should I quantify heterogeneous USP24 staining in paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Predefine the cell population and score viable cells separately from necrosis, section edges and non-target tissue (standard IHC practice). An H-score multiplies the percentage of cells at each intensity by its 0–3 intensity grade, yielding 0–300 (standard IHC practice). Report percent positive cells alongside H-score, and keep nuclear and cytoplasmic scores separate if both appear (standard IHC practice; HPA: tissue and subcellular profiles). Normalise counts to the number of eligible cells, or positive-cell density to evaluated viable area in mm² (standard IHC practice). Apply identical thresholds, counterstain settings and compartment definitions across specimens, recording batch and control performance (standard IHC practice).
How do I distinguish true USP24 signal from tissue artefact?
Prioritise reproducible staining within intact cells and assess whether its compartment fits the mainly cytoplasmic tissue profile (HPA: tissue IHC profile; standard IHC practice). Use high-staining hepatocytes or colon glandular cells as contextual positives, while recognising that adipocytes are reported as undetected (HPA: tissue IHC profile). Treat edge enhancement, necrotic debris and staining present in no-primary controls as potential artefacts, including endogenous enzyme activity (standard IHC practice). The A08241 image includes peptide blocking in paraffin-embedded breast carcinoma, but its caption does not state the fixative (A08241 tissue-IHC caption). Qualify unexpected nuclear or cell-type patterns because the HPA tissue assessment has medium RNA concordance and awaits external verification (HPA: Approved reliability description).
Boster reagents
Best USP24 / Ubiquitin carboxyl-terminal hydrolase 24 IHC Antibodies
A08241 has IHC image evidence in paraffin-embedded human breast carcinoma (IHC image caption); IF/ICC applications and human and mouse reactivity are catalog-listed (catalog: applications and reactivity).
Real IHC data
Anti-USP24 Antibody
Cat # A08241
A08241 will render with an IHC image of paraffin-embedded human breast carcinoma and a peptide-blocked comparison (IHC image caption). IF and ICC are listed applications, but the payload provides no IF image (catalog: applications and IF image alts).
Which to pick: Choose A08241 for paraffin-section tissue IHC: its own image caption shows human breast carcinoma, while the fixative is unreported (IHC image caption). The same rabbit polyclonal antibody is listed for IF/ICC and for human and mouse reactivity, but its image evidence is limited to human tissue IHC (catalog: host, clone, applications and reactivity; IHC image caption; IF image alts).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9UPU5 (UBP24_HUMAN, Ubiquitin carboxyl-terminal hydrolase 24).
- Human Protein Atlas. USP24 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. USP24 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol..
- Human Protein Atlas. USP24 antibody validation summary (3 antibodies).
- Ubiquitin-specific peptidase 24 accelerates aerobic glycolysis and tumor progression in gastric carcinoma through stabilizing PLK1 to activate NOTCH1. Cancer science 2023 — PMC10394155.
- USP24-dependent STAT2 stabilization mediates physiologic and pathologic bone formation. Cell death & disease 2026 — PMC13433775.
- USP24 promotes autophagy-dependent ferroptosis in hepatocellular carcinoma by reducing the K48-linked ubiquitination of Beclin1. Communications biology 2024 — PMC11461744.
- USP24-GSDMB complex promotes bladder cancer proliferation via activation of the STAT3 pathway. International journal of biological sciences 2021 — PMC8315027.
- PubMed PMID:16710414 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:10470851 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for USP24 (Ubiquitin carboxyl-terminal hydrolase 24). Expected localisation Cytoplasmic staining in most cell types (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A08241 at 1:100 - 1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast carcinoma tissue; fixative not specified (datasheet A08241); 2. Fixation: Image fixative and duration unreported (datasheet A08241); 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-USP24, 1:100 - 1:300 (datasheet A08241); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep formalin fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Adipocytes can be unstained despite the broad profile (HPA tissue IHC); Isoform / epitope: No annotated isoforms, signal peptide or processed chain (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt Q9UPU5, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A08241, PMC10394155, PMC13433775. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/USP24/immunohistochemistry-ihc-guide.