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TSG101 / Tumor susceptibility gene 101 protein · IHC design guide
Design Immunohistochemistry for TSG101
Plan TSG101 paraffin-section IHC around the cytoplasmic and membranous tissue pattern reported by HPA (HPA tissue IHC). Use high-staining glandular tissue as a reference and account for unstained adipocytes when interpreting results (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TSG101 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 and membranous tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic and membranous staining in glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Appendix+4 more · see all | |
| Negative control | Adipose tissue |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes may be unstained despite broad tissue expression (HPA tissue IHC) | |
| Regulation | No staining-intensity regulator specified (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended TSG101 IHC & IF Protocols
The catalog antibody protocol is accompanied by published TSG101 IHC methods for cervical and renal samples (PMC3997686; PMC6332891).
| Sample | Paraffin-embedded Rat liver tissue; fixative not specified (datasheet M01233) |
| Fixation | Image fixative and duration unreported (datasheet M01233); 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 IDG-20) anti-TSG101, 1:50 recommended; image 1:500 (datasheet M01233) |
| 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 | TSG101-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic and membranous expression. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min (page antigen-retrieval rule); neither excerpt specifies a retrieval method.
Section 2
What Is the Expected TSG101 Staining Pattern?
In paraffin IHC, expect mainly cytoplasmic staining, with possible membranous staining (HPA: general cytoplasmic and membranous expression; UniProt Q99816: mainly cytoplasmic, membrane associated when active). Glandular, respiratory epithelial, hematopoietic and neuronal cells provide documented high staining examples (HPA: tissue IHC). TSG101 has no transmembrane segment (UniProt Q99816 topology). HPA rates its tissue IHC pattern Approved, with medium consistency between staining and RNA data (HPA: reliability).
| Cytoplasmic chromogen in appendix, breast, cervix, colon or duodenal glandular cells. | This fits the reported compartment and high staining in those cells (HPA: general cytoplasmic expression; High in the listed glandular cells). Assess signal in the named cells rather than calling an entire tissue uniformly positive; HPA assigns staining levels to cell populations (HPA: tissue IHC). |
| Cytoplasmic staining, sometimes with a membranous component, in bronchial epithelium, marrow cells or cortical neurons. | Respiratory epithelial, hematopoietic and neuronal cells are each reported High in the corresponding tissue (HPA: bronchus, bone marrow and cerebral cortex). A membranous component is compatible with the tissue profile and membrane association during activity (HPA: tissue IHC; UniProt Q99816: subcellular location). |
| Predominantly nuclear staining, or a sharply defined surface rim with little cytoplasm, across many cells. | Recheck compartment assignment and staining controls before accepting this as the tissue IHC pattern: HPA describes general cytoplasmic and membranous staining, while UniProt calls TSG101 mainly cytoplasmic (HPA: tissue IHC; UniProt Q99816). Nuclear localization can depend on the cell cycle, so an isolated nuclear signal is not decisive by itself (UniProt Q99816). |
| Strong adipocyte staining while documented high staining populations remain weak or absent. | Treat this as a specificity concern, including possible cross reactivity or endogenous detection activity, rather than proof of a new positive cell type (HPA: adipocytes Not detected; High in listed positive cells). Check the detection only control and cell identity; one discordant section cannot establish the cause (general IHC practice). |
| Diffuse chromogen over tissue, stroma and cell free areas, or no staining in a documented high staining population. | Diffuse deposit lacks the cell and compartment pattern needed for interpretation; absent signal in a documented positive population makes a negative call unreliable (HPA: tissue IHC). Review background controls, section quality and assay steps before attributing either result to TSG101 distribution (general IHC practice). |
💡Expected TSG101 appearanceCall a section positive when identifiable cells show mainly cytoplasmic, sometimes membranous, chromogen, with high staining in a documented high staining population; widespread cell independent deposit or isolated staining of HPA negative adipocytes is suspect (HPA: tissue IHC; UniProt Q99816: mainly cytoplasmic).
| Cell population and tissue context | HPA reports High staining in specified glandular, respiratory epithelial, hematopoietic and neuronal cells, but Not detected in adipocytes (HPA: tissue IHC). Select and score the relevant cells; low tissue RNA specificity does not make every cell an IHC positive control (HPA: RNA specificity and tissue IHC). |
| Compartment and membrane association | TSG101 is mainly cytoplasmic and can associate with early and late endosome membranes; it has no transmembrane segment (UniProt Q99816: subcellular location and topology). HPA's general membranous tissue profile supports a membrane component, without specifying a required continuous cell surface rim (HPA: tissue IHC). |
| Cell cycle context | Nuclear localization is cell cycle dependent, and CEP55 interaction is required for midbody localization during cytokinesis (UniProt Q99816: subcellular location). These contexts can explain occasional noncytoplasmic signal, but do not replace the general tissue IHC expectation of cytoplasmic and membranous staining (HPA: tissue IHC). |
| Strength of tissue IHC evidence | The tissue profile is Approved with medium consistency between antibody staining and RNA expression data; HPA006161 and CAB004283 each have Approved IHC status (HPA: tissue IHC reliability; HPA: antibodies). Use the reported pattern as a reference while resolving unexpected staining with controls, rather than treating every departure as biological (general IHC practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a documented high staining cell population. | The assay may have failed, or the sampled section may lack enough identifiable target cells; HPA reports High staining only for the specified cell populations (HPA: tissue IHC). | Verify that the expected cells are present, then review primary antibody, retrieval, detection and counterstain steps against the validated IHC workflow (general IHC practice). Do not call the specimen negative until a positive control stains. |
| Broad haze obscures cytoplasm and cell borders. | Nonspecific antibody binding, excessive detection deposit or incomplete washing can obscure localization (general IHC practice). This appearance cannot be matched confidently to HPA's cytoplasmic and membranous profile (HPA: tissue IHC). | Inspect a detection only control, blocking and wash steps; adjust the validated staining workflow as needed, then rescore identifiable cells (general IHC practice). |
| Adipocytes stain strongly, especially when expected positive cells do not. | Cross reactivity or endogenous detection activity is possible; adipocytes are reported Not detected (HPA: adipose tissue IHC). The discordance alone does not identify which mechanism is responsible. | Confirm cell identity and compare detection only and positive control sections before interpreting adipocyte signal (general IHC practice; HPA: tissue IHC). |
| Signal appears mainly nuclear or as a uniform cell surface outline. | This differs from the mainly cytoplasmic expectation, although cell cycle dependent nuclear localization and membrane association are reported (UniProt Q99816: subcellular location; HPA: tissue IHC). | Check whether cytoplasmic signal is present in documented high staining cells, and review compartment boundaries and background controls before assigning a novel localization (HPA: tissue IHC; general IHC practice). |
| A slide stains, but intensity varies between sampled cell types. | HPA's High and Not detected calls are cell specific, and its tissue IHC profile has medium consistency with RNA data (HPA: tissue IHC reliability and cell levels). | Score each identifiable cell population separately and compare it with its own HPA reference; avoid interpreting whole section intensity as a single TSG101 result (HPA: tissue IHC; general IHC practice). |
| Q: IF/ICC shows nucleolar or plasma membrane signal. Does that invalidate the IHC slide? | A: No. HPA reports cytosol as the supported main IF location and nucleoli and plasma membrane as approved additional locations (HPA: subcellular ICC-IF). | Judge the paraffin IHC slide against its own cytoplasmic and membranous tissue profile and controls (HPA: tissue IHC; general IHC practice). Use the separate IF/ICC guide for imaging decisions. |
Experiment evidence
IHC & IF images showing TSG101 staining
M01233 · IHCImmunohistochemical analysis of paraffin-embedded Rat liver, using the Antibody at 1:500 dilution.
M01233 · IHCImmunohistochemical analysis of paraffin-embedded Human prostate cancer, using the Antibody at 1:1000 dilution.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000074319-TSG101/tissue/cerebral+cortex#imid_1411076
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000074319-TSG101/tissue/colon#imid_1411010
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000074319-TSG101/tissue/liver#imid_2185727
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000074319-TSG101/tissue/kidney#imid_1410996
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006161; green = TSG101, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006161; green = TSG101, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TSG101 IHC & IF
🧪Run breast first: its glandular cells should stain (HPA: High in breast glandular cells). Run adipose tissue as the negative comparator (HPA: Not detected in adipocytes); adipocytes within the breast section should provide a local background check, although their staining in that section is not established by the supplied HPA row.
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 TSG101 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 catalog antibody’s host and class (same subclass for a monoclonal), and TSG101 knockout material as a biological negative. For chromogenic IHC, block endogenous peroxidase and check the breast section for residual background signal (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and retrieval dependency are unreported in the supplied evidence; optimize antigen retrieval empirically for paraffin sections. The selected M01233 rat-liver paraffin-section caption reports 1:500 dilution but leaves the fixative unreported (selected-SKU caption). IF images exist in A-431, U-251MG and U2OS, with cytosol as the supported main location (HPA: subcellular); the supplied evidence does not establish whether frozen sections or IF are easier, or identify a breast-specific artefact.
HPA tissue IHC evidence for TSG101
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TSG101 IHC Tips
Troubleshoot TSG101 staining in paraffin section chromogenic IHC by checking retrieval, compartment, controls and scoring before interpreting differences between samples.
How should I retrieve TSG101 in paraffin sections when staining is weak?
Use Tris-EDTA pH 9.0 heat-induced epitope retrieval at 95–98 °C for 20 min (page retrieval setting). Allow sections to cool in retrieval buffer, then wash consistently before blocking so that comparisons reflect staining rather than handling differences (standard IHC practice). If signal remains weak, test a citrate buffer at pH 6.0 on adjacent sections as a fallback, with the same antibody and detection conditions (standard IHC practice). Include a positive reference section in each run because TSG101 is generally cytoplasmic and membranous in tissue IHC (HPA: tissue IHC profile). Compare signal and tissue preservation together; stronger staining after harsher retrieval does not alone establish specificity (standard IHC practice).
Could fixation explain inconsistent TSG101 staining between paraffin blocks?
Target-specific fixation sensitivity for TSG101 is unknown from the supplied evidence; the catalog antibody’s rat liver paraffin caption does not state a fixative (catalog antibody M01233: tissue-IHC caption). Record each block’s fixative, fixation duration and processing history before comparing staining intensity across cases (standard IHC practice). For a new tissue series, keep fixation and section thickness consistent, and process a reference block alongside study sections (standard IHC practice). If blocks differ in fixation history, compare retrieval conditions on adjacent sections while holding antibody incubation and chromogenic detection constant (standard IHC practice). Do not assign a fixation effect to TSG101 from its localisation, topology or modifications alone (UniProt Q99816: localisation, topology and modified residues).
Where should convincing TSG101 staining appear in tissue sections?
Expect predominantly cytoplasmic staining, with possible membranous staining, when assessing chromogenic tissue sections (HPA: general cytoplasmic and membranous expression; UniProt Q99816: mainly cytoplasmic). TSG101 associates with early and late endosome membranes during its activity, so granular cytoplasmic signal can be plausible (UniProt Q99816: subcellular localisation). Nuclear localisation can vary with the cell cycle, and midbody localisation depends on CEP55 during cytokinesis (UniProt Q99816: subcellular localisation). Assess these less common patterns against cell morphology and a matched reference section before reporting them as dominant tissue patterns (standard IHC practice). Diffuse staining confined to damaged edges or necrotic regions should prompt a technical review before compartment scoring (standard IHC practice).
Can isoforms or epitope accessibility explain different TSG101 patterns?
TSG101 has 2 listed isoforms, while its documented UEV and SB domains span residues 2–145 and 322–390, respectively (UniProt Q99816: isoforms and domains). The supplied catalog caption does not identify the antibody epitope, so these data cannot establish which isoforms its tissue staining detects (catalog antibody M01233: tissue-IHC caption; UniProt Q99816: isoforms). If an epitope map becomes available, check whether its sequence is retained in each isoform before comparing staining across specimens (standard antibody validation practice). Also consider retrieval-dependent epitope accessibility when adjacent sections show different intensities under otherwise matched conditions (standard IHC practice). Do not attribute a nuclear or granular pattern to a particular isoform without independent isoform-specific evidence (UniProt Q99816: isoforms and localisation).
How can I check a TSG101 pattern by multiplex IF?
Use IF as a separate localisation check, since the catalog evidence supplied here documents paraffin section IHC rather than IF performance (catalog antibody M01233: tissue-IHC caption). Pair TSG101 with a marker identifying the cell population being scored, such as a glandular cell marker when evaluating glandular staining (HPA: high staining in glandular cells of appendix, breast, cervix, colon and duodenum; standard IF practice). Choose spectrally separated fluorophores and place the weaker signal in a channel with lower tissue autofluorescence after checking unstained sections (standard IF practice). Use a mild permeabilisation condition that exposes the cytosolic side of endosome membranes, because TSG101 lacks a transmembrane segment and is mainly cytoplasmic (UniProt Q99816: topology and localisation). Confirm the IF fixation and permeabilisation conditions experimentally; the paraffin IHC caption provides no IF fixation evidence (catalog antibody M01233: tissue-IHC caption).
What should I check when TSG101 DAB staining looks widespread?
Compare stained sections with a no-primary control and inspect whether colour follows cells or accumulates over tissue edges and damaged areas (standard chromogenic IHC practice). Block endogenous peroxidase before horseradish peroxidase detection, and include an appropriate protein block to reduce nonspecific binding (standard chromogenic IHC practice). Titrate the catalog antibody around its reported 1:500 rat liver paraffin-image dilution while keeping development time and retrieval fixed (catalog antibody M01233: tissue-IHC caption; standard IHC practice). Widespread cytoplasmic signal is not automatically background because tissue IHC reports general cytoplasmic and membranous expression (HPA: tissue IHC profile). Judge suspicious signal against the no-primary control and expected cell boundaries before reducing antibody concentration further (standard IHC practice).
How should I quantify TSG101 staining across tissue samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment to be scored before reviewing cases, using cytoplasmic staining as the main tissue readout (HPA: tissue IHC profile; standard IHC practice). For a semiquantitative result, report the percentage of positive cells and an H-score from 0–300, calculated from percentages at intensity grades 0–3 (standard IHC scoring practice). Apply one threshold, imaging setup and region selection rule across the series, and exclude necrosis and section edges from analysis (standard IHC practice). Normalise positive counts to the number of eligible cells, or positive-cell density to the evaluated tissue area in mm² (standard image analysis practice). Report nuclear or membranous scores separately when those compartments are a study endpoint (UniProt Q99816: localisation; standard IHC scoring practice).
How do I distinguish true TSG101 staining from an artefact?
Look first for cell-associated, predominantly cytoplasmic staining, allowing a membranous component rather than requiring an exclusively punctate pattern (HPA: tissue IHC profile; UniProt Q99816: localisation). High staining in appendix glandular cells or bone marrow hematopoietic cells can provide context, while adipocytes are reported as not detected (HPA: tissue IHC cell-level observations). A nuclear signal needs careful compartment assessment because TSG101 nuclear localisation is cell cycle-dependent, not a uniform tissue expectation (UniProt Q99816: subcellular localisation). Reject apparent positives concentrated at cut edges, necrosis or the no-primary control, and investigate residual endogenous peroxidase before interpreting DAB colour (standard chromogenic IHC practice). HPA rates its tissue staining as Approved with medium consistency against RNA expression, so use controls and morphology to support case-level conclusions (HPA: tissue IHC reliability).
Boster reagents
Best TSG101 / Tumor susceptibility gene 101 protein IHC Antibodies
M01233 has paraffin-section IHC images from rat liver, human prostate cancer, mouse kidney, and rat stomach (catalog: IHC image captions). Both antibodies list IF and human, mouse, and rat reactivity (catalog: applications/reactivity).
Real IHC data
Anti-TSG101 Rabbit Monoclonal Antibody
Cat # M01233
M01233 lists IHC and ICC/IF for human, mouse, and rat, with paraffin-section IHC images from rat liver, human prostate cancer, mouse kidney, and rat stomach (catalog: applications/reactivity/IHC image captions). M01233-2 lists IF for human, mouse, and rat; no IHC or IF image caption is supplied for it (catalog: applications/reactivity/image captions).
Which to pick: Choose M01233 for tissue IHC: its own captions document paraffin-section staining in the named human, mouse, and rat samples (catalog: IHC image captions); the fixative is unreported (catalog: IHC image captions). For IF/ICC, M01233 lists both applications and is a rabbit monoclonal clone IDG-20 (catalog: applications/host/clone), while M01233-2 is described as a recombinant rabbit monoclonal and lists IF only (catalog: short description/applications). Both list human, mouse, and rat reactivity for cross-species work (catalog: reactivity); M01233 alone supplies IHC images across those species (catalog: IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q99816 (TS101_HUMAN, Tumor susceptibility gene 101 protein).
- Human Protein Atlas. TSG101 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. TSG101 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoli and plasma membrane..
- Human Protein Atlas. TSG101 antibody validation summary (2 antibodies).
- Expression of TSG101 protein and LSF transcription factor in HPV-positive cervical cancer cells. Oncology letters 2014 — PMC3997686.
- Endothelial cell-derived exosomes protect SH-SY5Y nerve cells against ischemia/reperfusion injury. International journal of molecular medicine 2017 — PMC5593464.
- siRNA against TSG101 reduces proliferation and induces G0/G1 arrest in renal cell carcinoma - involvement of c-myc, cyclin E1, and CDK2. Cellular & molecular biology letters 2019 — PMC6332891.
- Analysis of the Expression of LSF Transcription Factor in the Regulation of Transcription and TSG101 during the Neoplastic Transformation of Endometrial Cells. Cells 2024 — PMC11011417.
- PubMed PMID:9019400 — UniProt-cited evidence.
- PubMed PMID:9867424 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TSG101 (Tumor susceptibility gene 101 protein). Expected localisation Cytoplasmic and membranous tissue staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: M01233 at 1:50 recommended; image 1:500 (the product's own IHC validation image is shown). Controls (HPA): positive Appendix (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded Rat liver tissue; fixative not specified (datasheet M01233); 2. Fixation: Image fixative and duration unreported (datasheet M01233); 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 IDG-20) anti-TSG101, 1:50 recommended; image 1:500 (datasheet M01233); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Adipocytes may be unstained despite broad tissue expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unspecified (UniProt); Regulation: No staining-intensity regulator specified (UniProt). Evidence: UniProt Q99816, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the…, Datasheet M01233, PMC3997686, PMC6332891. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TSG101/immunohistochemistry-ihc-guide.