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TSC1 / Hamartin · IHC design guide
Design Immunohistochemistry for TSC1
Plan chromogenic TSC1 IHC in paraffin sections around widespread cytoplasmic staining (HPA tissue IHC) and a primary antibody starting range of 2–5 µg/mL (datasheet A00365-2). Compare matched sections and control background when scoring (standard IHC practice).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TSC1 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 tissue sections (HPA tissue IHC) | |
| Staining pattern | Widespread cytoplasmic staining across cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A00365-2) | |
| Positive control | Bone marrow+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A00365-2) | |
| Caveat | Endogenous peroxidase may confound bone marrow staining (HPA tissue IHC; standard IHC practice) | |
| Regulation | Nutrients redistribute TSC1 within cells (UniProt) | |
| Isoform / epitope | Two isoforms; antibody epitope coverage is unspecified (UniProt) |
Section 1
Recommended TSC1 IHC & IF Protocols
The catalog antibody protocol uses EDTA retrieval (datasheet A00365-2). The published IHC protocols below report conditions for renal carcinoma, paraffin tissue, and bladder tumour samples (PMC7156823; PMC11607531; PMC2427143).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A00365-2) |
| Fixation | Image fixative and duration unreported (datasheet A00365-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A00365-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A00365-2) |
| Primary antibody | Rabbit anti-TSC1, 2-5 μg/ml (datasheet A00365-2) |
| Primary incubation | Overnight at 4 °C (datasheet A00365-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A00365-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | TSC1-positive staining in hematopoietic cells of bone marrow (HPA tissue IHC: High). HPA tissue profile: Ubiquitous cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A00365-2). Use citrate retrieval when reproducing the corresponding published protocols (PMC7156823; PMC2427143).
Section 2
What Is the Expected TSC1 Staining Pattern?
TSC1 should show predominantly cytoplasmic staining across many cell types in paraffin sections (HPA tissue IHC: ubiquitous cytoplasmic expression; reliability Supported, with medium consistency between staining and RNA). Strong examples include skeletal myocytes, cardiomyocytes and placental cytotrophoblasts (HPA tissue IHC: High). A cytosolic pattern is consistent with its lack of a transmembrane segment (UniProt Q92574: cytosol; no transmembrane segment).
| Cytoplasmic staining in skeletal myocytes, cardiomyocytes or placental cytotrophoblasts. | This matches reported high staining in those cells (HPA tissue IHC: High). Compare signal within the relevant cell type; the tissue-wide average may obscure which cells are positive (standard IHC interpretation). |
| Predominantly nuclear or sharply cell-surface staining with little cytoplasmic signal. | This conflicts with the reported cytoplasmic pattern (HPA tissue IHC: ubiquitous cytoplasmic expression) and cytosolic location (UniProt Q92574). Review staining specificity and morphology before scoring it as TSC1 (standard IHC practice). |
| Strong staining confined to adipocytes or soft-tissue fibroblasts while expected positive cells are faint. | HPA reports low staining in those cell types and high staining in several others (HPA tissue IHC: Low in adipocytes and soft-tissue fibroblasts; High in skeletal myocytes). Consider cross-reactivity or endogenous detection activity, then compare controls (standard IHC practice). |
| Broad haze over cells and empty areas, with poorly defined cytoplasm. | A pattern that does not track cell boundaries is difficult to interpret against the reported cytoplasmic distribution (HPA tissue IHC). Check reagent background and detection controls before assigning a TSC1 score (standard IHC practice). |
| No detectable signal in a well-preserved, known-positive cell population. | Absence in cells reported as High, such as cardiomyocytes, makes the run inconclusive without controls (HPA tissue IHC: High in cardiomyocytes). Review the positive control and detection steps; HPA's Supported reliability does not guarantee every section will stain (HPA tissue IHC; standard IHC practice). |
💡Expected TSC1 appearanceA convincing positive is cell-associated, predominantly cytoplasmic staining, potentially strong in HPA High cell types such as skeletal myocytes; isolated nuclear or cell-free color is suspect (HPA tissue IHC: ubiquitous cytoplasmic expression; High in skeletal myocytes; standard IHC interpretation).
| Cell type and tissue context (HPA tissue IHC) | HPA describes ubiquitous cytoplasmic expression but different levels among cells: High in bone-marrow hematopoietic cells and lung alveolar type I cells, Medium in adrenal glandular cells, and Low in adipocytes (HPA tissue IHC). Interpret intensity by cell population, not tissue name alone (standard IHC practice). |
| Antibody validation (HPA antibody records) | Two listed antibodies, CAB011568 and CAB012481, have Supported IHC status; HPA074132 has no listed IHC status (HPA antibodies). The tissue profile has Supported reliability with medium staining–RNA consistency (HPA tissue IHC). These qualifications limit certainty for an unexpected pattern; they do not validate an unspecified antibody. |
| Cytosol and nutrient-dependent location (UniProt Q92574) | TSC1 is listed in the cytosol and at lysosomal membranes; recruitment to lysosomal membranes occurs without nutrients, with relocalization to cytosol after nutrients return (UniProt Q92574). Routine tissue IHC should therefore be read primarily against the cytoplasmic pattern observed by HPA (HPA tissue IHC), without assuming a visible lysosomal rim. |
| Isoforms and antigen identity (UniProt Q92574) | UniProt lists two TSC1 isoforms and one annotated Hamartin chain spanning residues 1–1164, with no signal peptide or propeptide (UniProt Q92574). No antibody epitope or isoform coverage is supplied, so these entries cannot predict whether a particular IHC reagent detects both isoforms. |
| Detection chemistry (standard chromogenic IHC practice) | Endogenous enzyme activity or nonspecific detection reagents can produce color unrelated to antibody binding (standard chromogenic IHC practice). A suitable negative reagent control helps identify that contribution; it does not establish the identity of otherwise plausible cytoplasmic staining (standard IHC interpretation). |
| Situation | Likely cause | Next action |
|---|---|---|
| High-background section with color outside recognizable cells. | Reagent deposition, inadequate blocking or endogenous detection activity may contribute (standard chromogenic IHC practice). | Inspect a matched negative reagent control and the distribution of color; adjust blocking, washing or detection conditions as indicated by that control (standard IHC practice). |
| No cytoplasmic signal in skeletal myocytes or cardiomyocytes. | These are High cell types in HPA, so an absent signal raises concern about the staining run, antibody performance or specimen quality (HPA tissue IHC; standard IHC interpretation). | Check a same-run positive control, primary antibody use, retrieval conditions and detection reagents before calling the specimen negative (standard IHC practice). |
| Nuclear staining dominates the section. | The compartment conflicts with HPA's ubiquitous cytoplasmic pattern and UniProt's cytosolic annotation (HPA tissue IHC; UniProt Q92574). | Compare the negative reagent control and an HPA High cell population; reassess antibody specificity if the nuclear pattern persists (HPA tissue IHC; standard IHC practice). |
| Strong color appears mainly in adipocytes or soft-tissue fibroblasts. | HPA assigns both cell populations Low staining, so disproportionate signal may reflect nonspecific binding or endogenous activity (HPA tissue IHC; standard IHC practice). | Confirm cell identity and compare negative controls with a reported High cell population in the same run; avoid treating Low as an absolute negative (HPA tissue IHC; standard IHC practice). |
| Patchy cytoplasmic staining makes scoring uncertain. | The HPA profile is ubiquitous but cell levels vary, and its reliability is Supported with medium staining–RNA consistency (HPA tissue IHC). Section quality can also complicate scoring (standard IHC practice). | Score identifiable cell populations separately and compare an appropriate positive control; record patchiness rather than converting it to a tissue-wide absence (standard IHC interpretation). |
| IF/ICC question: should a matching cell image show only diffuse cytosol? | HPA supports cytosol as the main location; its additional lipid-droplet, cilium-transition-zone and basal-body locations are uncertain (HPA subcellular ICC-IF). | Use cytosol as the primary comparison and treat puncta or those additional locations cautiously; assess them with appropriate imaging controls (HPA subcellular ICC-IF; standard IF practice). |
Experiment evidence
IHC & IF images showing TSC1 staining
A00365-2 · IHCIHC analysis of Hamartin/TSC1 using anti-Hamartin/TSC1 antibody (A00365-2). Hamartin/TSC1 was detected in a paraffin-embedded section of hum
A00365-2 · IHCIHC analysis of Hamartin/TSC1 using anti-Hamartin/TSC1 antibody (A00365-2). Hamartin/TSC1 was detected in a paraffin-embedded section of hum
A00365-2 · IFIF analysis of Hamartin/TSC1 using anti-Hamartin/TSC1 antibody (A00365-2). Hamartin/TSC1 was detected in an immunocytochemical section of A5
PA2012 · IHCAnti-Hamartin antibody, PA2012, IHC(P) IHC(P): Human Mammary Cancer Tissue
PA2012 · IHCAnti-Hamartin antibody, PA2012, IHC(P) IHC(P): Rat Brain Tissue
A00365 · IFImmunocytochemistry of TSC1 in EL4 cells with TSC1 antibody at 2 μg/mL
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000165699-TSC1/tissue/cerebral+cortex#imid_3800976
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000165699-TSC1/tissue/colon#imid_3800910
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000165699-TSC1/tissue/liver#imid_3800919
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000165699-TSC1/tissue/kidney#imid_3800912
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB011568; green = TSC1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB011568; green = TSC1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TSC1 IHC & IF
🧪Run heart muscle first: cardiomyocytes should show TSC1 staining (HPA: High in cardiomyocytes). HPA detects TSC1 in all 45 scored tissues, so there is no supported negative tissue; use no-primary and isotype controls, and treat any unstained cells within the heart section as background-level observations rather than validated TSC1-negative cells (HPA: no negative tissue rows).
Positive control tissue: Bone marrow (Hematopoietic cells, HPA High)
Negative control tissue: None in HPA: TSC1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TSC1 in A-431, U-251MG, U2OS, HaCaT, Rh30, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide; a concentration-matched rabbit IgG isotype control matched to the primary antibody’s clonality; and, if available, a matched TSC1-knockout specimen or validated peptide-block control (selected-SKU IHC caption: rabbit primary). Quench endogenous peroxidase in blood-containing heart sections before interpreting DAB staining (standard IHC practice).
⚠️Feasibility: The selected-SKU paraffin-section caption uses heat retrieval in EDTA at pH 8.0 and DAB detection, but does not establish that retrieval is required (selected-SKU IHC caption). A target-specific fixation window or fixation effect is unreported, and the caption does not state a fixative (selected-SKU IHC caption). Whether frozen sections or IF are easier is unreported; blood-associated endogenous peroxidase can add DAB background in heart sections (standard IHC practice).
HPA tissue IHC evidence for TSC1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Cerebellum | Cells in molecular layer | High | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | High | Protein (IHC) | HPA → |
| Lung | Alveolar cells type I | High | Protein (IHC) | HPA → |
| Placenta | Cytotrophoblasts | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: TSC1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced TSC1 IHC Tips
Troubleshoot TSC1 chromogenic IHC in paraffin sections by checking retrieval, cell identity and cytoplasmic staining before interpreting signal intensity.
Which retrieval conditions should I try first for weak TSC1 staining?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A00365-2). The selected tissue image used this treatment before 2 μg/ml primary antibody overnight at 4°C; reproduce those conditions and include a section without primary antibody (datasheet A00365-2; standard IHC practice). If staining remains weak, compare another retrieval buffer or heating duration on adjacent sections while holding detection conditions constant, and inspect tissue morphology (standard IHC practice). Favor conditions that preserve interpretable cytoplasmic staining over those that produce diffuse color or tissue damage, because TSC1 is mainly cytosolic (HPA subcellular; standard IHC practice).
Could fixation explain variable TSC1 staining between paraffin sections?
The selected paraffin-section caption does not state the fixative, so target-specific fixation sensitivity is unknown (datasheet A00365-2). Record the fixative and processing history for each specimen, and compare sections processed together before attributing a staining difference to TSC1 abundance (standard IHC practice). Keep the EDTA pH 8.0 retrieval and 2 μg/ml primary concentration constant while checking whether a processing batch also differs in tissue preservation or background (datasheet A00365-2; standard IHC practice). If processing histories differ, repeat staining on matched sections when available and report the limitation rather than assigning a target-specific fixation effect (standard IHC practice).
Where should convincing TSC1 staining appear in a tissue section?
Look first for cytoplasmic staining: tissue IHC shows ubiquitous cytoplasmic expression, and subcellular data support a mainly cytosolic location (HPA tissue IHC; HPA subcellular). TSC1 can associate with lysosomal membranes when nutrients are absent and return to the cytosol in response to nutrients, so some redistribution is biologically plausible (UniProt Q92574 localisation, PubMed:24529379). It has no annotated transmembrane segment, and a DAB section alone cannot establish which side of a lysosomal membrane carries the signal (UniProt Q92574 topology; standard IHC practice). Treat predominantly nuclear staining as suspect and check cell morphology and controls before assigning it to TSC1 (HPA subcellular; standard IHC practice).
Can this antibody distinguish TSC1 isoforms or phosphorylation states in IHC?
TSC1 has 2 annotated isoforms, but the supplied antibody caption does not identify its epitope or establish isoform selectivity (UniProt Q92574 isoforms; datasheet A00365-2). The protein also has annotated phosphoserines, including residues 487, 505 and 511; those annotations do not establish phospho-specific staining by this antibody (UniProt Q92574 modified residues; datasheet A00365-2). Check the antibody's documented immunogen or epitope against both isoforms before interpreting a staining difference as isoform expression (standard IHC practice). Report this stain as total detectable TSC1 unless epitope mapping or an independent validation supports a narrower claim (standard IHC practice).
How should I assess TSC1 in a multiplex IF experiment?
For IF/ICC, pair TSC1 with a marker identifying the expected cell type, such as myocytes in skeletal muscle, where tissue IHC reports high TSC1 staining (HPA tissue IHC; standard IF practice). Choose a fluorophore channel with low tissue autofluorescence and inspect single-channel controls before interpreting overlap (standard IF practice). Permeabilise sufficiently to access cytosolic and lysosome-associated epitopes, since TSC1 is mainly cytosolic and can be recruited to lysosomal membranes (HPA subcellular; UniProt Q92574 localisation; standard IF practice). The selected 2 μg/ml overnight condition comes from chromogenic paraffin-section IHC and does not establish an IF concentration or IF validation (datasheet A00365-2).
How can I separate TSC1 signal from diffuse DAB background?
The selected image used 10% goat serum blocking, a peroxidase-conjugated secondary for 30 minutes at 37°C, and DAB development (datasheet A00365-2). Compare a section without primary antibody and use an endogenous peroxidase block as part of the chromogenic workflow to locate background from detection chemistry (standard IHC practice). If color is widespread, check whether blocking, washing or DAB development produces similar staining in the control before changing the primary concentration (standard IHC practice). Fibroblasts and adipocytes have low reported staining, but neither is a validated negative control; use their appearance only as contextual evidence (HPA tissue IHC).
What is a defensible way to score TSC1 staining across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Score cytoplasmic staining within a defined cell population, since tissue IHC describes ubiquitous cytoplasmic TSC1 expression (HPA tissue IHC; standard IHC practice). Record the percentage of cells at each intensity from 0 to 3 and calculate an H-score from 0 to 300, or report percentage positive using a prespecified threshold (standard IHC practice). Normalize counts to the number of evaluable cells of that type, or use positive-cell density per mm² when tissue area is the appropriate denominator (standard IHC practice). Keep retrieval, detection, exposure and scoring thresholds consistent across specimens, and exclude damaged or necrotic regions (standard IHC practice).
When is apparent TSC1 staining likely to be artefactual?
Convincing staining should favor the cytoplasm of identifiable cells; high staining is reported in skeletal-muscle myocytes, heart cardiomyocytes and bone-marrow hematopoietic cells (HPA tissue IHC). Question nuclear-only color, signal confined to section edges or necrotic regions, and color that persists without primary antibody; these patterns warrant morphology and detection-control checks (HPA subcellular; standard IHC practice). Endogenous peroxidase can contribute DAB color, so evaluate the peroxidase-block control before assigning diffuse staining to TSC1 (standard IHC practice). Because nutrient state can shift the TSC1 complex between lysosomal membranes and cytosol, staining distribution alone does not measure mTORC1 activity (UniProt Q92574 localisation and function).
Boster reagents
Best TSC1 / Hamartin IHC Antibodies
Anti-TSC1 catalog antibodies have IHC images from human paraffin sections and rat brain tissue, plus IF/ICC images from cultured cells (catalog image captions).
Real IHC data
Anti-Hamartin/TSC1 Antibody ®
Cat # A00365-2
Real IHC data
Anti-Hamartin/TSC1 Antibody ®
Cat # PA2012
Real IF data
Anti-Hamartin TSC1 Antibody
Cat # A00365
A00365-2 has IHC images from human paraffin sections and an IF image from A549 cells; PA2012 has IHC images from human mammary cancer and rat brain tissue (catalog image captions). A00365 has ICC and IF images from EL4 and L1210 cells, respectively (A00365 image captions).
Which to pick: Choose A00365-2 for human paraffin-section chromogenic IHC: its caption documents EDTA retrieval at pH 8.0, 2 μg/ml primary antibody, and peroxidase/DAB detection; the fixative is unreported (A00365-2 IHC caption). For IF/ICC, A00365-2 has a human A549 IF image, while A00365 has EL4 ICC and L1210 IF images (A00365-2 and A00365 image captions). For cross-species tissue IHC, PA2012 lists human, mouse, and rat reactivity and IHC-P use, with images from human mammary cancer and rat brain tissue (PA2012 catalog applications, reactivity, and IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q92574 (TSC1_HUMAN, Hamartin).
- Human Protein Atlas. TSC1 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. TSC1 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the lipid droplets, primary cilium transition zone and basal body..
- Human Protein Atlas. TSC1 antibody validation summary (3 antibodies).
- MicroRNA-92b-3p is a prognostic oncomiR that targets TSC1 in clear cell renal cell carcinoma. Cancer science 2020 — PMC7156823.
- Familial multifocal micronodular pneumocyte hyperplasia with a novel splicing mutation in TSC1: Three cases in one family. PloS one 2019 — PMC6386448.
- Helicobacter pylori activates SLFN4(+)MDSCs to accelerate gastric intestinal metaplasia. iScience 2024 — PMC11607531.
- Bladder tumour-derived somatic TSC1 missense mutations cause loss of function via distinct mechanisms. Human molecular genetics 2008 — PMC2427143.
- PubMed PMID:9242607 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15164053 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TSC1 (Hamartin). Expected localisation Cytoplasmic staining in tissue sections (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A00365-2). Validated IHC antibody: A00365-2 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Bone marrow (Hematopoietic cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A00365-2); 2. Fixation: Image fixative and duration unreported (datasheet A00365-2); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: EDTA pH 8.0 (datasheet A00365-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A00365-2); 8. Primary antibody: Rabbit anti-TSC1, 2-5 μg/ml (datasheet A00365-2); 9. Primary incubation: Overnight at 4 °C (datasheet A00365-2). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A00365-2); Caveat: Endogenous peroxidase may confound bone marrow staining (HPA tissue IHC; standard IHC practice); Isoform / epitope: Two isoforms; antibody epitope coverage is unspecified (UniProt); Regulation: Nutrients redistribute TSC1 within cells (UniProt). Evidence: UniProt Q92574, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the lipid…, Datasheet A00365-2, PMC7156823, PMC11607531. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TSC1/immunohistochemistry-ihc-guide.