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CLTB / Clathrin light chain B · IHC design guide
Design Immunohistochemistry for CLTB
Plan chromogenic CLTB IHC on paraffin sections using 2–5 μg/ml of the IHC-validated antibody (datasheet A09071-1). Compare staining with the often granular cytoplasmic tissue pattern and account for its medium consistency with RNA data (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CLTB 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 | Granular cytoplasmic staining in tissue (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining, often granular, across cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A09071-1) | |
| Positive control | Epididymis+4 more · see all | |
| Negative control | Caudate |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent (standard IHC practice; not target-specific) | |
| Caveat | Staining has medium consistency with RNA data (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | 2 isoforms, brain and non-brain; map the epitope (UniProt) |
Section 1
Recommended CLTB IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: A09071-1) is accompanied by published paraffin-section methods from two CLTB studies (PMC12622436; PMC10157171).
| Sample | Paraffin-embedded human endometrial cancer tissue; fixative not specified (datasheet A09071-1) |
| Fixation | Image fixative and duration unreported (datasheet A09071-1); 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 A09071-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A09071-1) |
| Primary antibody | Rabbit anti-CLTB, 2-5 μg/ml (datasheet A09071-1) |
| Primary incubation | Overnight at 4 °C (datasheet A09071-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A09071-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CLTB-positive staining in glandular cells of epididymis (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression often with a granular pattern. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet: A09071-1); neither article specifies its retrieval conditions (PMC12622436; PMC10157171).
Section 2
What Is the Expected CLTB Staining Pattern?
CLTB sits on the cytoplasmic face of clathrin-coated pits and vesicles; it has no transmembrane segment (UniProt P09497 topology). In paraffin sections, expect generally cytoplasmic, often granular staining, including strong staining in epididymal and small-intestinal glandular cells (HPA tissue IHC: High). HPA rates the tissue pattern Approved, with medium consistency between staining and RNA data and external verification pending (HPA tissue IHC reliability).
| Granular cytoplasmic staining in epididymal or small-intestinal glandular cells. | This matches the reported High staining in those cells (HPA tissue IHC). Granularity fits CLTB's association with coated vesicles (UniProt P09497 localization). Judge the pattern in the cells, rather than treating uniform colour across the whole section as the expected result. |
| Predominantly nuclear staining, with little cytoplasmic signal. | A nuclear-only pattern conflicts with the reported cytoplasmic tissue pattern and cytoplasmic-face localization (HPA tissue IHC; UniProt P09497 topology). Treat it as suspect and check staining controls and antibody specificity before calling the section CLTB-positive (standard IHC practice). |
| Strong staining in caudate glial cells, with otherwise weak expected staining. | HPA reports CLTB as Not detected in caudate glial cells (HPA tissue IHC). Strong staining there warrants investigation for cross-reactivity or endogenous chromogen-producing activity; that single observation does not prove either cause (standard IHC interpretation). |
| Diffuse colour over cells, stroma, and empty spaces. | This obscures the general cytoplasmic, often granular pattern reported for CLTB (HPA tissue IHC). Consider nonspecific background, residual detection activity, or excessive development, using a no-primary control to locate signal generated by the detection system (standard chromogenic IHC practice). |
| No visible staining in epididymal or small-intestinal glandular cells. | Absence in these reported High reference populations makes a negative result difficult to interpret (HPA tissue IHC). Review section quality, antigen retrieval, primary-antibody conditions, and detection controls before concluding that the test specimen lacks CLTB (standard IHC practice). |
💡Expected CLTB appearanceCall a section positive when glandular cells show predominantly granular cytoplasmic staining, expected to be High in epididymis or small intestine (HPA tissue IHC), while nuclear-only colour or widespread diffuse background is suspect given CLTB's cytoplasmic-face localization (UniProt P09497 topology; standard IHC interpretation).
| Subcellular location and topology (UniProt P09497) | CLTB associates with the cytoplasmic face of coated pits and vesicles and has no transmembrane segment (UniProt P09497). Score a cytoplasmic pattern; the annotation does not specify an IHC antigen-retrieval setting or establish fixation sensitivity. |
| Reference tissue and cell population (HPA tissue IHC) | Epididymal and small-intestinal glandular cells are reported High; adrenal glandular cells and bone-marrow hematopoietic cells are Medium (HPA tissue IHC). Compare the same cell population when assessing intensity, because HPA also reports lower staining in some tissues. |
| Strength of tissue-pattern evidence (HPA tissue IHC) | The tissue staining is Approved, with medium consistency against RNA expression and external verification pending (HPA tissue IHC reliability). Use it as a reference pattern, while keeping a discordant result open to technical or biological explanations. |
| Brain and non-brain isoforms (UniProt P09497) | UniProt lists 2 isoforms, Brain and Non-brain (UniProt P09497 isoforms). The supplied evidence does not identify the catalog antibody's epitope or establish equal recognition of both, so do not infer isoform-specific staining from intensity alone. |
| IF/ICC Q: what localization is expected? (HPA subcellular ICC-IF) | A: Mainly vesicles, with additional plasma-membrane and cytosolic localization (HPA subcellular ICC-IF). Those cell-image observations can inform compartment interpretation; they do not establish the antibody's paraffin-section performance (HPA antibody validation). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive glandular cells appear unstained. | The expected reference is High in epididymis and small intestine (HPA tissue IHC); absent colour could reflect a failed staining step (standard IHC practice). | Check section integrity and detection controls, then review the antibody's IHC-P conditions and antigen retrieval before interpreting the specimen as negative (standard IHC practice). |
| Staining is weak but confined to the expected cytoplasm. | HPA reports intensity differences among tissues, including Medium and Low populations (HPA tissue IHC); weak colour alone cannot identify the cause. | Compare matching cell populations and a reported High reference on the same run; assess signal against background before changing IHC conditions (HPA tissue IHC; standard IHC practice). |
| Nuclei stain more strongly than cytoplasm. | Nuclear dominance conflicts with CLTB's reported cytoplasmic tissue pattern and coated-pit or vesicle association (HPA tissue IHC; UniProt P09497 localization). | Inspect a no-primary control and review antibody concentration and detection conditions; withhold a positive localization call until cytoplasmic staining is convincing (standard IHC practice). |
| Brown colour persists in the no-primary control. | Signal without primary antibody points to the detection system or endogenous activity, rather than establishing CLTB binding (standard chromogenic IHC practice). | Check the detection reagents and endogenous-enzyme blocking appropriate to the chromogen system; repeat with the no-primary control alongside the specimen (standard chromogenic IHC practice). |
| Colour is widespread and masks cellular detail. | Diffuse background makes HPA's often granular cytoplasmic pattern hard to assess (HPA tissue IHC); nonspecific staining or overdevelopment are possible (standard IHC practice). | Compare the no-primary control, review blocking and washes, and adjust primary-antibody or chromogen conditions using the antibody's IHC-P guidance (standard IHC practice). |
| An IF/ICC vesicle pattern is used to justify an IHC-P result. | HPA reports vesicles as the main ICC-IF location, while its antibody validation records ICC and IHC separately (HPA subcellular ICC-IF; HPA antibody validation). | Interpret the paraffin section against HPA tissue IHC cell and compartment patterns and its own controls; use the separate IF/ICC guide for fluorescence-specific decisions (HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing CLTB staining
A09071-1 · IHCIHC analysis of CLTB using anti-CLTB antibody (A09071-1). CLTB was detected in a paraffin-embedded section of human endometrial cancer tissu
A09071-1 · IHCIHC analysis of CLTB using anti-CLTB antibody (A09071-1). CLTB was detected in a paraffin-embedded section of human pancreas cancer tissue.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000175416-CLTB/tissue/cerebral+cortex#imid_1267644
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000175416-CLTB/tissue/colon#imid_1267586
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000175416-CLTB/tissue/liver#imid_1267579
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000175416-CLTB/tissue/kidney#imid_1267580
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036458; green = CLTB, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036458; green = CLTB, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CLTB IHC & IF
🧪Run small intestine first: glandular cells should stain strongly (HPA: High in small-intestine glandular cells). Use caudate glial cells as the negative tissue (HPA: Not detected); on the positive slide, treat adjacent nonglandular cells with little or no staining as an internal background reference, without assuming they are CLTB-negative.
Positive control tissue: Epididymis (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CLTB in A-431, MCF-7, U2OS, NIH 3T3, with annotated localisation: Vesicles (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, matched nonimmune rabbit IgG control, and CLTB-knockout material processed in parallel as a biological negative (selected IHC caption: rabbit primary antibody). For small-intestine IHC, quench endogenous peroxidase and block endogenous biotin if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A09071-1 paraffin-section caption does not state the fixative. That caption used heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required. IF/ICC can assess the expected vesicular pattern (HPA: vesicles supported); the supplied evidence does not establish whether IF or frozen sections are easier than paraffin IHC, and endogenous biotin may cause small-intestine background with avidin–biotin detection (standard IHC practice).
HPA tissue IHC evidence for CLTB
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 |
|---|---|---|---|---|
| Epididymis | Glandular cells | High | Protein (IHC) | HPA → |
| Small intestine | Glandular cells | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CLTB IHC Tips
Use the catalog antibody’s paraffin-section conditions as a starting point, then judge CLTB staining by its expected compartment and tissue pattern.
Which retrieval condition should I start with for weak CLTB staining in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A09071-1). The selected tissue-IHC example used that condition before incubation with 2 μg/ml primary antibody overnight at 4°C (caption A09071-1). If staining remains weak, adjust heating duration in small steps while keeping buffer and detection conditions constant, and compare intact tissue regions across slides (standard IHC practice). Assess whether the change restores a granular cytoplasmic pattern, rather than merely increasing diffuse color or tissue damage (HPA: general cytoplasmic expression often granular; standard IHC practice).
Could fixation explain inconsistent CLTB staining between paraffin blocks?
Yes, differences in fixation can change antigen accessibility in paraffin sections, so document fixative, fixation duration, processing, and block age for each comparison (standard IHC practice). The selected CLTB caption does not state a fixative, so target-specific fixation sensitivity is unknown (caption A09071-1). Compare sections with matched processing first, using EDTA retrieval at pH 8.0 and the same antibody incubation to limit other sources of variation (datasheet A09071-1; standard IHC practice). If a block stains poorly, inspect morphology and include a concurrently processed positive control before attributing the result to CLTB abundance (standard IHC practice).
What CLTB distribution should I expect in a positive tissue section?
Look for cytoplasmic staining that may appear granular in positive cells (HPA: general cytoplasmic expression often granular). CLTB associates with the cytoplasmic face of coated pits and vesicles and has no transmembrane segment, so staining need not trace a continuous cell boundary (UniProt P09497: localisation and topology). Vesicles are its main supported subcellular location, with additional plasma-membrane and cytosolic localisation reported (HPA: subcellular localisation). Use glandular cells in epididymis or small intestine as contextual positive comparators where available, while comparing staining within equivalent tissue compartments (HPA: high glandular-cell staining; standard IHC practice).
Could isoforms or epitope accessibility change my CLTB IHC result?
CLTB has brain and non-brain isoforms, so an unmapped antibody epitope leaves isoform coverage uncertain (UniProt P09497: isoforms; supplied antibody evidence: no epitope map). It also has annotated modified residues, including phosphoserines at 11 and 13 and phosphothreonine at 187, but their effect on this antibody has not been established (UniProt P09497: modified residues; supplied antibody evidence: no epitope map). Keep retrieval and staining conditions matched when comparing regions, because an intensity difference alone cannot identify an isoform (standard IHC practice). If isoform specificity matters, obtain epitope information and validate with an independent assay before assigning a staining pattern to either isoform (standard IHC practice).
How should I assess CLTB localisation by multiplex IF?
Pair CLTB with a marker of the cell type being assessed; glandular cells are a useful tissue context because they show high IHC staining in epididymis and small intestine (HPA: tissue IHC). Choose spectrally separated fluorophores and place the weaker signal in a red or far-red channel after checking tissue autofluorescence and single-label controls (standard IF practice). If permeabilisation is required, titrate it gently because CLTB lies on the cytoplasmic face of coated pits and vesicles, where antibody access matters (UniProt P09497: topology; standard IF practice). Interpret puncta against the reported vesicular, plasma-membrane, and cytosolic locations, while treating IF performance of this catalog antibody as unestablished by the supplied tissue-IHC caption (HPA: subcellular localisation; caption A09071-1).
How can I reduce diffuse or widespread DAB background in CLTB sections?
First compare a no-primary control with the stained section to locate background from detection reagents, endogenous peroxidase, or tissue pigment (standard chromogenic IHC practice). The selected example blocked with 10% goat serum and used a peroxidase-conjugated secondary for 30 minutes at 37°C, followed by DAB development (caption A09071-1). For a new tissue, include an endogenous-peroxidase block and optimise washes, primary concentration, and DAB development while changing one variable at a time (standard IHC practice). Retain conditions that reveal cell-associated granular cytoplasmic signal without uniform extracellular or section-wide color (HPA: tissue-IHC profile; standard IHC practice).
How should I quantify CLTB IHC across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the tissue compartment and eligible cells before scoring, then record the percentage of positive cells and staining intensity using the same thresholds across slides (standard IHC practice). An H-score can combine intensity grades 0–3 with their cell percentages, yielding 0–300; alternatively, measure positive-cell density per mm² in a fixed region of interest (standard IHC practice). Normalise counts to viable tissue area or the number of eligible cells, and keep imaging, counterstain, and DAB thresholds consistent (standard IHC practice). Report granular cytoplasmic staining separately from nonspecific diffuse signal, since the reported tissue pattern is often granular and cytoplasmic (HPA: tissue-IHC profile).
How do I distinguish genuine CLTB staining from an IHC artefact?
A plausible positive signal is cell-associated and cytoplasmic, often granular, consistent with CLTB at coated pits and vesicles (HPA: tissue-IHC profile; UniProt P09497: localisation). Compare the cell type with the tissue context: glandular cells stain highly in epididymis and small intestine, whereas caudate glial cells were reported as not detected (HPA: tissue IHC). Treat strong nuclear-only color, section-edge enhancement, and staining confined to necrotic areas as suspect until controls support it (UniProt P09497: localisation; standard IHC practice). Check a no-primary slide for endogenous peroxidase or other DAB signal, and interpret any negative result alongside section quality and a positive control (standard chromogenic IHC practice).
Boster reagents
Best CLTB / Clathrin light chain B IHC Antibodies
A09071-1 has real IHC images from paraffin sections of human endometrial and pancreas cancers and mouse and rat cerebellum (catalog image captions); no IF images are supplied (catalog payload).
Real IHC data
Anti-CLTB Antibody ®
Cat # A09071-1
A09071-1 is listed for IHC in human, mouse and rat samples (catalog applications and reactivity). Its IHC captions show paraffin sections of human endometrial and pancreas cancers, mouse cerebellum and rat cerebellum (catalog image captions).
Which to pick: For tissue IHC, choose A09071-1 at the listed 2–5 μg/ml; its images support paraffin sections, and the fixative is unreported (datasheet: 2–5 μg/ml; catalog image captions). No SKU in this payload is validated for IF/ICC, and A09071-1’s clonality is unreported (catalog applications; catalog clone field). For cross-species IHC, A09071-1 lists human, mouse and rat reactivity, with a paraffin-section image for each species (catalog reactivity; catalog image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P09497 (CLCB_HUMAN, Clathrin light chain B).
- Human Protein Atlas. CLTB tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CLTB subcellular location (ICC-IF): Mainly localized to vesicles. In addition localized to the plasma membrane and cytosol..
- Human Protein Atlas. CLTB antibody validation summary (2 antibodies).
- Clathrin Light Chain B Drives Hepatocellular Carcinoma Progression Through Dual Mechanisms: Small Extracellular Vesicle-Mediated Angiogenesis and the NF-κB-PCLAF Signaling Axis. Advanced science (Weinheim, Baden-Wurttemberg, Germany) 2025 — PMC12622436.
- A novel 25-ferroptosis-related gene signature for the prognosis of gliomas. Frontiers in oncology 2023 — PMC10157171.
- PubMed PMID:3267234 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:11889126 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CLTB (Clathrin light chain B). Expected localisation Granular cytoplasmic staining in tissue (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A09071-1). Validated IHC antibody: A09071-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Epididymis (Glandular cells); negative Caudate. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human endometrial cancer tissue; fixative not specified (datasheet A09071-1); 2. Fixation: Image fixative and duration unreported (datasheet A09071-1); 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 A09071-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A09071-1); 8. Primary antibody: Rabbit anti-CLTB, 2-5 μg/ml (datasheet A09071-1); 9. Primary incubation: Overnight at 4 °C (datasheet A09071-1). Watch for: Fixation: Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Caveat: Staining has medium consistency with RNA data (HPA tissue IHC); Isoform / epitope: 2 isoforms, brain and non-brain; map the epitope (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt P09497, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to vesicles. In addition localized to the plasma…, Datasheet A09071-1, PMC12622436, PMC10157171. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CLTB/immunohistochemistry-ihc-guide.