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TTLL12 / Tubulin--tyrosine ligase-like protein 12 · IHC design guide
Design Immunohistochemistry for TTLL12
Plan chromogenic TTLL12 IHC in paraffin sections using the cytoplasmic and nuclear tissue pattern (HPA tissue IHC). Kidney tubular and prostate glandular cells provide high-staining references (HPA tissue IHC); start with 2–5 μg/ml of the IHC-validated antibody (datasheet A14589-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TTLL12 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 nuclear staining in most tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic and nuclear; high in kidney tubular cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A14589-1) | |
| Positive control | Kidney+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent between sections. (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes have no detectable signal (HPA tissue IHC) | |
| Regulation | Increased in prostatic intraepithelial neoplasia and metastases (UniProt) | |
| Isoform / epitope | No isoforms; one 1–644 chain, no extracellular domain (UniProt) |
Section 1
Recommended TTLL12 IHC & IF Protocols
The catalog antibody's IHC-P protocol (datasheet A14589-1) is followed by 2 published TTLL12 paraffin-section IHC protocols (PMC13474887; PMC12142039).
| Sample | Paraffin-embedded human colorectal adenocarcinoma tissue; fixative not specified (datasheet A14589-1) |
| Fixation | Image fixative and duration unreported (datasheet A14589-1); 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 A14589-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A14589-1) |
| Primary antibody | Rabbit anti-TTLL12, 2-5 μg/ml (datasheet A14589-1) |
| Primary incubation | Overnight at 4 °C (datasheet A14589-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A14589-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | TTLL12-positive staining in cells in tubules of kidney (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nuclear expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A14589-1). Both published protocols used citrate retrieval (PMC13474887; PMC12142039).
Section 2
What Is the Expected TTLL12 Staining Pattern?
TTLL12 is predominantly cytoplasmic, with nuclear localization also reported; it has no transmembrane segment (UniProt Q14166 localization and topology). In paraffin-section IHC, expect cytoplasmic and nuclear staining in many tissues, especially kidney tubular cells and prostate glandular cells (HPA tissue IHC: High in both). HPA rates the tissue pattern Approved, pending external verification (HPA tissue IHC reliability).
| Kidney tubules or prostate glands show clear cytoplasmic staining, with some nuclear staining. | This fits the reported high staining in kidney tubular cells and prostate glandular cells and the broader cytoplasmic and nuclear tissue pattern (HPA tissue IHC). Score the identified cells and compartments separately; a single field does not establish uniform expression throughout the tissue (general IHC practice). |
| Signal is confined to extracellular material, a gland lumen, or an isolated tissue edge. | Those locations do not match the reported intracellular pattern (HPA tissue IHC; UniProt Q14166 localization). Treat the result as suspect until a known-positive section and detection controls reproduce the expected cellular distribution (general IHC practice). Nuclear staining alone warrants review because nuclear localization is reported (HPA tissue IHC; UniProt Q14166 localization). |
| Adipocytes, cardiomyocytes, or skeletal myocytes stain strongly while expected positive cells do not. | HPA reports TTLL12 as not detected in these cell types (HPA tissue IHC). Check cell identification and compare the staining with a matched negative detection control; cross-reactivity or endogenous detection activity can produce apparent positives (general IHC practice). |
| A diffuse chromogen haze covers cells, stroma, and empty areas, obscuring cell boundaries. | A field-wide haze cannot be scored reliably as TTLL12 because HPA describes cellular cytoplasmic and nuclear staining (HPA tissue IHC). Background can arise from detection reagents or inadequate blocking or washing; assess a negative detection control before interpreting intensity (general IHC practice). |
| Kidney tubular cells and prostate glandular cells show no interpretable signal. | These are reported high-staining comparators, so a blank result calls for a technical check before concluding that the tested sample lacks TTLL12 (HPA tissue IHC). Review section integrity, retrieval, antibody incubation, and chromogenic detection using the validated IHC workflow (general IHC practice). |
💡Expected TTLL12 appearanceCall a positive result when kidney tubular cells or prostate glandular cells show clear, high cytoplasmic staining with permissible nuclear staining; isolated extracellular deposits or broad cell-free haze are suspect (HPA tissue IHC: High in both; HPA tissue profile; general IHC practice).
| Tissue and cell selection (HPA tissue IHC) | Kidney tubular cells and prostate glandular cells are high-staining comparators; adrenal and breast glandular cells are reported medium (HPA tissue IHC). Choose an identified cell population for scoring, since a tissue-wide average can hide differences among cells (general IHC practice). |
| Subcellular interpretation (HPA tissue IHC; UniProt Q14166) | The tissue profile supports cytoplasmic and nuclear staining, while UniProt describes predominantly cytoplasmic localization and also lists nucleus, centrosome, spindle, and midbody (HPA tissue IHC; UniProt Q14166 localization). These annotations do not require every structure to be resolved in chromogenic sections (general IHC practice). |
| Antibody evidence (HPA antibody validation; HPA tissue IHC) | HPA003054 and CAB046029 each have Approved IHC status (HPA antibody validation). The tissue profile is Approved but pending external verification (HPA tissue IHC reliability); do not treat either designation as proof that every observed signal is specific. |
| Topology and processing (UniProt Q14166) | TTLL12 has no transmembrane segment, signal peptide, or propeptide, and the listed chain spans residues 1–644 (UniProt Q14166 topology and processing). A sharply extracellular or luminal pattern therefore needs control-based review rather than an assumption of secreted target (UniProt Q14166 topology; general IHC practice). |
| IF/ICC question: what localization is expected? (HPA subcellular ICC-IF) | Mainly cytosol, with supported nucleoplasm and uncertain plasma membrane localization (HPA subcellular ICC-IF). This answers the localization question for IF/ICC; the present staining interpretation concerns paraffin-section IHC (HPA tissue IHC). |
| Fixation sensitivity (source scope) | Target-specific fixation effects are not established by the supplied assay evidence. Verify with a matched IHC source before attributing a result to fixation. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive kidney tubules are blank. | The reported high-staining comparator failed, so sample or assay failure is possible (HPA tissue IHC: High in kidney tubular cells; general IHC practice). | Confirm that tubules are present and preserved; review retrieval, primary-antibody incubation, and chromogenic detection against the antibody's IHC workflow (general IHC practice). |
| Prostate section looks negative despite intact glands. | HPA reports high staining in prostate glandular cells, but a section-level negative needs cell-specific assessment (HPA tissue IHC). | Inspect the glandular cells at suitable magnification and run a known-positive comparator through the same staining batch (HPA tissue IHC; general IHC practice). |
| Nuclei stain but cytoplasm is weak. | Nuclear signal can be genuine, although the reported tissue profile includes cytoplasm and UniProt describes predominantly cytoplasmic localization (HPA tissue IHC; UniProt Q14166 localization). | Compare staining across identified positive cells, then check the detection control and IHC workflow before scoring an exclusively nuclear pattern (general IHC practice). |
| Adipocytes or muscle cells appear strongly positive. | Adipocytes, cardiomyocytes, and skeletal myocytes are reported as not detected; apparent staining may reflect background or cross-reactivity (HPA tissue IHC; general IHC practice). | Verify cell identity and compare with a matched negative detection control and a high-staining kidney or prostate comparator (HPA tissue IHC; general IHC practice). |
| Brown haze obscures cellular boundaries. | Diffuse background prevents confident assignment to cytoplasm or nuclei (HPA tissue IHC profile; general IHC practice). | Check a negative detection control; review blocking, washes, primary-antibody concentration, and development time within the IHC workflow (general IHC practice). |
| Signal appears mainly in luminal debris or along damaged edges. | This distribution differs from the reported intracellular localization and may reflect staining artefact (HPA tissue IHC; UniProt Q14166 localization; general IHC practice). | Exclude damaged regions from scoring, compare intact cells on the same section, and repeat with the IHC-validated antibody workflow if the pattern persists (general IHC practice). |
Experiment evidence
IHC & IF images showing TTLL12 staining
A14589-1 · IHCIHC analysis of TTLL12 using anti-TTLL12 antibody (A14589-1). TTLL12 was detected in a paraffin-embedded section of human colorectal adenoca
A14589-1 · IHCIHC analysis of TTLL12 using anti-TTLL12 antibody (A14589-1). TTLL12 was detected in a paraffin-embedded section of human thyroid cancer tis
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000100304-TTLL12/tissue/cerebral+cortex#imid_1060256
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000100304-TTLL12/tissue/colon#imid_1060182
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000100304-TTLL12/tissue/liver#imid_1060183
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000100304-TTLL12/tissue/kidney#imid_1060184
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003054; green = TTLL12, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003054; green = TTLL12, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TTLL12 IHC & IF
🧪Run kidney first: tubular cells should stain strongly (HPA: High in kidney cells in tubules). Use adipose tissue as a negative comparator, with little or no staining in adipocytes (HPA: Not detected in adipocytes); no cell type within the kidney slide is established as an internal negative by the supplied HPA rows, so assess adjacent cells for background without assuming they lack TTLL12.
Positive control tissue: Kidney (Cells in tubules, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TTLL12 in A-431, U-251MG, U2OS, RPTEC/TERT1, KOLF2.1J, with annotated localisation: Cytosol (supported) (HPA subcellular).
Technical controls: Include a no-primary (secondary-only) control, a concentration-matched rabbit IgG isotype control, and TTLL12 knockout material or a cognate-peptide block if available (selected-SKU caption: rabbit primary antibody; standard IHC practice). For kidney HRP/DAB staining, quench endogenous peroxidase and check any residual tubular signal in the no-primary control (standard IHC practice).
⚠️Feasibility: Paraffin-section IHC is demonstrated with heat retrieval in EDTA at pH 8.0, but the selected-SKU image used colorectal adenocarcinoma, so its conditions do not establish retrieval requirements for kidney (selected-SKU tissue-IHC caption). A target-specific fixation window or fixation effect is unreported, and the caption does not state a fixative (selected-SKU tissue-IHC caption); the supplied evidence also does not establish that frozen sections or IF are easier. Kidney tubules can show endogenous detection background, so interpret tubular staining against the no-primary slide (standard IHC practice).
HPA tissue IHC evidence for TTLL12
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Prostate | Glandular cells | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Salivary gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TTLL12 IHC Tips
Troubleshoot TTLL12 chromogenic IHC in paraffin sections using the catalog antibody’s tissue result and compartment-specific controls.
What retrieval should I use if TTLL12 staining is weak in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet A14589-1). The catalog antibody detected TTLL12 in a paraffin section after this retrieval, followed by 2 μg/ml primary antibody overnight at 4°C (datasheet A14589-1). If staining is weak, compare retrieval durations on adjacent sections while keeping antibody concentration and detection constant (standard IHC practice). If that fails, test citrate at pH 6.0 as a fallback, with a matched positive control (standard IHC practice). Judge improvement by cellular staining and tissue preservation, since excessive heating can damage morphology (standard IHC practice).
How should I troubleshoot variable TTLL12 staining between paraffin blocks?
Target-specific fixation sensitivity is unknown: the selected tissue caption describes a paraffin section but does not state its fixative (datasheet A14589-1). Record each block’s fixative and fixation duration, and compare sections processed with the same EDTA pH 8.0 retrieval and detection conditions (standard IHC practice; datasheet A14589-1). Include a positive control section in each run so a global loss of signal is distinguishable from a block-specific problem (standard IHC practice). Check whether tissue morphology and the staining pattern remain interpretable before changing antibody concentration (standard IHC practice). Do not use tissue expression patterns to infer a TTLL12-specific fixation effect.
Where should convincing TTLL12 signal appear within stained cells?
Expect predominantly cytoplasmic staining, with possible nuclear staining, when evaluating TTLL12 IHC (UniProt Q14166 subcellular location; HPA: cytoplasmic and nuclear expression in most tissues). HPA subcellular imaging supports cytosolic and nucleoplasmic localisation, while its plasma-membrane assignment is uncertain (HPA subcellular). UniProt also places TTLL12 at the midbody, centrosome, and spindle, so focal intracellular signal may merit examination in mitotic cells (UniProt Q14166 subcellular location). Assess compartment and cell identity together, preferably against a matched positive section (standard IHC practice). Treat isolated crisp membrane-only DAB as questionable until an independent control supports it (HPA subcellular: plasma membrane uncertain; standard IHC practice).
Could epitope position explain different cytoplasmic and nuclear IHC patterns?
Check the antibody’s immunogen or epitope documentation before attributing a pattern difference to epitope selection (standard IHC practice). The supplied record lists 0 annotated isoforms, a 300–644 TTL domain, and a 1–644 chain, but gives no antibody epitope position (UniProt Q14166). TTLL12 has no annotated transmembrane segment, signal peptide, or glycosylation site in this record (UniProt Q14166). Those annotations cannot establish whether this antibody detects a fixation-sensitive epitope or explain a compartment shift (standard IHC interpretation). Compare antibodies with documented, distinct epitopes on adjacent sections if epitope dependence remains a concern (standard IHC practice).
How can I examine TTLL12 localisation by multiplex IF alongside IHC?
Use IF as a separate assay and verify its antibody conditions independently; the selected catalog evidence is paraffin-section chromogenic IHC (datasheet A14589-1). For prostate glandular cells, pair TTLL12 with an epithelial marker such as pan-cytokeratin to identify the scored population (HPA: high in prostate glandular cells; standard IF practice). Choose spectrally separated fluorophores and place the weaker target in a channel with low tissue autofluorescence, checking single-stain controls for bleed-through (standard IF practice). Because TTLL12 is intracellular and has no annotated transmembrane segment, validate permeabilisation that permits cytosolic and nuclear access while preserving morphology (UniProt Q14166; standard IF practice). Compare compartment patterns with IHC, accounting for assay-specific detection (standard IF practice).
What should I check when TTLL12 DAB staining looks diffuse or widespread?
Begin with a no-primary control to assess secondary-reagent and chromogen background, and use a peroxidase block before DAB development (standard IHC practice). The selected demonstration used 10% goat-serum blocking, 2 μg/ml primary antibody overnight at 4°C, and an anti-rabbit secondary for 30 minutes at 37°C (datasheet A14589-1). If background persists, titrate primary concentration and development time while retaining a positive control and identical section handling (standard IHC practice). Diffuse cytoplasmic staining alone is insufficient evidence of artefact because TTLL12 is predominantly cytoplasmic (UniProt Q14166). Compare cell boundaries, nuclear counterstain, and no-primary signal before deciding what to score (standard IHC practice).
How should I quantify TTLL12 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because TTLL12 can appear in both cytoplasm and nucleus (HPA tissue IHC; UniProt Q14166 subcellular location). For a defined region, report the percentage of positive cells and a compartment-specific H-score using intensity classes 0–3 (standard IHC practice). If counting focal positive cells, report density per mm² of viable tissue and state the sampled area (standard IHC practice). Normalise counts to the number of eligible cells or viable area, and apply the same threshold, counterstain, and imaging settings across sections (standard IHC practice). Record excluded necrosis and edge regions before comparing groups (standard IHC practice).
How do I distinguish true TTLL12 positivity from staining artefacts?
A credible result should show cellular cytoplasmic signal, with possible nuclear signal, rather than an isolated membrane-only outline (UniProt Q14166 subcellular location; HPA subcellular: plasma membrane uncertain). Use a positive control with documented expression, such as prostate glandular cells or kidney tubular cells, while recognizing that HPA tissue IHC is approved pending external verification (HPA: high in both cell populations; HPA reliability). Check whether apparent positives follow viable cells instead of section edges or necrotic areas (standard IHC practice). Compare a no-primary section and the peroxidase-blocked section to identify nonspecific DAB or endogenous enzyme signal (standard IHC practice). Interpret unexpected cell populations cautiously and confirm them with an independent control (standard IHC practice).
Boster reagents
Best TTLL12 / Tubulin--tyrosine ligase-like protein 12 IHC Antibodies
A14589-1 has real IHC images from human paraffin-embedded colorectal adenocarcinoma and thyroid cancer sections (catalog IHC image captions); no IF image is supplied (catalog IF image alts).
Real IHC data
Anti-TTLL12 Antibody ®
Cat # A14589-1
A14589-1 is listed for human IHC (catalog applications and reactivity). Its images show staining in paraffin-embedded human colorectal adenocarcinoma and thyroid cancer sections (catalog IHC image captions).
Which to pick: Choose A14589-1 for human paraffin-section IHC: it is a rabbit antibody listed for human IHC, with images from those sections (catalog host, applications, reactivity, and IHC image captions). No SKU in the payload lists IF/ICC or nonhuman reactivity, so neither use has catalog support here (catalog applications and reactivity). The captions do not report a fixative; fixation and tissue processing cannot be inferred (catalog IHC image captions). The selected A14589-1 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image A14589-1).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q14166 (TTL12_HUMAN, Tubulin--tyrosine ligase-like protein 12).
- Human Protein Atlas. TTLL12 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. TTLL12 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoplasm and plasma membrane..
- Human Protein Atlas. TTLL12 antibody validation summary (2 antibodies).
- Tubulin-tyrosine ligase-like protein 12(TTLL12): elevated expression in oral squamous cell carcinoma and its clinical prognostic implications. BMC oral health 2026 — PMC13474887.
- Tumor-intrinsic TTLL12 drives resistance to cancer immunotherapy via modulating myeloid-derived suppressor cells. Journal for immunotherapy of cancer 2025 — PMC12142039.
- Downregulated genes by silencing MYC pathway identified with RNA-SEQ analysis as potential prognostic biomarkers in gastric adenocarcinoma. Aging 2020 — PMC7803532.
- PubMed PMID:15461802 — UniProt-cited evidence.
- PubMed PMID:10591208 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TTLL12 (Tubulin--tyrosine ligase-like protein 12). Expected localisation Cytoplasmic and nuclear staining in most tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A14589-1). Validated IHC antibody: A14589-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Kidney (Cells in tubules); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colorectal adenocarcinoma tissue; fixative not specified (datasheet A14589-1); 2. Fixation: Image fixative and duration unreported (datasheet A14589-1); 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 A14589-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A14589-1); 8. Primary antibody: Rabbit anti-TTLL12, 2-5 μg/ml (datasheet A14589-1); 9. Primary incubation: Overnight at 4 °C (datasheet A14589-1). Watch for: Fixation: Keep fixation consistent between sections. (standard IHC practice; not target-specific); Caveat: Adipocytes have no detectable signal (HPA tissue IHC); Isoform / epitope: No isoforms; one 1–644 chain, no extracellular domain (UniProt); Regulation: Increased in prostatic intraepithelial neoplasia and metastases (UniProt). Evidence: UniProt Q14166, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the…, Datasheet A14589-1, PMC13474887, PMC12142039. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TTLL12/immunohistochemistry-ihc-guide.