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TUBB2A / Tubulin beta-2A chain · IHC design guide
Design Immunohistochemistry for TUBB2A
Plan chromogenic TUBB2A IHC in paraffin sections around the cytoplasmic staining reported in CNS and peripheral nerves (HPA tissue IHC). Compare cell populations with reported staining levels, and interpret positive signal in light of HPA's warning that the antibody targets products from more than one gene (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TUBB2A 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 tissue staining (HPA tissue IHC) | |
| Staining pattern | General cytoplasmic staining, strongest in CNS and peripheral nerves (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, heat-mediated (datasheet M06868) | |
| Positive control | Placenta+3 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 across sections (standard IHC practice; not target-specific). Selected-image fixative and duration unreported (datasheet M06868); verify before use. | |
| Caveat | Staining may include products from other genes (HPA tissue IHC) | |
| Regulation | Brain enriched expression (UniProt) | |
| Isoform / epitope | No annotated isoforms; one 1–445 chain (UniProt) |
Section 1
Recommended TUBB2A IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published protocols for hepatocellular carcinoma tissue (PMC13033567) and mouse xenograft tumors (PMC5390261).
| Sample | Paraffin-embedded Human colon tissue; fixative not specified (datasheet M06868) |
| Fixation | Image fixative and duration unreported (datasheet M06868); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6.0, 20min (datasheet M06868) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Mouse monoclonal (clone Mix) anti-TUBB2A, 1:50 recommended; image 1:200 (datasheet M06868) |
| 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 | TUBB2A-positive staining in endothelial cells of placenta (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression, most abundant in CNS and peripheral nerves. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6.0 (datasheet M06868); neither article reports retrieval conditions sufficient to reproduce its step (PMC13033567; PMC5390261).
Section 2
What Is the Expected TUBB2A Staining Pattern?
TUBB2A is a cytoplasmic microtubule protein with no transmembrane segment (UniProt Q13885). In IHC, expect cytoplasmic staining, including signal in peripheral nerves and CNS neuropil, with prominent staining reported in placental endothelial cells, skin granular layer cells and pachytene spermatocytes (HPA tissue IHC). Interpret positive staining cautiously: HPA rates the tissue pattern Supported, but notes that its antibodies can recognize proteins from more than one gene and that presumed off-target staining was disregarded (HPA tissue IHC).
| Cytoplasmic staining in placental endothelial cells, skin granular layer cells or pachytene spermatocytes. | This matches HPA's High IHC observations in those cell populations (HPA tissue IHC). Check the identity of stained cells and the surrounding architecture; a strong color reaction alone cannot establish TUBB2A specificity because HPA reports recognition of proteins from more than one gene (HPA tissue IHC). |
| Medium signal in cerebral cortex neuropil, or weak staining in hippocampal neuronal cells. | These intensities fit the respective Medium and Low HPA observations (HPA tissue IHC). Do not require every brain cell to stain strongly: HPA reports no detection in caudate glial cells or cerebellar granular layer cells, despite describing CNS enrichment overall (HPA tissue IHC). |
| Predominantly nuclear, membranous or extracellular staining, with little cytoplasmic signal. | The compartment disagrees with UniProt's cytoplasm and cytoskeleton assignment and its lack of a transmembrane segment (UniProt Q13885). Review slide morphology and detection controls before scoring it as specific; a compartment mismatch alone does not identify the artifact's cause (standard IHC practice). |
| Strong color in an HPA-negative population, or widespread color outside recognizable cells. | Adipocytes and adrenal glandular cells are among HPA's Not detected populations (HPA tissue IHC). Staining there warrants a specificity or endogenous detection check; widespread color may be background from the staining workflow (standard IHC practice). Neither appearance proves a particular cause. |
| No cytoplasmic signal in a well-preserved, expected-positive control section. | Compare the control with HPA's High placental endothelial, skin granular layer or pachytene spermatocyte observations (HPA tissue IHC). If it is also blank, check the primary antibody, retrieval, detection and counterstain workflow before interpreting a test section as negative (standard IHC practice). |
💡Expected TUBB2A appearanceA convincing positive has cytoplasmic staining in the expected cell population, potentially High in placental endothelial cells, skin granular layer cells or pachytene spermatocytes; isolated nuclear color or staining confined to an HPA-negative population is suspect (UniProt Q13885; HPA tissue IHC).
| Cell population and intensity | HPA reports High staining in three specified populations, Medium in cerebral cortex neuropil, Low in hippocampal neuronal cells and Not detected in several other populations (HPA tissue IHC). Score the identified cell population rather than assigning one intensity to an entire tissue. |
| Antibody specificity | The HPA tissue profile is Supported, and three listed antibodies have Supported IHC status (HPA antibodies; HPA tissue IHC). HPA also cautions that antibodies may recognize proteins from more than one gene and that presumed off-target staining was disregarded; morphology and controls therefore matter. |
| Localization and processing | UniProt places TUBB2A in the cytoplasm and cytoskeleton, with no transmembrane segment, signal peptide or propeptide; the annotated chain spans residues 1–445 (UniProt Q13885). These annotations support a cytoplasmic interpretation, but do not predict antigen retrieval requirements or fixation sensitivity. |
| Antigen retrieval in paraffin IHC | Use a documented, working retrieval condition and compare positive controls when optimizing staining (standard IHC practice). The supplied UniProt and HPA records provide no TUBB2A-specific fixation or retrieval sensitivity, so a weak result cannot be attributed to epitope masking from these sources. |
| IF/ICC Q&A: where should fluorescence appear? | Primarily along microtubules; HPA also reports the mitotic spindle, cytokinetic bridge and specified ciliary or sperm structures (HPA subcellular ICC-IF). This is an IF/ICC localization reference, while the tissue intensities above come from IHC (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected-positive cells are blank. | A failed staining run or unsuitable working conditions are possible; the supplied sources establish expected cell populations but no TUBB2A-specific fixation effect (HPA tissue IHC; standard IHC practice). | Confirm morphology, primary antibody application, retrieval and detection with a positive control containing an HPA High population; assess the test section only after that control works (HPA tissue IHC; standard IHC practice). |
| Nuclear color dominates the section. | That pattern conflicts with the cytoplasmic and cytoskeletal annotation (UniProt Q13885); the image alone cannot distinguish nonspecific binding from a workflow artifact (standard IHC practice). | Check compartment boundaries against the counterstain and compare a no-primary control. Score cytoplasmic signal separately from nuclear color (standard IHC practice). |
| An HPA Not detected cell population stains strongly. | Possible cross-reactivity merits attention because HPA cautions that its antibodies may recognize proteins from more than one gene (HPA tissue IHC). Endogenous chromogenic activity is another general IHC possibility (standard IHC practice). | Verify cell identity, compare an expected-positive population and run a no-primary detection control; investigate persistent cell-associated color with another validated antibody if available (HPA tissue IHC; standard IHC practice). |
| Color is diffuse across tissue and empty spaces. | This distribution is inconsistent with the expected cell-associated cytoplasmic pattern (UniProt Q13885; HPA tissue IHC). Background from blocking, washing or detection can produce diffuse color (standard IHC practice). | Review a no-primary control, washing and blocking, then reassess the cytoplasmic pattern in identifiable positive cells before assigning a score (standard IHC practice). |
| All brain cells appear equally intense. | HPA describes abundant CNS expression overall, yet reports Medium cortex neuropil, Low hippocampal neuronal cells and no detection in listed caudate glial and cerebellar granular layer cells (HPA tissue IHC). | Score each identified cell population and compartment separately; verify that strong, uniform color is absent from the no-primary control before calling it specific (HPA tissue IHC; standard IHC practice). |
| Two IHC antibodies give different patterns. | HPA lists three antibodies with Supported IHC status but cautions about recognition of proteins from more than one gene (HPA antibodies; HPA tissue IHC). Discordance therefore needs review rather than an automatic positive call. | Compare localization and expected cell populations on matched sections, check detection controls, and report the disagreement when assigning confidence to the result (UniProt Q13885; HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing TUBB2A staining
M06868 · IHCImmunohistochemical analysis of paraffin-embedded Human-colon tissue. 1, β II tubulin Monoclonal Antibody (Mix) was diluted at 1:200 (4°C, o
M06868 · IHCImmunohistochemical analysis of paraffin-embedded Rat-kidney tissue. 1, β II tubulin Monoclonal Antibody (Mix) was diluted at 1:200 (4°C, ov
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000137267-TUBB2A/tissue/cerebral+cortex#imid_13050179
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000137267-TUBB2A/tissue/colon#imid_13050393
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000137267-TUBB2A/tissue/liver#imid_13050394
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000137267-TUBB2A/tissue/kidney#imid_13050395
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB015339; green = TUBB2A, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB015339; green = TUBB2A, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TUBB2A IHC & IF
🧪Run placenta first: endothelial cells should stain strongly (HPA: High in placenta endothelial cells). Use adipose tissue as the negative comparator, with adipocytes at background level (HPA: Not detected in adipocytes); on the placenta slide, cells without specific staining should show only background chromogen, although HPA does not identify a particular internal negative cell type.
Positive control tissue: Placenta (Endothelial cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show TUBB2A in A-431, U-251MG, U2OS, A-549, HEL, HaCaT, MCF-7, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), Sperm, with annotated localisation: Microtubules (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and an isotype control matched to the primary antibody’s host species and clonality (standard IHC practice). Use TUBB2A knockout material as a biological negative, or an immunizing-peptide block if the peptide is available (standard IHC practice). For placenta chromogenic IHC, quench endogenous peroxidase and inspect vascular regions for background staining (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected SKU M06868 tissue-IHC caption does not state a fixative (caption: fixative not stated). The paraffin-section caption uses sodium citrate retrieval at pH 6.0 for 20 minutes above 98°C, but does not establish that retrieval is required; it reports a primary dilution of 1:200 overnight at 4°C (datasheet caption: M06868). Frozen sections or IF cannot be judged easier from the supplied evidence; in placenta IHC, vascular background from endogenous peroxidase needs attention (standard IHC practice).
HPA tissue IHC evidence for TUBB2A
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Caution, targets protein from more than one gene. Presumed off target binding observed and disregarded. External characterization data supports antibody staining.). 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 |
|---|---|---|---|---|
| Placenta | Endothelial cells | High | Protein (IHC) | HPA → |
| Skin | Cells in granular layer | High | Protein (IHC) | HPA → |
| Testis | Pachytene spermatocytes | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuropil | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced TUBB2A IHC Tips
Use the IHC-validated antibody’s paraffin-section conditions as a starting point, then judge staining by cellular location, tissue context and controls.
Which retrieval conditions should I try first for TUBB2A IHC?
Start with heat-mediated citrate retrieval at pH 6.0 (datasheet M06868). The catalog antibody’s paraffin-section image used sodium citrate above 98°C for 20 minutes, followed by primary antibody at 1:200 overnight at 4°C (caption M06868). Keep section thickness, cooling and detection conditions consistent while comparing retrieval runs, because these variables affect chromogenic signal (standard IHC practice). If staining remains weak, test a different retrieval buffer or heating duration on matched sections, with the citrate condition alongside it as a reference (standard IHC practice). Judge improvement by cytoplasmic staining in appropriate cells, rather than by stronger background alone (UniProt Q13885 localisation; HPA tissue IHC).
Could fixation explain weak TUBB2A staining in paraffin sections?
Target-specific sensitivity to fixation is unknown from the supplied evidence; the selected paraffin-section caption does not state its fixative (caption M06868). Record the actual fixative and fixation duration for each specimen, then compare sections processed together before changing antibody concentration (standard IHC practice). Start the comparison with citrate retrieval at pH 6.0 and the caption’s 1:200 primary dilution, keeping detection conditions constant (datasheet M06868; caption M06868). If a processing difference tracks signal loss, compare matched material with a controlled fixation interval while retaining a positive tissue control (standard IHC practice). Do not infer fixation sensitivity from tissue staining patterns or protein modifications (HPA tissue IHC; UniProt Q13885).
What staining pattern fits TUBB2A in a tissue section?
Expect primarily cytoplasmic staining, consistent with TUBB2A’s cytoskeletal assignment and HPA’s general tissue profile (UniProt Q13885 localisation; HPA tissue IHC). Microtubules are its supported main subcellular location, although fine structures may be hard to resolve with chromogenic tissue IHC (HPA subcellular; standard IHC practice). Evaluate signal within the relevant cell compartment against a secondary-only control, which was included in the catalog image (caption M06868). Strong nuclear-only staining warrants checking counterstain, detection background and antibody specificity before assigning it to TUBB2A (UniProt Q13885 localisation; standard IHC practice). Compare matched sections under the same citrate pH 6.0 retrieval and 1:200 primary conditions before interpreting subtle differences (caption M06868).
How can I assess whether this antibody distinguishes TUBB2A from other beta-tubulins?
Treat an apparent TUBB2A signal as an antibody-specificity question: HPA reports that its tissue antibody targets protein from more than one gene (HPA tissue IHC). The supplied record lists 0 isoforms and no epitope sequence for the catalog antibody, so it does not establish discrimination among beta-tubulin family members (UniProt Q13885 record; caption M06868). TUBB2A has annotated modified residues, but their effects on this antibody’s binding are unknown without epitope information (UniProt Q13885). Compare staining with an independent, characterized reagent or a suitable loss-of-target control where available (standard IHC practice). Report signal as antibody staining until those controls support a TUBB2A-specific assignment (HPA tissue IHC; standard IHC practice).
How should I use IF to investigate an ambiguous IHC pattern?
Use IF/ICC as a separate localisation check, without treating the catalog antibody’s paraffin IHC conditions as an IF protocol (caption M06868; standard IF practice). Multiplex with a marker of the expected cell type, such as an endothelial marker when examining placental endothelial cells, which HPA scores high (HPA tissue IHC). Choose fluorophores and imaging channels after checking the specimen’s autofluorescence and single-stain controls (standard IF practice). Because TUBB2A is cytoplasmic and has no transmembrane segment, plan permeabilisation to expose an intracellular epitope, then optimise it for the specimen and antibody (UniProt Q13885 topology and localisation; standard IF practice). Compare any filamentous signal with HPA’s supported microtubule localisation, while assessing antibody specificity independently (HPA subcellular; HPA tissue IHC).
What should I check when TUBB2A IHC has diffuse background?
First compare the section with a secondary-only control, as used in the catalog’s paraffin-section image, to separate primary-dependent staining from detection background (caption M06868; standard IHC practice). Check blocking, washes, endogenous peroxidase suppression and chromogen development on matched sections before increasing primary antibody concentration (standard IHC practice). The catalog image used primary antibody at 1:200 overnight at 4°C and secondary at 1:200 for 30 minutes at room temperature (caption M06868). Reassess retrieval if background rises after heating, using citrate pH 6.0 as the documented reference condition (datasheet M06868; standard IHC practice). Interpret widespread staining cautiously because HPA flags binding to protein from more than one gene (HPA tissue IHC).
How should I quantify TUBB2A staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the tissue compartment and cell population before scoring, since HPA reports general cytoplasmic expression with cell-dependent intensity (HPA tissue IHC). For cellular staining, record the percentage of positive cells and intensity, or calculate an H-score from consistently defined intensity bins (standard IHC practice). For structures such as cerebral-cortex neuropil, quantify positive area or signal density per mm² of annotated tissue rather than forcing a cell count (HPA tissue IHC; standard IHC practice). Normalise comparisons to the same eligible tissue area or cell population, and keep retrieval, staining batch and image thresholds consistent (standard IHC practice). Include a reference section and background control in each batch, and report scoring rules before comparing groups (standard IHC practice).
When is TUBB2A staining convincing rather than artefactual?
A convincing pattern is predominantly cytoplasmic in an anatomically appropriate cell population, consistent with TUBB2A’s cytoskeletal localisation and HPA’s tissue profile (UniProt Q13885 localisation; HPA tissue IHC). For example, HPA scores placental endothelial cells high and adipocytes as not detected, although those observations do not validate every antibody or specimen (HPA tissue IHC). Inspect nuclear-only signal, section-edge staining and necrotic areas against intact neighbouring tissue and the secondary-only control (caption M06868; standard IHC practice). Check endogenous peroxidase activity when interpreting chromogenic deposits, and avoid counting counterstain or pigment as positive signal (standard IHC practice). Because HPA warns of binding to protein from more than one gene, require independent specificity evidence for a TUBB2A-specific conclusion (HPA tissue IHC).
Boster reagents
Best TUBB2A / Tubulin beta-2A chain IHC Antibodies
M06868 has IHC images from paraffin-embedded human colon, rat kidney and mouse testis (catalog image captions); IF is listed as an application, with no IF image supplied (catalog).
Real IHC data
Anti-beta II tubulin TUBB2A Monoclonal Antibody
Cat # M06868
M06868 has IHC images from paraffin-embedded human colon, rat kidney and mouse testis (catalog image captions). M06868 lists IF among its applications and human, mouse and rat reactivity, but supplies no IF image (catalog).
Which to pick: Choose M06868 for tissue IHC: its own captions document staining of paraffin-embedded human colon, rat kidney and mouse testis at 1:200 (catalog image captions). For IF/ICC, M06868 lists IF and a 1:50 dilution, but supplies no IF image or ICC validation (catalog); its listed human, mouse and rat reactivity makes it the cross-species option (catalog). The IHC captions do not report the fixative (catalog image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q13885 (TBB2A_HUMAN, Tubulin beta-2A chain).
- Human Protein Atlas. TUBB2A tissue IHC expression (reliability: Supported).
- Human Protein Atlas. TUBB2A subcellular location (ICC-IF): Mainly localized to the microtubules. In addition localized to the cytokinetic bridge, mitotic spindle, primary cilium, primary cilium tip, basal body, flagellar centriole, principal piece and end piece..
- Human Protein Atlas. TUBB2A antibody validation summary (3 antibodies).
- TUBB2A expression and its prognostic significance in hepatocellular carcinoma revealed by cholesterol-metabolism-related gene profiling. Frontiers in molecular biosciences 2026 — PMC13033567.
- Differential requirements of tubulin genes in mammalian forebrain development. PLoS genetics 2019 — PMC6697361.
- Knockdown of stem cell regulator Oct4A in ovarian cancer reveals cellular reprogramming associated with key regulators of cytoskeleton-extracellular matrix remodelling. Scientific reports 2017 — PMC5390261.
- Multiple programmed cell death patterns predict the prognosis and drug sensitivity in gastric cancer. Frontiers in immunology 2025 — PMC11832517.
- PubMed PMID:14574404 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:16371510 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TUBB2A (Tubulin beta-2A chain). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, heat-mediated (datasheet M06868). Validated IHC antibody: M06868 at 1:50 recommended; image 1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Placenta (Endothelial cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded Human colon tissue; fixative not specified (datasheet M06868); 2. Fixation: Image fixative and duration unreported (datasheet M06868); 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: Citrate pH 6.0 , 20min (datasheet M06868); 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: Mouse monoclonal (clone Mix) anti-TUBB2A, 1:50 recommended; image 1:200 (datasheet M06868); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific). Selected-image fixative and duration unreported (datasheet M06868); verify before use.; Caveat: Staining may include products from other genes (HPA tissue IHC); Isoform / epitope: No annotated isoforms; one 1–445 chain (UniProt); Regulation: Brain enriched expression (UniProt). Evidence: UniProt Q13885, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the microtubules. In addition localized to the…, Datasheet M06868, PMC13033567, PMC5390261. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TUBB2A/immunohistochemistry-ihc-guide.