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TBCA / Tubulin-specific chaperone A · IHC design guide
Design Immunohistochemistry for TBCA
Plan TBCA staining in paraffin sections using its observed cytoplasmic and nuclear tissue pattern (HPA tissue IHC). This guide covers fixation, antibody dilution, detection and interpretation for chromogenic IHC.
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
TBCA 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 tissue staining (HPA tissue IHC) | |
| Staining pattern | General cytoplasmic and nuclear staining, including glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+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 | Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining has medium consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Low tissue specificity; no regulator reported (HPA tissue IHC; UniProt) | |
| Isoform / epitope | 2 isoforms; epitope differences are unreported (UniProt) |
Section 1
Recommended TBCA IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet) with published chromogenic TBCA IHC methods for zebra finch brain (PMC3754799; PMC4422980).
| Sample | Paraffin-embedded rat brain tissue; fixative not specified (datasheet A06926) |
| Fixation | Image fixative and duration unreported (datasheet A06926); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-TBCA, 1:50-1:200 (datasheet A06926) |
| 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 | TBCA-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic and nuclear expression. No signal in the no-primary control. |
💡Decision noteFor paraffin sections, start with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page retrieval); the published methods used thaw-mounted sections (PMC3754799; PMC4422980).
Section 2
What Is the Expected TBCA Staining Pattern?
TBCA is a cytoplasmic, cytoskeleton-associated tubulin-folding protein with no transmembrane segment (UniProt O75347). In paraffin-section IHC, expect staining in glandular, epithelial and muscle cells reported by HPA; HPA also describes general cytoplasmic and nuclear expression (HPA tissue IHC). Treat that pattern as a guide rather than proof of specificity: HPA rates the tissue staining Approved, with medium consistency against RNA data (HPA tissue IHC).
| Cytoplasmic staining in adrenal or parathyroid glandular cells, or pancreatic exocrine cells. | These are reported High-staining populations (HPA tissue IHC). Compare the intended cells with adjacent tissue and controls before scoring: a brown signal in the expected cells supports the pattern, but staining alone does not establish antibody specificity (HPA tissue IHC; standard IHC practice). |
| Cytoplasmic staining with some nuclear staining in an expected cell population. | HPA describes general cytoplasmic and nuclear tissue expression, while UniProt assigns TBCA to the cytoplasm and cytoskeleton (HPA tissue IHC; UniProt O75347). Record the compartments separately; nuclear staining can fit the tissue profile, but it should not replace assessment of the cytoplasmic signal. |
| Predominantly cell-surface, extracellular or tissue-edge staining. | A surface or extracellular pattern is discordant with the annotated cytoplasm and cytoskeleton and the absence of a transmembrane segment (UniProt O75347). Check morphology, section edges and negative controls for nonspecific signal before interpreting it as TBCA (standard IHC practice). |
| Strong staining in cells outside the reported HPA populations, without a convincing pattern in expected cells. | Consider cross-reactivity or endogenous chromogen activity (standard IHC practice). HPA reports low tissue specificity and provides selected examples, not an exhaustive list of positive or negative cell types (HPA tissue IHC); cell identity alone cannot establish that the new signal is false. |
| Diffuse brown haze across cells and extracellular areas, or no staining in a reported high-staining tissue. | Diffuse haze weakens compartment and cell-type scoring; absent signal in a reported positive calls the run into question (standard IHC practice; HPA tissue IHC). Interpret either finding with the negative and positive controls before assigning a biological difference. |
💡Expected TBCA appearanceA credible positive is chiefly cytoplasmic staining in a reported glandular, epithelial or cardiomyocyte population, potentially High in the listed HPA examples; isolated surface or extracellular colour is suspect (HPA tissue IHC; UniProt O75347).
| Cell population and tissue choice | HPA reports High staining in adrenal, appendix, gallbladder and parathyroid glandular cells, pancreatic exocrine cells, nasopharyngeal respiratory epithelium and cardiomyocytes; endometrial stromal cells are Medium (HPA tissue IHC). Use a listed high-staining population as a comparison when judging a weak test section. |
| Compartment interpretation | UniProt places TBCA in the cytoplasm and cytoskeleton; HPA tissue IHC also reports nuclear expression (UniProt O75347; HPA tissue IHC). Document both compartments when present. Their differing emphasis limits how confidently one can reject a nuclear component from tissue IHC alone. |
| Validation and low-signal reference | HPA rates antibody HPA036487 Approved for IHC and calls the tissue profile medium in RNA–staining consistency (HPA antibodies; HPA tissue IHC). Hippocampal glial cells are listed as Low, not negative; weak glial staining is therefore a poor absolute negative control (HPA tissue IHC). |
| Isoforms and processing | UniProt lists 2 isoforms, no signal peptide or propeptide, and a chain spanning residues 2–108 (UniProt O75347). The supplied record gives no IHC epitope position, so it does not show whether the IHC-validated antibody detects both isoforms equally (UniProt O75347; HPA antibodies). |
| IF/ICC Q&A: What localisation should I expect? | Mainly microtubules (supported), with additional nucleolar localisation marked uncertain (HPA ICC-IF). This is an IF/ICC observation, not a requirement that chromogenic tissue IHC resolve microtubules or nucleoli. HPA lists HPA036487 as Supported for ICC and Approved for IHC (HPA antibodies). |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a listed high-staining population. | The staining run may have failed, or signal may be below detection; HPA's High designation is a tissue observation, not a guarantee for every section (HPA tissue IHC; standard IHC practice). | Check that the positive control stained and the detection reagents worked; then review the catalog antibody's IHC-P instructions for the chosen retrieval and dilution (standard IHC practice). |
| Weak signal in the test tissue, with a working positive control. | Expression may be lower in the sampled cell population: HPA reports Low in hippocampal glia and Medium in endometrial stromal cells (HPA tissue IHC). | Identify and score the relevant cells before adjusting staining conditions; compare with a listed high-staining population in the same run (HPA tissue IHC; standard IHC practice). |
| Nuclear staining dominates while cytoplasm is faint. | HPA reports nuclear as well as cytoplasmic tissue expression, so nuclear staining alone does not diagnose an artefact (HPA tissue IHC). UniProt's localisation remains cytoplasm and cytoskeleton (UniProt O75347). | Score nuclear and cytoplasmic signal separately, review cell morphology, and compare both compartments with negative controls (standard IHC practice). |
| Brown signal appears in many unrelated cells or outside cells. | Nonspecific binding or endogenous detection activity can obscure the expected cellular pattern (standard IHC practice). Such distribution needs control-based assessment rather than a TBCA call. | Inspect a no-primary control and review blocking, washing and endogenous enzyme suppression appropriate to the chromogenic system (standard IHC practice). |
| Staining is concentrated at tissue edges or on damaged areas. | Uneven reagent exposure or damaged-section background can create a spatial pattern unrelated to the expected cell distribution (standard IHC practice). | Exclude visibly damaged areas from scoring and compare intact regions with the positive and negative controls (standard IHC practice). |
| An IF/ICC image shows nucleolar signal, but tissue IHC does not. | HPA labels nucleoli an uncertain additional IF/ICC location; its supported main IF/ICC location is microtubules (HPA ICC-IF). | Do not require nucleolar staining for a positive IHC call. Judge the tissue section by its cell populations, cytoplasmic pattern and run controls (HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing TBCA staining
A06926 · IHCImmunohistochemistry of paraffin-embedded rat brain using TBCA antibody at dilution of 1:100 .Perform microwave antigen retrieval with 10 mM
A06926 · IHCImmunohistochemistry of paraffin-embedded mouse spinal cord using TBCA antibody at dilution of 1:100 .Perform microwave antigen retrieval wi
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000171530-TBCA/tissue/cerebral+cortex#imid_9260072
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000171530-TBCA/tissue/colon#imid_9259286
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000171530-TBCA/tissue/liver#imid_9259287
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000171530-TBCA/tissue/kidney#imid_9259272
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036487; green = TBCA, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036487; green = TBCA, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for TBCA IHC & IF
🧪Run appendix first and score its glandular cells, which show High TBCA staining (HPA: High in appendix glandular cells). HPA detects TBCA in all 45 scored tissues, so use no-primary and isotype controls for the negative comparison; cells on the positive slide that lack visible signal should show counterstain only, but no cell population there is established as TBCA-negative (HPA: no negative tissue rows; standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: TBCA 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 TBCA in A-431, U-251MG, U2OS, with annotated localisation: Microtubules (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide, a host- and clonality-matched isotype control, and a biological specificity control using TBCA knockout material or peptide blocking when the immunogen is available (standard IHC practice). Check endogenous peroxidase activity in appendix tissue before chromogenic detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A06926 rat-brain paraffin-section caption does not state a fixative (selected tissue-IHC caption). That caption uses microwave retrieval in 10 mM PBS at pH 7.2 and a 1:100 antibody dilution, but it does not establish that retrieval is required for appendix tissue (selected tissue-IHC caption). The supplied evidence does not establish whether frozen-section IHC or IF is easier for TBCA; for appendix, inspect glandular lumens and tissue edges for nonspecific chromogenic deposit before scoring cells (standard IHC practice).
HPA tissue IHC evidence for TBCA
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Endometrium | Cells in endometrial stroma | Medium | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: TBCA is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced TBCA IHC Tips
Troubleshoot TBCA staining in paraffin sections by checking retrieval, localisation and controls before comparing signal intensity across samples.
Which retrieval conditions should I try first for TBCA paraffin IHC?
For TBCA paraffin IHC, start with citrate buffer at pH 6.0, 95–98 °C for 20 min (page IHC retrieval setting). If staining is weak, compare it on matched sections with microwave retrieval in 10 mM PBS at pH 7.2, the condition shown for catalog antibody A06926 in paraffin-embedded rat brain (A06926 tissue-IHC caption). Keep the antibody dilution and detection conditions constant during that comparison; the caption used 1:100 (A06926 tissue-IHC caption). Score signal within cells alongside tissue damage and diffuse background, since excessive heat can reduce interpretable morphology (standard IHC practice).
How should I investigate weak TBCA staining after fixation?
TBCA-specific fixation sensitivity is unknown from the supplied evidence; the A06926 paraffin-section caption does not state a fixative (A06926 tissue-IHC caption). Record the fixative and fixation duration for each specimen, then compare sections processed with the same retrieval, antibody dilution and detection conditions (standard IHC practice). Include a section from the same processing batch that previously stained well, if available, to distinguish a batch-wide failure from a specimen issue (standard IHC practice). Examine morphology and staining across the section before changing retrieval, because damaged or unevenly processed tissue can make intensity comparisons unreliable (standard IHC practice).
Where should TBCA staining appear, and how should nuclear signal be assessed?
Assess cytoplasmic staining first: TBCA is annotated in the cytoplasm and cytoskeleton, and supported cell-imaging evidence places it mainly at microtubules (UniProt O75347 subcellular; HPA subcellular). HPA tissue IHC also reports a general cytoplasmic and nuclear pattern, so nuclear staining alone does not resolve whether a section is specific (HPA tissue IHC). Nucleolar localisation has uncertain support and should be interpreted cautiously in chromogenic sections (HPA subcellular). Compare nuclear and cytoplasmic patterns within intact cells, review a no-primary control, and check whether apparent nuclear signal follows tissue folds or section edges (standard IHC practice).
Could isoforms or epitope accessibility explain differing TBCA staining patterns?
TBCA has 2 annotated isoforms, while the supplied evidence does not identify which epitope the catalog antibody recognises (UniProt O75347 isoforms; supplied A06926 caption). Do not assign a staining difference to one isoform without antibody epitope information or an independent isoform-specific assay (standard IHC interpretation). The annotated mature chain spans residues 2–108, with N-terminal acetylalanine at residue 2, but the effect of that modification on this antibody is unknown (UniProt O75347 processing; modified residues). Compare retrieval conditions on adjacent sections and keep detection constant before interpreting a changed pattern as biological (standard IHC practice).
How can IF help check the TBCA pattern seen by chromogenic IHC?
Use IF as a separate localisation check, since HPA cell imaging places TBCA mainly at microtubules and reports nucleolar localisation with uncertain support (HPA subcellular). Multiplex TBCA with a validated marker for the expected cell type in the sample, and choose fluorophores after inspecting that tissue’s autofluorescence in unstained and no-primary controls (standard IF practice). TBCA has no annotated transmembrane segment and is assigned to the cytoplasm and cytoskeleton, so permeabilise fixed cells to access intracellular epitopes, then optimise its strength empirically (UniProt O75347 topology; subcellular; standard IF practice). Keep the IF result distinct from the A06926 paraffin-IHC image when judging antibody performance (A06926 tissue-IHC caption).
What should I check when TBCA IHC shows widespread brown background?
First compare the stained section with a no-primary control and inspect whether brown signal follows cells or instead accumulates at edges, folds and damaged regions (standard IHC practice). Apply a peroxidase block before chromogenic detection and check for residual endogenous enzyme activity; this is a general IHC control, not TBCA-specific evidence (standard IHC practice). Reassess protein blocking, washing, antibody concentration and DAB development time one variable at a time (standard IHC practice). Widespread cellular signal is plausible because HPA reports low tissue specificity for TBCA RNA, but that observation does not validate every brown deposit as TBCA (HPA tissue IHC).
How should I quantify TBCA staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because HPA tissue IHC describes both cytoplasmic and nuclear expression (HPA tissue IHC). For intact cells, report an H-score or the percentage of positive cells with intensity categories; record cytoplasmic and nuclear scores separately when both are assessed (standard IHC practice). For spatial counts, report positive cells per mm² of viable tissue and exclude folds, necrosis and section edges by a prespecified rule (standard IHC practice). Normalise comparisons to the same eligible cell population or viable tissue area, with matched staining and imaging conditions across sections (standard IHC practice).
How can I distinguish credible TBCA staining from artefact?
A credible result should show cellular staining consistent with TBCA’s cytoplasmic and cytoskeletal annotation, while considering the nuclear expression reported in HPA tissue IHC (UniProt O75347 subcellular; HPA tissue IHC). Supported microtubule localisation strengthens a structured cytoplasmic pattern, whereas nucleolar staining alone has uncertain support (HPA subcellular). Check the identity of stained cells against the sampled tissue: HPA reports high staining in several listed glandular cell populations, but its profile does not establish a universal positive cell type (HPA tissue IHC). Treat signal concentrated at edges or necrosis, or reproduced in a no-primary control, as possible processing or endogenous enzyme artefact (standard IHC practice).
Boster reagents
Best TBCA / Tubulin-specific chaperone A IHC Antibodies
A06926 has IHC images of paraffin-embedded rat brain and mouse spinal cord (A06926 image captions); human, mouse, and rat reactivity is listed (catalog: A06926 reactivity).
Real IHC data
Anti-Tubulin-specific chaperone A TBCA Antibody
Cat # A06926
A06926 is the sole SKU and is listed for IHC (catalog: A06926 applications). Its images show paraffin-embedded rat brain and mouse spinal cord stained at 1:100 (A06926 image captions).
Which to pick: For paraffin-section tissue IHC, choose A06926; its images use microwave antigen retrieval in 10 mM PBS, pH 7.2, and the fixative is unreported (A06926 image captions). No IF/ICC option is documented: A06926 does not list IF/ICC or show IF images (catalog: A06926 applications and image alts). For work across species, A06926 is a rabbit polyclonal listed as reactive with human, mouse, and rat, with IHC images for rat and mouse (catalog: A06926 host, dilution_raw, reactivity, and image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O75347 (TBCA_HUMAN, Tubulin-specific chaperone A).
- Human Protein Atlas. TBCA tissue IHC expression (reliability: Approved).
- Human Protein Atlas. TBCA subcellular location (ICC-IF): Mainly localized to the microtubules. In addition localized to the nucleoli..
- Human Protein Atlas. TBCA antibody validation summary (1 antibodies).
- Sexually dimorphic and developmentally regulated expression of tubulin-specific chaperone protein A in the LMAN of zebra finches. Neuroscience 2013 — PMC3754799.
- Masculinisation of the zebra finch song system: roles of oestradiol and the Z-chromosome gene tubulin-specific chaperone protein A. Journal of neuroendocrinology 2015 — PMC4422980.
- Identifying space-resolved proteins of the murine thymus, by combining MALDI-MSI and proteomics. Life science alliance 2026 — PMC12634821.
- PubMed PMID:9653160 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15372022 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for TBCA (Tubulin-specific chaperone A). Expected localisation Cytoplasmic and nuclear tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A06926 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded rat brain tissue; fixative not specified (datasheet A06926); 2. Fixation: Image fixative and duration unreported (datasheet A06926); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-TBCA, 1:50-1:200 (datasheet A06926); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has medium consistency with RNA expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope differences are unreported (UniProt); Regulation: Low tissue specificity; no regulator reported (HPA tissue IHC; UniProt). Evidence: UniProt O75347, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the microtubules. In addition localized to the…, Datasheet A06926, PMC3754799, PMC4422980. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/TBCA/immunohistochemistry-ihc-guide.