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CCT3 / T-complex protein 1 subunit gamma · IHC design guide
Design Immunohistochemistry for CCT3
Plan CCT3 staining in paraffin sections using the mainly cytoplasmic lymphoid tissue pattern as a reference (HPA tissue IHC). Start with the IHC-validated antibody at 0.5–1 µg/mL (datasheet PB9926) and compare staining across cell types (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CCT3 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 | Mainly cytoplasmic in lymphoid tissues (HPA tissue IHC) | |
| Staining pattern | Lymphoid tissue: mainly cytoplasmic staining (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet PB9926) | |
| 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 conditions consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Staining intensity varies across tissues and cell types (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | 2 isoforms; check epitope coverage across variants (UniProt) |
Section 1
Recommended CCT3 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: PB9926) is accompanied by published CCT3 tissue IHC protocols from PMC13514722 and PMC5958781.
| Sample | Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet PB9926) |
| Fixation | Image fixative and duration unreported (datasheet PB9926); 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, 20 min (datasheet PB9926) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB9926) |
| Primary antibody | Rabbit anti-CCT3, 0.5-1μg/ml (datasheet PB9926) |
| Primary incubation | Overnight at 4 °C (datasheet PB9926) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9926) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CCT3-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Mainly cytoplasmic expression in lymphoid tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet: PB9926); the lung study used 10 mM citrate for 15–20 min (PMC13514722).
Section 2
What Is the Expected CCT3 Staining Pattern?
CCT3 is cytoplasmic and has no transmembrane segment (UniProt P49368). In paraffin sections, expect mainly cytoplasmic staining where tissue expression is observed, including glandular cells in breast and duodenum, glial cells in cerebral cortex, and cervical squamous epithelial cells (HPA: High). HPA rates its tissue IHC evidence Approved, with medium consistency between staining and RNA expression (HPA: tissue IHC).
| Cytoplasmic signal in breast or duodenal glandular cells, glia, or cervical squamous cells. | This fits the reported compartment and cell patterns (UniProt P49368: Cytoplasm; HPA: High in those cells). Compare cells within the same section; HPA's High rating does not require identical intensity in every cell. |
| Nuclear staining dominates while the expected cytoplasmic signal is absent. | That pattern conflicts with the annotated location (UniProt P49368: Cytoplasm). Treat it as questionable; inspect morphology and controls before assigning CCT3 positivity. Nuclear counterstain alone is not target staining (general IHC practice). |
| Strong staining appears chiefly in ovarian stroma or smooth muscle cells. | These are reported as Low, not negative (HPA: ovarian stroma and smooth muscle). Unexpectedly strong signal warrants a specificity check; the tissue profile alone cannot prove cross-reactivity or identify its source. |
| Brown color covers many cell types, extracellular areas, or blank spaces without a cellular pattern. | This does not resemble the mainly cytoplasmic tissue pattern (HPA: tissue IHC). Consider background from detection reagents, endogenous enzyme activity, or nonspecific binding; use controls to distinguish them (general IHC practice). |
| A breast glandular compartment expected to stain shows no detectable signal. | Breast glandular cells are rated High (HPA: tissue IHC), so check that the relevant cells are present. A blank result may reflect staining workflow failure or sample variation; one section does not overturn the HPA pattern. |
💡Expected CCT3 appearanceCall positive a discernible cytoplasmic signal in the expected cells, potentially strong in HPA High examples; nuclear-only, extracellular, or uniform brown color is suspect (UniProt P49368: Cytoplasm; HPA: tissue IHC; general IHC practice).
| Compartment and topology | CCT3 is annotated in the cytoplasm, with no transmembrane segment (UniProt P49368). Judge paraffin IHC by cellular cytoplasmic staining; a crisp surface outline alone is not the expected tissue pattern. |
| Tissue and cell context | HPA rates glandular cells High in adrenal gland, breast, duodenum, endometrium, and epididymis; glia High in caudate and cerebral cortex (HPA: tissue IHC). Low entries remain possible weak positives, not validated negatives. |
| Strength of pattern evidence | Tissue IHC is Approved but has medium consistency with RNA data (HPA: tissue IHC). HPA006543 is IHC Approved; the supplied antibody list gives no IHC status for HPA074177 (HPA: antibodies). Interpret isolated discordant staining cautiously. |
| Isoforms and epitope coverage | UniProt lists 2 isoforms but supplies no epitope or isoform coverage for the antibody (UniProt P49368). Isoform status therefore cannot explain a weak section or justify predicting a different compartment. |
| Antigen retrieval and detection | Retrieval conditions, blocking, and detection can affect paraffin IHC background and visibility (general IHC practice). No source here reports CCT3-specific fixation sensitivity or an optimal retrieval condition; assess workflow controls without attributing a result to CCT3 epitope masking. |
| IF/ICC Q: Should fluorescence match IHC exactly? | HPA approves cytosol as the main ICC-IF location and vesicles and plasma membrane as additional locations (HPA: subcellular). That informs an IF check; the tissue IHC expectation remains mainly cytoplasmic (HPA: tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a breast section's glandular cells. | Those cells are rated High (HPA: tissue IHC); an absent signal may reflect missing target cells, section quality, or an IHC workflow problem (general IHC practice). | Confirm glandular cells on the counterstain and run a known-positive section alongside reagent controls. Review the validated IHC workflow before changing conditions; no CCT3-specific fixation effect is documented here. |
| Nuclei stain strongly but cytoplasm stays clear. | A predominantly nuclear result conflicts with CCT3's annotated cytoplasmic location (UniProt P49368). Counterstain or nonspecific signal may be mistaken for chromogen (general IHC practice). | Distinguish counterstain color from chromogen, inspect a reagent control, and repeat the comparison in an HPA High cell compartment. Do not score nuclear-only color as expected CCT3 staining. |
| Uniform brown haze obscures cell borders. | Diffuse color lacks the mainly cytoplasmic distribution reported for CCT3 (HPA: tissue IHC). Endogenous detection activity or nonspecific binding can produce such background (general IHC practice). | Check a no-primary control and the blocking and detection steps. Judge a repeated stain only where cell boundaries and cytoplasm remain readable (general IHC practice). |
| Ovarian stroma or smooth muscle looks stronger than a chosen positive compartment. | Both cell types are listed Low (HPA: tissue IHC). The mismatch raises a specificity question, although HPA levels do not define a guaranteed intensity ratio between individual sections. | Compare sections processed together and inspect no-primary controls. Reassess cell identity and morphology before calling the unexpected staining CCT3-specific (general IHC practice). |
| Stain is strongest only at tissue edges or in damaged areas. | An edge-dominant pattern is not the reported cellular pattern (HPA: tissue IHC). Uneven reagent exposure or section damage can complicate interpretation (general IHC practice). | Score intact interior tissue with identifiable cells and compare the control section. Repeat staining if an intact cytoplasmic compartment cannot be assessed (general IHC practice). |
| A Low tissue is blank and is being used as a negative control. | Low is a reported staining level, not absence of CCT3 (HPA: esophageal and vaginal squamous cells, ovarian stroma, smooth muscle, chondrocytes). HPA lists no negative tissue entries in this payload. | Use reagent controls to assess background and a High cell compartment to assess detection. Do not label a blank Low sample a validated biological negative (HPA: tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing CCT3 staining
PB9926 · IHCIHC analysis of CCT3 using anti-CCT3 antibody (PB9926). CCT3 was detected in paraffin-embedded section of human lung cancer tissue. Heat med
PB9926 · IHCIHC analysis of CCT3 using anti-CCT3 antibody (PB9926). CCT3 was detected in paraffin-embedded section of human mammary cancer tissue. Heat
PB9926 · IFIF analysis of CCT3 using anti-CCT3 antibody (PB9926). CCT3 was detected in immunocytochemical section of A431 cell. Enzyme antigen retrieva
M05920 · IFIF analysis of CCT3 using anti-CCT3 antibody (M05920). CCT3 was detected in immunocytochemical section of A431 cells. Enzyme antigen retriev
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000163468-CCT3/tissue/cerebral+cortex#imid_2171204
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000163468-CCT3/tissue/colon#imid_2171122
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000163468-CCT3/tissue/liver#imid_2171131
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000163468-CCT3/tissue/kidney#imid_2171140
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006543; green = CCT3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA006543; green = CCT3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CCT3 IHC & IF
🧪Run cerebral cortex first and expect staining in glial cells (HPA: High in cerebral cortex glial cells). HPA detects CCT3 in all 45 scored tissues, so no negative tissue is established; no-primary and isotype controls carry the negative comparison, and apparently unstained cells on the positive slide should be treated as background comparators rather than validated CCT3-negative cells (HPA: no negative tissue rows; standard IHC practice).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: CCT3 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 CCT3 in A-431, U-251MG, U2OS, K-562, OE19, with annotated localisation: Cytosol (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a host- and clonality-matched isotype control; for PB9926, the host is rabbit (caption: rabbit anti-CCT3). Use a CCT3 knockout preparation as a biological negative, or cognate peptide competition if the peptide is available; quench endogenous peroxidase for chromogenic detection and assess cortical autofluorescence if performing IF (standard IHC/IF practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected paraffin-section caption leaves the fixative unreported (caption: fixative not stated). The caption uses citrate buffer at pH 6 for 20 minutes, but does not establish that retrieval is required (caption: heat-mediated antigen retrieval). HPA provides ICC-IF images but no basis to conclude frozen sections or IF are easier than paraffin IHC; cortical lipofuscin can complicate IF background assessment (HPA: ICC-IF images; standard IF practice).
HPA tissue IHC evidence for CCT3
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 → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Caudate | Glial cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Glial cells | High | Protein (IHC) | HPA → |
| Cervix | Squamous epithelial cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: CCT3 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced CCT3 IHC Tips
Troubleshoot chromogenic CCT3 staining in paraffin sections by checking retrieval, compartment, tissue context, and controls before comparing staining scores.
Which retrieval condition should I start with for CCT3 in paraffin sections?
Start with heat-mediated antigen retrieval in citrate buffer at pH 6 (datasheet PB9926). The selected tissue-IHC image used 20 minutes of retrieval in that buffer, followed by 10% goat serum blocking and overnight primary-antibody incubation at 4°C (PB9926 tissue-IHC caption). If cytoplasmic staining is weak, first check section adhesion, heating consistency, and whether the positive control stains under the same run conditions (standard IHC practice). Only then compare a carefully controlled alternative retrieval condition, recording its effect on both specific staining and tissue morphology (standard IHC practice).
Could fixation explain weak or uneven CCT3 staining?
CCT3-specific sensitivity to fixation is unknown from the supplied evidence; the selected paraffin-section caption does not state a fixative (PB9926 tissue-IHC caption). Record the actual fixative, fixation duration, and processing history for each specimen before comparing staining across cases (standard IHC practice). Uneven preservation can alter antigen accessibility or morphology, so inspect the affected area against a well-preserved area and a processed positive control (standard IHC practice). If results differ between batches, hold retrieval, antibody concentration, and detection conditions constant while comparing specimens with documented processing (standard IHC practice). Do not infer fixation tolerance from CCT3 localisation or modification annotations (UniProt P49368).
Where should convincing CCT3 staining appear?
Expect predominantly cytoplasmic staining when evaluating CCT3 in paraffin sections (UniProt P49368 subcellular location; HPA tissue IHC profile). HPA reports mainly cytoplasmic expression in lymphoid tissues, while its subcellular IF assessment identifies the cytosol as the main location and vesicles and plasma membrane as additional locations (HPA tissue IHC; HPA subcellular). CCT3 has no annotated transmembrane segment, so an exclusively crisp membrane outline deserves scrutiny before it is called specific (UniProt P49368 topology). Compare the pattern with cell morphology and a known positive section, and assess diffuse nuclear or stromal staining against control slides (standard IHC practice).
Can the CCT3 antibody distinguish isoforms or modification-dependent epitopes?
Two CCT3 isoforms are listed, but the supplied evidence does not map the catalog antibody’s epitope or establish isoform selectivity (UniProt P49368 isoforms; PB9926 tissue-IHC caption). CCT3 also has annotated modified residues, including phosphoserines at 11, 170, and 243 and acetyllysine at 222 (UniProt P49368 modified residues). Those annotations alone cannot explain a staining difference or show that retrieval exposes a particular epitope (UniProt P49368; standard IHC interpretation). For an isoform-specific claim, obtain epitope information and validate discrimination with appropriate reference material before assigning compartment or intensity differences to an isoform (standard IHC practice).
How should I check CCT3 localisation by IF alongside this IHC result?
Use IF as a separate localisation check: HPA reports mainly cytosolic CCT3, with additional vesicle and plasma-membrane localisation (HPA subcellular). Multiplex CCT3 with a validated marker defining the cell population of interest, then compare signals within the same cells rather than relying on adjacent-cell staining (standard IF practice). Select a fluorophore channel with low tissue autofluorescence, assess an unstained control, and keep exposure settings consistent when comparing samples (standard IF practice). Because CCT3 is primarily cytosolic and has no annotated transmembrane segment, include permeabilisation when testing access to a cytosolic epitope; the antibody’s epitope is not specified here (HPA subcellular; UniProt P49368 topology; standard IF practice).
How can I reduce diffuse brown background without losing CCT3 signal?
First distinguish background from the expected predominantly cytoplasmic pattern using a positive section and a no-primary control in the same staining run (UniProt P49368 subcellular location; standard IHC practice). The selected image used 10% goat serum, 1 µg/mL primary antibody overnight at 4°C, and biotin-based detection with DAB (PB9926 tissue-IHC caption). Check that serum blocking, washes, and chromogen development are consistent, then titrate antibody concentration if the no-primary control is clean but tissue staining remains diffuse (standard IHC practice). For peroxidase detection, check the peroxidase block and endogenous biotin controls as appropriate to the detection system (standard IHC practice).
How should I score CCT3 IHC across heterogeneous sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and cytoplasmic scoring rule before reviewing case labels, because expected CCT3 localisation is predominantly cytoplasmic (UniProt P49368 subcellular location; standard IHC practice). Report the percentage of positive target cells and staining intensity, or calculate an H-score from intensity categories 0–3 for a possible range of 0–300 (standard IHC practice). If cell abundance varies, normalise positive-cell counts to the number of evaluable target cells, or report positive-cell density per mm² of viable tissue (standard IHC practice). Apply one threshold, exposure-independent scoring method, and exclusion rule for folds, necrosis, and section edges across all slides (standard IHC practice).
When is strong CCT3 staining likely to be artefactual?
Give greatest weight to staining in intact cells with the expected predominantly cytoplasmic distribution (UniProt P49368 subcellular location; HPA tissue IHC profile). HPA reports high staining in several listed cell populations and low staining in others, so identify the stained cell type before attributing a whole-section signal to the cells under study (HPA tissue IHC). Be cautious with isolated nuclear staining, sharp edge accentuation, necrotic deposits, or signal present in a no-primary control; examine morphology and repeat with matched controls (standard IHC practice). Brown signal driven by endogenous peroxidase or the detection system should be resolved through the relevant controls before it is interpreted as CCT3 (standard IHC practice).
Boster reagents
Best CCT3 / T-complex protein 1 subunit gamma IHC Antibodies
The catalog includes CCT3 staining in human paraffin sections and A431 cells (PB9926 and M05920 image captions); PB9926 also lists mouse and rat reactivity (catalog: PB9926 reactivity).
Real IHC data
Anti-CCT3 Antibody ®
Cat # PB9926
Real IF data
Anti-CCT3 Antibody ® (monoclonal, 12H4)
Cat # M05920
PB9926 has IHC images from human lung, mammary and intestinal cancer, and placenta paraffin sections, plus IF images from A431 cells (PB9926 image captions). M05920 has IF images from A431 cells and lists human reactivity (M05920 image captions; catalog: M05920 reactivity).
Which to pick: Choose PB9926 for tissue IHC: its rabbit antibody has a listed IHC application and 0.5–1 μg/ml IHC dilution, with paraffin section images; the fixative is unreported (catalog: PB9926 applications and dilution; PB9926 IHC captions). For IF/ICC, PB9926 has A431 images and a listed 2 μg/ml IF dilution, while mouse monoclonal M05920 (clone 12H4) has A431 IF images and a captioned 5 μg/ml concentration (PB9926 and M05920 IF captions; catalog: M05920 clone). For mouse or rat samples, PB9926 lists reactivity with both species, although its pictured IHC sections are human; M05920 lists human reactivity only (catalog: PB9926 and M05920 reactivity; PB9926 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P49368 (TCPG_HUMAN, T-complex protein 1 subunit gamma).
- Human Protein Atlas. CCT3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CCT3 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the plasma membrane and vesicles..
- Human Protein Atlas. CCT3 antibody validation summary (2 antibodies).
- Expression patterns and clinical implications of chaperonin subunit 3 mRNA and protein in laryngeal squamous cell carcinoma. World journal of clinical oncology 2025 — PMC12754133.
- CCT3 Enhances Lung Adenocarcinoma Progression by Promoting p53 Ubiquitination and Degradation. Medicina (Kaunas, Lithuania) 2026 — PMC13514722.
- Insights into the roles and driving forces of CCT3 in human tumors. Frontiers in pharmacology 2022 — PMC9596777.
- Suppression of CCT3 inhibits malignant proliferation of human papillary thyroid carcinoma cell. Oncology letters 2018 — PMC5958781.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:17974005 — UniProt-cited evidence.
- PubMed PMID:16710414 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CCT3 (T-complex protein 1 subunit gamma). Expected localisation Mainly cytoplasmic in lymphoid tissues (HPA tissue IHC). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet PB9926). Validated IHC antibody: PB9926 at 0.5-1μg/ml (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-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet PB9926); 2. Fixation: Image fixative and duration unreported (datasheet PB9926); 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 , 20 min (datasheet PB9926); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB9926); 8. Primary antibody: Rabbit anti-CCT3, 0.5-1μg/ml (datasheet PB9926); 9. Primary incubation: Overnight at 4 °C (datasheet PB9926). Watch for: Fixation: Keep fixation conditions consistent across sections (standard IHC practice; not target-specific); Caveat: Staining intensity varies across tissues and cell types (HPA tissue IHC); Isoform / epitope: 2 isoforms; check epitope coverage across variants (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt P49368, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the cytosol. In addition localized to the plasma…, Datasheet PB9926, PMC13514722, PMC5958781. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CCT3/immunohistochemistry-ihc-guide.