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CBX8 / Chromobox protein homolog 8 · IHC design guide
Design Immunohistochemistry for CBX8
Plan CBX8 paraffin-section IHC with its nuclear location (UniProt) and the nuclear and cytoplasmic tissue staining reported by HPA (HPA tissue IHC) in mind. This guide covers fixation, chromogenic detection and compartment-aware scoring, with an IHC antibody range of 0.5–1 μg/ml (datasheet A05234-2).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CBX8 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 | Nuclear and cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Nuclear and cytoplasmic signal in multiple cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A05234-2) | |
| Positive control | Bone marrow+4 more · see all | |
| Negative control | Hippocampus+3 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) | |
| Caveat | Cytoplasmic signal may complicate nuclear scoring (HPA tissue IHC) | |
| Regulation | No expression regulator annotated (UniProt) | |
| Isoform / epitope | No isoforms or processing; no extracellular domain (UniProt) |
Section 1
Recommended CBX8 IHC & IF Protocols
The catalog antibody has an IHC-P protocol (datasheet A05234-2); the published options below describe CBX8 staining in HCC and CRC sections (PMC6361915; PMC4279409).
| Sample | Paraffin-embedded human rectal cancer tissue; fixative not specified (datasheet A05234-2) |
| Fixation | Image fixative and duration unreported (datasheet A05234-2); 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 A05234-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A05234-2) |
| Primary antibody | Rabbit anti-CBX8, 0.5-1μg/ml (datasheet A05234-2) |
| Primary incubation | Overnight at 4 °C (datasheet A05234-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A05234-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CBX8-positive staining in hematopoietic cells of bone marrow (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic and nuclear expression. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 for the catalog antibody (datasheet A05234-2); evaluate nuclear staining (UniProt Q9HC52).
Section 2
What Is the Expected CBX8 Staining Pattern?
CBX8 is a nuclear chromatin regulator without a transmembrane segment (UniProt Q9HC52). In paraffin-section IHC, expect nuclear staining in hematopoietic cells of bone marrow, colon and rectal glandular cells, and ciliated cells of bronchus, fallopian tube and nasopharynx; HPA reports these cell groups as High (HPA tissue IHC). HPA also describes general cytoplasmic and nuclear expression, with Approved IHC reliability and medium consistency between staining and RNA data (HPA tissue IHC).
| Strong nuclear staining in bone-marrow hematopoietic cells or colon glandular cells (HPA: High in both). | This fits the expected cellular signal and CBX8’s nuclear location (UniProt Q9HC52). Assess the named cells individually; an HPA level describes observed staining in a cell population, not every cell in the section (HPA tissue IHC). |
| Predominantly membrane-bound or extracellular deposits, without a convincing nuclear pattern (UniProt Q9HC52: nuclear; no transmembrane segment). | Treat this as suspect localisation and review morphology and controls (general IHC practice). HPA reports some cytoplasmic staining, so cytoplasmic colour alone does not prove an artefact (HPA tissue IHC). |
| Strong signal in hippocampal glial cells or liver cholangiocytes (HPA: Not detected in these cells). | Check cell identification, antibody specificity and endogenous detection activity before assigning CBX8 positivity (general IHC practice). HPA’s negative calls apply to the named cell types, not every cell in those tissues (HPA tissue IHC). |
| Diffuse chromogen across tissue, including spaces outside recognisable cells (general IHC practice). | An indistinct deposit cannot establish CBX8 localisation; assess no-primary and detection controls, blocking and washes (general IHC practice). Compare any retained cellular signal with the nuclear expectation (UniProt Q9HC52). |
| No signal in the expected hematopoietic cells of bone marrow (HPA: High in hematopoietic cells). | First confirm that the relevant cells and an interpretable section are present (general IHC practice). A blank result conflicts with this HPA reference pattern but alone cannot distinguish technical failure from sample variation (HPA tissue IHC). |
💡Expected CBX8 appearanceCall a result positive when strong cellular staining includes nuclei of the named HPA High cell populations and remains morphologically interpretable (HPA tissue IHC; UniProt Q9HC52); isolated membrane or extracellular colour is suspect (UniProt Q9HC52; general IHC practice).
| Compartment reference: CBX8 is nuclear (UniProt Q9HC52); HPA describes general cytoplasmic and nuclear IHC staining (HPA tissue IHC). | Use a nuclear component as the clearest localisation check, while recording cytoplasmic signal separately (UniProt Q9HC52; HPA tissue IHC). Neither source establishes that every positive cell must be exclusively nuclear. |
| Cell-specific reference levels include High in skin melanocytes and testis round or early spermatids, but Not detected in ovarian stromal cells and parathyroid glandular cells (HPA tissue IHC). | Score the specified cells rather than treating a whole organ as uniformly positive or negative (HPA tissue IHC). This distinction matters when adjacent cell populations differ in staining (general IHC practice). |
| IHC evidence is Approved with medium staining-to-RNA consistency; antibody HPA031462 is IHC Approved (HPA tissue IHC; HPA antibodies). | Use the reported pattern as a reference with that stated reliability, not as proof of specificity in every specimen (HPA tissue IHC). Validate an unexpected result with appropriate controls (general IHC practice). |
| CBX8 has one annotated 1–389 chain, no annotated signal peptide or propeptide, and no listed isoforms (UniProt Q9HC52). | These annotations offer no supplied basis for predicting a shed extracellular product or an isoform-specific IHC pattern (UniProt Q9HC52). The supplied record gives no epitope position or target-specific retrieval response. |
| IF/ICC Q&A: Where should CBX8 appear? HPA reports enhanced nucleoplasmic localisation in ICC-IF (HPA subcellular). | Nucleoplasmic signal agrees with the nuclear UniProt annotation (UniProt Q9HC52; HPA subcellular). This is an interpretation cross-check; IF/ICC preparation belongs to its own guide page. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected bone-marrow hematopoietic cells are blank (HPA: High in hematopoietic cells). | The selected field may lack the named cells, or the IHC run may have failed (general IHC practice). HPA does not identify a CBX8-specific fixation or retrieval failure mode. | Confirm morphology, then examine a suitable positive section and run controls (general IHC practice). Review the established IHC workflow if controls fail; do not infer a CBX8 fixation effect from the blank field. |
| Only diffuse or extracellular colour is visible (UniProt Q9HC52: nuclear). | Non-specific chromogen deposition or incomplete washing can obscure cellular localisation (general IHC practice); the supplied sources do not assign this pattern to CBX8 biology. | Review the no-primary control, blocking, washes and detection step (general IHC practice). Score CBX8 only where cell boundaries and the expected nuclear component can be assessed (UniProt Q9HC52). |
| Staining seems universal in a section that includes HPA Not detected cell types (HPA tissue IHC). | Cell identity may be mistaken, or background and endogenous detection activity may be mistaken for antibody signal (general IHC practice). The HPA negative calls concern specified cells. | Recheck morphology in those cells and compare no-primary and detection controls (general IHC practice). Document any reproducible deviation from the HPA cell-specific pattern rather than assigning whole-tissue positivity. |
| Cytoplasmic staining is present but nuclei are faint (HPA: general cytoplasmic and nuclear IHC expression). | The pattern is ambiguous against UniProt’s nuclear annotation; HPA’s cytoplasmic observation means it cannot be dismissed solely on compartment (UniProt Q9HC52; HPA tissue IHC). | Score nuclear and cytoplasmic components separately, compare HPA High cell populations, and check control staining (general IHC practice; HPA tissue IHC). Avoid converting a cytoplasmic-only field into a definitive nuclear result. |
| A presumed negative control stains (HPA tissue IHC: Not detected in selected cell types). | The control may contain other cells with CBX8 staining, or detection background may be present (HPA tissue IHC; general IHC practice). A tissue name alone does not specify its cellular composition. | Inspect the particular HPA Not detected cells, such as liver cholangiocytes, and check the no-primary control (HPA tissue IHC; general IHC practice). Interpret other cells independently. |
| IHC and IF/ICC localisation appear different (HPA: cytoplasmic and nuclear IHC; enhanced nucleoplasmic ICC-IF). | The supplied HPA observations describe different applications; they do not establish why their apparent compartments differ (HPA tissue IHC; HPA subcellular). | Report each application’s observed compartment and compare nuclear signal with UniProt’s annotation (UniProt Q9HC52). Use the separate IF/ICC guide for preparation decisions; avoid inferring an IHC fixation effect. |
Experiment evidence
IHC & IF images showing CBX8 staining
A05234-2 · IHCIHC analysis of Cbx8 using anti-Cbx8 antibody (A05234-2). Cbx8 was detected in paraffin-embedded section of human rectal cancer tissue. Heat
A05234-2 · IHCIHC analysis of Cbx8 using anti-Cbx8 antibody (A05234-2). Cbx8 was detected in paraffin-embedded section of human rectal cancer tissue. Heat
A05234-2 · IFIF analysis of Cbx8 using anti-Cbx8 antibody (A05234-2). Cbx8 was detected in immunocytochemical section of A549 cells. Enzyme antigen retri
M05234 · IHCIHC analysis of Cbx8 using anti-Cbx8 antibody (M05234). Cbx8 was detected in paraffin-embedded section of human gallbladder adenocarcinoma t
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000141570-CBX8/tissue/cerebral+cortex#imid_8036751
HPA · BronchusHuman Protein Atlas IHC, Bronchus. Source: https://www.proteinatlas.org/ENSG00000141570-CBX8/tissue/bronchus#imid_8036732
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000141570-CBX8/tissue/colon#imid_8037386
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000141570-CBX8/tissue/liver#imid_8037379
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA031462; green = CBX8, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA031462; green = CBX8, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CBX8 IHC & IF
🧪Run colon first: glandular cells should show CBX8 staining (HPA: High in colon glandular cells). Use parathyroid gland as the negative tissue (HPA: Not detected in parathyroid gland glandular cells); non-glandular cells on the colon slide are only candidate internal negatives and should be scored as such after their CBX8 status is verified.
Positive control tissue: Bone marrow (Hematopoietic cells, HPA High)
Negative control tissue: Hippocampus (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CBX8 in A-431, U-251MG, U2OS, HeLa BAC 5991, HeLa , with annotated localisation: Nucleoplasm (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, a host- and clonality-matched control immunoglobulin (selected caption: rabbit primary antibody), and CBX8 knockout material processed in parallel as a biological negative. For the biotin–DAB detection used in the selected caption, quench endogenous peroxidase and check for endogenous biotin in the colon section (selected caption: biotinylated secondary and SABC–DAB).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A05234-2 paraffin-section caption does not state a fixative. Heat retrieval in EDTA at pH 8.0 was used in that caption; retrieval dependence and whether frozen sections or IF are easier remain unreported. In colon, distinguish glandular nuclear signal from luminal mucus or other nonspecific chromogen deposits (UniProt: nucleus; HPA: nucleoplasm).
HPA tissue IHC evidence for CBX8
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 |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Fallopian tube | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Nasopharynx | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Parathyroid gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CBX8 IHC Tips
Troubleshoot CBX8 staining in paraffin sections by checking retrieval, nuclear localisation, controls and scoring before interpreting the chromogenic signal.
What retrieval should I start with for CBX8 in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA buffer at pH 8.0 (datasheet A05234-2). That condition was used on a paraffin section of human rectal cancer before incubation with 1 μg/mL antibody overnight at 4°C (datasheet A05234-2). If nuclear staining is weak, compare retrieval duration or a second buffer on adjacent sections while holding antibody concentration, detection and exposure to DAB constant; treat those changes as optimisation, not validated CBX8 conditions. Check that any gain appears in nuclei rather than across the section, because CBX8 is nuclear (UniProt Q9HC52) and nucleoplasmic localisation is supported independently (HPA subcellular).
How should I assess whether fixation is causing weak CBX8 staining?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative (datasheet A05234-2). Record each specimen’s fixative and fixation time, then compare sections processed with the same retrieval, antibody incubation and chromogenic development before attributing weak staining to fixation. For a reference condition, the documented workflow uses EDTA at pH 8.0 and 1 μg/mL antibody overnight at 4°C (datasheet A05234-2). If staining varies, inspect morphology and nuclear counterstaining, and test consistently processed controls; neither the tissue patterns (HPA tissue IHC) nor CBX8’s topology or modifications (UniProt Q9HC52) establish a fixation effect.
Should cytoplasmic CBX8 staining count as a positive IHC result?
Prioritise a nuclear pattern when judging CBX8 staining: the protein is annotated as nuclear (UniProt Q9HC52), with enhanced nucleoplasmic localisation in cell imaging (HPA subcellular). Tissue IHC also reports general cytoplasmic and nuclear expression (HPA tissue IHC), so cytoplasmic colour warrants review rather than automatic exclusion. Compare the signal with a nuclear counterstain, section morphology and a no-primary control; check whether cytoplasmic colour tracks cell boundaries, pigment or diffuse background. Score nuclear and cytoplasmic staining separately, and avoid calling a section positive solely because DAB appears outside nuclei when its control or staining pattern is unconvincing.
How can I evaluate an apparent epitope-dependent staining difference?
The supplied record lists a single 1–389 amino-acid chain and no annotated isoforms (UniProt Q9HC52); it does not identify the catalog antibody’s binding epitope. CBX8 has a chromo domain at residues 11–69 and annotated phosphorylation sites, including Ser110 and Ser130 (UniProt Q9HC52), but those facts do not establish antibody sensitivity to modification. When staining changes, first hold retrieval, antibody concentration and detection constant across adjacent sections, then compare an independently validated antibody if available. Describe any epitope or modification explanation as a hypothesis until mapping or an appropriate specificity experiment supports it.
How can I check CBX8 localisation by multiplex IF?
For the separate IF/ICC workflow, pair CBX8 with a marker for the expected cell type and a nuclear counterstain; its expected nucleoplasmic location is supported by cell imaging (HPA subcellular). Choose well-separated fluorophores, assign a longer-wavelength channel to the weaker target where tissue autofluorescence interferes, and inspect single-label and no-primary controls before interpreting overlap. Because CBX8 is nuclear and has no transmembrane segment (UniProt Q9HC52), permeabilisation must allow antibody access through the cell and nuclear boundaries; optimise it for the specimen and epitope. Do not transfer the paraffin-section EDTA pH 8.0 retrieval or overnight 4°C incubation to IF as validated IF settings (datasheet A05234-2).
What should I check when DAB background obscures CBX8-positive nuclei?
Inspect a no-primary section and the tissue edges first, then compare them with nuclear staining in the test section; CBX8’s expected location is nuclear (UniProt Q9HC52). In chromogenic IHC, check the general workflow’s endogenous peroxidase block, secondary-antibody background and DAB development, especially where blood, pigment or damaged tissue complicates interpretation. The documented section used 10% goat serum blocking, a biotinylated secondary for 30 minutes at 37°C, and a streptavidin–biotin detection system with DAB (datasheet A05234-2). If background persists, compare shorter development or adjusted blocking on matched sections while keeping retrieval at EDTA pH 8.0 initially (datasheet A05234-2).
How should I quantify CBX8 IHC across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, and use the same nuclear counterstain and DAB threshold across the comparison; CBX8 is nuclear (UniProt Q9HC52). Report either the percentage of positive nuclei and an intensity-based H-score, or positive-cell density per mm² when tissue area is the appropriate denominator. Normalise cell-based scores to the number of evaluable cells in the specified compartment, and area-based scores to viable, analysed tissue area; exclude folds and necrosis by a prespecified rule. Keep cytoplasmic scores separate because tissue IHC describes both cytoplasmic and nuclear expression (HPA tissue IHC), and document any threshold set using controls.
How do I distinguish genuine CBX8 staining from artefact?
A credible positive pattern should show interpretable nuclear staining in morphologically intact cells, consistent with nuclear annotation (UniProt Q9HC52) and nucleoplasmic cell imaging (HPA subcellular). Context helps: rectal glandular cells are reported as high, while liver cholangiocytes are reported as not detected (HPA tissue IHC); these patterns are reference observations, not universal controls. Examine edge-heavy colour, necrotic regions, pigment and no-primary sections before assigning weak DAB signal to CBX8, and check the general peroxidase block for endogenous enzyme activity. If a signal is confined to an unexpected compartment or cell population, record it separately and seek an independent specificity check before making a biological claim.
Boster reagents
Best CBX8 / Chromobox protein homolog 8 IHC Antibodies
IHC images show CBX8 staining in human rectal cancer, mouse and rat small intestine (A05234-2 captions), and human gallbladder adenocarcinoma (M05234 caption); IF images show A549 cells and human rectal cancer (A05234-2 captions).
Real IHC data
Anti-Cbx8 Antibody ®
Cat # A05234-2
Real IHC data
Anti-Cbx8 Antibody ® (monoclonal, 8G7)
Cat # M05234
A05234-2 has IHC images from paraffin sections of human rectal cancer and mouse and rat small intestine, plus IF images from A549 cells and human rectal cancer (A05234-2 captions). M05234 has an IHC image from a paraffin section of human gallbladder adenocarcinoma (M05234 caption).
Which to pick: For human tissue IHC, either SKU has a paraffin-section example: A05234-2 in rectal cancer and M05234, a mouse monoclonal clone 8G7, in gallbladder adenocarcinoma (catalog; A05234-2 and M05234 captions). Choose A05234-2 for IF/ICC or mouse and rat IHC because those applications and species are listed for it, with corresponding IF and IHC images; M05234 lists human IHC and no IF/ICC application (catalog; A05234-2 captions). Both IHC captions specify paraffin sections but leave the fixative unreported (A05234-2 and M05234 captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9HC52 (CBX8_HUMAN, Chromobox protein homolog 8).
- Human Protein Atlas. CBX8 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CBX8 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. CBX8 antibody validation summary (1 antibodies).
- The co-expression of CBX8 and PD-L1 and prognostic value in cervical cancer. Medicine 2021 — PMC10545086.
- CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma. Cell death & disease 2019 — PMC6361915.
- Paradoxical role of CBX8 in proliferation and metastasis of colorectal cancer. Oncotarget 2014 — PMC4279409.
- Correlations between chromobox homolog 8 and key factors of epithelial-mesenchymal transition in hepatocellular carcinoma. Cancer cell international 2019 — PMC6916084.
- PubMed PMID:10825164 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CBX8 (Chromobox protein homolog 8). Expected localisation Nuclear and cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A05234-2). Validated IHC antibody: A05234-2 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Bone marrow (Hematopoietic cells); negative Hippocampus. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human rectal cancer tissue; fixative not specified (datasheet A05234-2); 2. Fixation: Image fixative and duration unreported (datasheet A05234-2); 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 A05234-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A05234-2); 8. Primary antibody: Rabbit anti-CBX8, 0.5-1μg/ml (datasheet A05234-2); 9. Primary incubation: Overnight at 4 °C (datasheet A05234-2). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Cytoplasmic signal may complicate nuclear scoring (HPA tissue IHC); Isoform / epitope: No isoforms or processing; no extracellular domain (UniProt); Regulation: No expression regulator annotated (UniProt). Evidence: UniProt Q9HC52, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet A05234-2, PMC6361915, PMC4279409. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CBX8/immunohistochemistry-ihc-guide.