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POLR3G / DNA-directed RNA polymerase III subunit RPC7 · IHC design guide
Design Immunohistochemistry for POLR3G
Plan POLR3G chromogenic IHC in paraffin sections using the observed nuclear tissue pattern as a guide (HPA tissue IHC). Compare testis and squamous epithelia with an appropriate negative control, and interpret weak staining cautiously because tissue staining awaits external verification (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
POLR3G 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 staining observed in tissue (HPA tissue IHC) | |
| Staining pattern | Nuclei in seminiferous ducts and squamous epithelia (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Skin+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A10669) | |
| Caveat | Tissue staining awaits external verification (HPA tissue IHC) | |
| Regulation | Expressed in dividing cells (UniProt) | |
| Isoform / epitope | No annotated isoforms or processing changes (UniProt) |
Section 1
Recommended POLR3G IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet) with one published POLR3G IHC protocol in bladder cancer tissue (PMC13092746).
| Sample | Paraffin-embedded Mouse Brain tissue; fixative not specified (datasheet A10669) |
| Fixation | Image fixative and duration unreported (datasheet A10669); 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 Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear 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-POLR3G, 1:100-1:300 (datasheet A10669) |
| 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 | POLR3G-positive staining in keratinocytes of skin (HPA tissue IHC: Medium). HPA tissue profile: Nuclear expression in several different tissue types, most abundant in seminiferous ducts of testis and squamous epithelia. No signal in the no-primary control. |
💡Decision noteStart with Tris-EDTA pH 9.0 retrieval at 95–98 °C for 20 min (page retrieval rule); the published alternative uses citrate in an autoclave (PMC13092746).
Section 2
What Is the Expected POLR3G Staining Pattern?
POLR3G should stain mainly in nuclei, with nucleolar exclusion (UniProt O15318; HPA: nuclear tissue expression). Expect the clearest tissue signal in testis preleptotene spermatocytes and several squamous epithelia (HPA: Medium). POLR3G has no transmembrane segment (UniProt O15318 topology). HPA rates the tissue IHC profile Enhanced, while describing medium agreement with RNA data and pending external verification (HPA: tissue reliability).
| Nuclear staining in testis preleptotene spermatocytes or squamous epithelial cells; nucleoli remain unstained. | This fits the reported tissue distribution and nuclear pattern (HPA: Medium in these cells; UniProt O15318: excluded from nucleoli). Judge the cells and compartment together: nuclear color elsewhere alone is less informative because HPA reports nuclear expression across several tissue types (HPA: tissue profile). |
| Prominent nucleolar staining, or predominantly cytoplasmic staining in the listed adult positive tissues. | This conflicts with the expected dominant nuclear pattern and nucleolar exclusion (HPA: tissue profile; UniProt O15318). Check morphology, counterstain and detection controls before calling it POLR3G. Cytoplasmic localisation is recorded by UniProt, so its presence alone is not proof of an artefact (UniProt O15318: cytoplasm). |
| Strong staining in HPA-listed negative cells, such as adipocytes or adrenal glandular cells. | HPA reports POLR3G as Not detected in those cells (HPA: adipose tissue and adrenal gland IHC). Consider cross-reactivity or endogenous detection activity, especially if the positive cells lack the expected nuclear pattern (general IHC practice). A single discrepant field does not identify the cause. |
| Diffuse color over tissue and empty spaces obscures the nuclei. | That distribution cannot be scored as a specific nuclear result (HPA: nuclear tissue profile). Review background in the no-primary and detection controls, then assess blocking, washing and detection conditions (general IHC practice). Interpret any remaining cell signal only after the background is sufficiently low. |
| No nuclear signal in an interpretable testis section containing preleptotene spermatocytes. | HPA reports Medium staining in these cells, making this a useful check of the run (HPA: testis IHC). Confirm the cells are present and the section and detection controls worked (general IHC practice). Absence in one run cannot by itself establish that POLR3G is absent from the sample. |
💡Expected POLR3G appearanceCall a positive result when the relevant cells show discernible nuclear staining, with nucleoli spared and about Medium signal in HPA-listed positive populations (HPA: tissue IHC; UniProt O15318); isolated strong staining in HPA-listed negative cells or diffuse background is suspect (HPA: negative cells; general IHC practice).
| Cell population and tissue | HPA reports Medium staining in testis preleptotene spermatocytes, keratinocytes, squamous epithelial cells, ovarian follicle cells and splenic red-pulp cells (HPA: tissue IHC). Its Not detected calls apply to the named cells, not every cell in those organs (HPA: tissue IHC). |
| Compartment and nucleoli | The tissue profile is nuclear, while UniProt records both nucleus and cytoplasm and specifically excludes nucleoli (HPA: tissue IHC; UniProt O15318). Score the nuclear pattern first; interpret cytoplasmic color in the context of the sample and its controls. |
| Strength and validation limits | HPA labels the tissue profile Enhanced and reports Medium staining in its listed positive cells (HPA: tissue IHC). It also notes medium agreement with RNA data and pending external verification; neither designation guarantees that every section or antibody run will reproduce the profile (HPA: reliability description). |
| Antibody-specific evidence | Among the listed antibodies, CAB010109 has Enhanced IHC validation; HPA052658 and HPA074330 have no IHC status supplied (HPA: antibody records). Treat ICC support for all three as evidence for that application only (HPA: antibody records). |
| IF/ICC Q: Where should signal appear? | A: Mainly in the nucleoplasm; nuclear bodies are additionally supported, while cytosol is uncertain (HPA: subcellular ICC-IF). This is an interpretation cue for IF/ICC, not an IHC-P protocol or a reason to score diffuse cytoplasmic chromogen as positive. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected nuclei are blank in testis preleptotene spermatocytes. | The positive cells may be absent from the viewed field, or the IHC run may have failed (HPA: Medium in preleptotene spermatocytes; general IHC practice). | Find the relevant cells on the counterstained section; check a working positive control and detection reagents, then review the catalog antibody's IHC-P conditions (general IHC practice). Do not infer target-specific fixation sensitivity: none is supplied here. |
| Nucleoli stain more strongly than surrounding nuclei. | That reverses the reported nucleolar exclusion (UniProt O15318). Misread nuclear detail or nonspecific staining are possibilities (general IHC practice). | Recheck nuclear boundaries against the counterstain and compare a no-primary control (general IHC practice). Score only a reproducible pattern consistent with nuclear staining and nucleolar sparing (HPA: tissue IHC; UniProt O15318). |
| Cytoplasm dominates the signal in an adult tissue section. | HPA's tissue profile is nuclear, although UniProt also lists cytoplasm (HPA: tissue IHC; UniProt O15318). Location alone cannot establish whether the color is specific. | Compare nuclear signal in the expected positive cells with the no-primary control (HPA: tissue IHC; general IHC practice). Record cytoplasmic staining separately until its specificity is supported; do not convert an IF/ICC observation into an IHC-P expectation. |
| HPA-listed negative cells stain strongly. | The result disagrees with the cell-specific Not detected calls (HPA: tissue IHC). Cross-reactivity or endogenous detection activity may contribute (general IHC practice). | Check a no-primary control and the relevant endogenous-activity control for the chromogenic system (general IHC practice). Reassess cell identity and nuclear localisation before treating the staining as POLR3G (HPA: tissue IHC). |
| Broad, uneven chromogen makes nuclear scoring difficult. | Background from blocking, washing or detection conditions can obscure the expected nuclear distribution (general IHC practice; HPA: tissue profile). | Compare tissue and blank areas with a no-primary control; optimize blocking and washing within the established IHC workflow (general IHC practice). Score cell-specific nuclear signal only when background permits a clear distinction. |
| Signal is weak in a cell population reported as Medium. | HPA levels describe its observed samples and do not set a guaranteed intensity for each section (HPA: tissue IHC). Weak detection or a poorly represented positive cell population are also possible (general IHC practice). | Confirm the named cells are present, compare a validated positive section, and review the catalog antibody's IHC-P dilution and retrieval instructions (general IHC practice). No POLR3G-specific retrieval or fixation effect is established by the supplied sources. |
Experiment evidence
IHC & IF images showing POLR3G staining
A10669 · IHCImmunohistochemistry (IHC) analysis of paraffin-embedded Mouse Brain, antibody was diluted at 1:100.
A10669 · IHCImmunohistochemistry (IHC) analysis of paraffin-embedded Mouse Brain, antibody was diluted at 1:100.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000113356-POLR3G/tissue/cerebral+cortex#imid_18248951
HPA · EsophagusHuman Protein Atlas IHC, Esophagus. Source: https://www.proteinatlas.org/ENSG00000113356-POLR3G/tissue/esophagus#imid_18248939
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000113356-POLR3G/tissue/colon#imid_18248893
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000113356-POLR3G/tissue/liver#imid_18248886
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB010109; green = POLR3G, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB010109; green = POLR3G, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for POLR3G IHC & IF
🧪Run skin first: keratinocytes should stain (HPA: Medium in skin keratinocytes). Use adipose tissue as the negative comparator; adipocytes should be unstained (HPA: Not detected in adipocytes), and score any unstained cells within the skin section as internal negatives only after confirming their cell identity.
Positive control tissue: Skin (Keratinocytes, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show POLR3G in A-431, U-251MG, U2OS, PC-3, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; an isotype control matched to the primary antibody’s host species and immunoglobulin class or subclass; and POLR3G knockout tissue or cells as a biological specificity control (standard IHC practice). On skin sections, block endogenous peroxidase and distinguish chromogenic signal from melanin pigment (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and retrieval requirement are unreported in the supplied evidence; optimize antigen retrieval for the chosen IHC-validated antibody (standard IHC practice). The selected A10669 mouse-brain paraffin-section caption reports a 1:100 dilution but does not report the fixative (selected-SKU IHC caption: fixative unreported), so it cannot establish fixation. Whether frozen sections or IF are easier is unreported; skin pigment can complicate chromogenic scoring (standard IHC practice). The selected A10669 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image A10669).
HPA tissue IHC evidence for POLR3G
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Pending external verification.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Skin | Keratinocytes | Medium | Protein (IHC) | HPA → |
| Cervix | Squamous epithelial cells | Medium | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Medium | Protein (IHC) | HPA → |
| Oral mucosa | Squamous epithelial cells | Medium | Protein (IHC) | HPA → |
| Ovary | Follicle cells | 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 → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced POLR3G IHC Tips
Troubleshoot POLR3G staining by checking nuclear localisation, tissue controls, and the processing details documented for the antibody.
What retrieval should I start with for POLR3G paraffin-section IHC?
Start with Tris-EDTA at pH 9.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule: nuclear antigen). Cool sections consistently, then compare a retrieved section with an otherwise identical section lacking retrieval to judge whether staining improves (standard IHC practice). If nuclear staining remains weak, vary heating time in small increments before trying another buffer as a fallback, while monitoring tissue damage (standard IHC practice). Score signal in intact nuclei because tissue IHC shows predominantly nuclear expression, with reported staining in testis and squamous epithelia (HPA: tissue IHC profile).
Could fixation explain weak POLR3G staining in my paraffin sections?
POLR3G-specific sensitivity to fixation is unknown from the supplied evidence; the catalog antibody image identifies paraffin-embedded mouse brain but does not report its fixative (A10669 IHC caption). Record fixative, fixation duration, section age, and retrieval conditions for each sample before comparing staining intensity across runs (standard IHC practice). When weak staining is widespread, compare similarly processed sections with different fixation histories and include a tissue expected to show nuclear signal (standard IHC practice; HPA: tissue IHC profile). Treat differences as processing effects to investigate, rather than evidence that a particular fixative masks POLR3G, until matched experiments establish that effect (standard IHC practice).
Should I accept cytoplasmic or nucleolar POLR3G staining?
Prioritise nuclear staining in paraffin-section IHC because the tissue profile describes nuclear expression, while UniProt also lists cytoplasmic localisation and exclusion from nucleoli (HPA: tissue IHC profile; UniProt O15318: subcellular location). Inspect the haematoxylin counterstain at high magnification to distinguish nucleoplasm from nucleoli and from pigment or precipitate overlying a nucleus (standard IHC practice). Diffuse cytoplasmic signal alone merits comparison with nuclear signal in a positive control, especially because HPA rates cytosolic localisation as uncertain in its subcellular data (HPA: subcellular location). Keep nuclear and cytoplasmic observations separate in the score rather than combining them into one positive call (standard IHC practice).
How can I assess epitope specificity against POLR3G and POLR3GL?
Do not interpret staining as POLR3G-specific solely from its nuclear pattern: POLR3G and POLR3GL occupy alternative Pol III complexes, and the record lists no POLR3G isoforms (UniProt O15318: function and isoforms). Obtain the catalog antibody’s immunogen or epitope information, then assess sequence similarity to POLR3GL before assigning molecular specificity (standard IHC practice; UniProt O15318: function). If specificity remains uncertain, compare staining after an independent POLR3G perturbation while preserving tissue morphology and detection settings (standard IHC practice). The annotated phosphorylation sites at residues 133 and 157 are reasons to document epitope placement if known, not evidence of altered IHC binding (UniProt O15318: modified residues).
How should I cross-check a POLR3G IHC pattern by IF?
Use IF as a separate cross-check and multiplex POLR3G with a marker identifying the cell population being scored, such as a validated squamous epithelial marker when examining squamous epithelium (HPA: tissue IHC profile; standard IF practice). Choose a far-red fluorophore for a channel with substantial tissue autofluorescence, and inspect single-channel controls before calling overlap (standard IF practice). POLR3G has no annotated transmembrane segment and is mainly nucleoplasmic, so permeabilise fixed cells sufficiently for antibody access to intracellular and nuclear epitopes without assuming the exact epitope is known (UniProt O15318: topology; HPA: subcellular location; standard IF practice). Keep IF fixation and permeabilisation records separate from the paraffin-section caption, which reports no fixative (A10669 IHC caption).
What should I change when POLR3G chromogenic staining is diffuse?
Check a no-primary control and inspect whether brown colour follows tissue edges, folds, pigment, or areas lacking intact nuclei before treating it as POLR3G signal (standard IHC practice). Block endogenous peroxidase before chromogenic detection, and compare background after adjusting primary-antibody concentration, incubation, and wash stringency one variable at a time (standard IHC practice). The catalog image reports antibody dilution of 1:100 in paraffin-embedded mouse brain, but provides no fixative or background assessment (A10669 IHC caption). Interpret a cleaner nuclear pattern against tissue controls because reported tissue staining is nuclear and includes cell populations with undetected signal (HPA: tissue IHC profile and negative entries).
How should I quantify POLR3G across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scored cell population and compartment before analysis, then report the percentage of positive nuclei or an H-score on a 0–300 scale using intensity grades 0–3 (standard IHC practice; HPA: tissue IHC profile). Normalise positive counts to the number of evaluable nuclei in the same annotated region, and report tissue area if using positive cells per mm² (standard IHC practice). Keep exposure, chromogen development, counterstain, and positivity thresholds consistent across compared sections (standard IHC practice). Report nuclear and cytoplasmic scores separately, and stratify by cell type because HPA shows differing staining levels among tissue compartments (HPA: tissue IHC profile; standard IHC practice).
When is a POLR3G-positive IHC result convincing?
A convincing result places signal in intact nuclei of the scored cells and shows an interpretable contrast with controls, consistent with the reported tissue pattern (HPA: tissue IHC profile; standard IHC practice). Compare expected positive populations, such as testicular preleptotene spermatocytes or skin keratinocytes, with a population reported as undetected, such as adipocytes (HPA: tissue IHC positive and negative entries). Treat nucleolar-only staining, necrotic regions, tissue-edge colour, and residual endogenous-enzyme signal as possible artefacts requiring control review (UniProt O15318: nucleolar exclusion; standard IHC practice). The tissue profile has enhanced reliability but only medium staining-to-RNA consistency and awaits external verification, so confirm consequential calls independently (HPA: reliability description).
Boster reagents
Best POLR3G / DNA-directed RNA polymerase III subunit RPC7 IHC Antibodies
A10669 lists IHC and IF applications (catalog: applications) and human and mouse reactivity (catalog: reactivity). Its IHC figure shows paraffin-embedded mouse brain at 1:100 (IHC image caption).
Real IHC data
Anti-POLR3G Antibody
Cat # A10669
A10669 will render with an IHC figure of paraffin-embedded mouse brain stained at 1:100 (IHC image caption). IF is listed as an application, and human and mouse are listed as reactive species (catalog: applications and reactivity); no IF figure is provided (catalog: IF image alts).
Which to pick: Choose A10669 for paraffin-section tissue IHC based on its mouse brain figure (IHC image caption); the fixative is unreported (IHC image caption). For IF, A10669 is listed (catalog: applications), but ICC validation is unreported (catalog: applications). For work across species, it lists human and mouse reactivity (catalog: reactivity), while the IHC figure documents mouse brain only (IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O15318 (RPC7_HUMAN, DNA-directed RNA polymerase III subunit RPC7).
- Human Protein Atlas. POLR3G tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. POLR3G subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the nuclear bodies and cytosol..
- Human Protein Atlas. POLR3G antibody validation summary (3 antibodies).
- Identification of MYC co-expression gene: POLR3G is associated with cell senescence, immunotherapy, chemotherapy responses, and clinical prognosis in bladder cancer patients. Translational oncology 2026 — PMC13092746.
- The POLR3G Subunit of Human RNA Polymerase III Regulates Tumorigenesis and Metastasis in Triple-Negative Breast Cancer. Cancers 2022 — PMC9735567.
- PubMed PMID:9171375 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:15372022 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for POLR3G (DNA-directed RNA polymerase III subunit RPC7). Expected localisation Nuclear staining observed in tissue (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A10669 at 1:100-1:300 (the product's own IHC validation image is shown). Controls (HPA): positive Skin (Keratinocytes); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded Mouse Brain tissue; fixative not specified (datasheet A10669); 2. Fixation: Image fixative and duration unreported (datasheet A10669); 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 Tris-EDTA buffer, pH 9.0 , 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-POLR3G, 1:100-1:300 (datasheet A10669); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A10669); Caveat: Tissue staining awaits external verification (HPA tissue IHC); Isoform / epitope: No annotated isoforms or processing changes (UniProt); Regulation: Expressed in dividing cells (UniProt). Evidence: UniProt O15318, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A10669, PMC13092746. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/POLR3G/immunohistochemistry-ihc-guide.