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RBM15B / Putative RNA-binding protein 15B · IHC design guide
Design Immunohistochemistry for RBM15B
Plan RBM15B IHC in paraffin sections around granular nuclear staining, using colon glandular cells as a medium-staining reference (HPA tissue IHC). The guide covers the catalog antibody’s IHC workflow (datasheet A11484-1) and the cytoplasmic positivity HPA disregarded (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
RBM15B 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 | Granular nuclear staining (HPA tissue IHC) | |
| Staining pattern | Granular nuclei in several tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A11484-1) | |
| Positive control | Caudate+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 paraffin-section fixation consistent (standard IHC practice; not target-specific) | |
| Caveat | Cytoplasmic positivity was disregarded (HPA tissue IHC) | |
| Regulation | Ubiquitously expressed (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended RBM15B IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by three published RBM15B IHC protocols (PMC8416726; PMC9760713; PMC10550633).
| Sample | Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A11484-1) |
| Fixation | Image fixative and duration unreported (datasheet A11484-1); 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 A11484-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A11484-1) |
| Primary antibody | Rabbit anti-RBM15B, 2-5 μg/ml (datasheet A11484-1) |
| Primary incubation | Overnight at 4 °C (datasheet A11484-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A11484-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | RBM15B-positive staining in neuronal cells of caudate (HPA tissue IHC: Medium). HPA tissue profile: Nuclear expression with a granular pattern in several tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA, pH 8.0 (datasheet A11484-1); citrate microwave retrieval is a published alternative (PMC8416726; PMC10550633).
Section 2
What Is the Expected RBM15B Staining Pattern?
RBM15B should appear mainly in nuclei, with granular nucleoplasmic staining and nucleolar exclusion (UniProt Q8NDT2; HPA tissue IHC). Look for medium staining in colon glandular cells, kidney tubular cells, or caudate neuronal cells (HPA tissue IHC). RBM15B has no transmembrane segment (UniProt Q8NDT2 topology). HPA rates its tissue IHC staining Approved, pending external verification, and explicitly disregards observed cytoplasmic positivity (HPA tissue IHC).
| Granular nuclear staining in colon glandular cells, with little apparent nucleolar staining. | This matches the reported medium signal in those cells (HPA tissue IHC) and granular nucleoplasmic, nucleolus-excluded localisation (UniProt Q8NDT2). Score the nuclear compartment, while recognising that the HPA tissue result remains pending external verification (HPA tissue IHC). |
| Prominent cytoplasmic stain, with weak or absent nuclear stain. | Treat cytoplasmic positivity as suspect: HPA observed and disregarded it in its tissue IHC assessment (HPA tissue IHC). RBM15B is reported in nuclear compartments and has no transmembrane segment (UniProt Q8NDT2). Review detection background and antibody specificity before assigning a biological interpretation (general IHC practice). |
| Strong stain in a cell population recorded as undetected by HPA, such as adipocytes. | HPA reports RBM15B as not detected in adipocytes (HPA tissue IHC). Check cell identification and compare the nuclear pattern with a known-positive cell population; unexpected staining can reflect cross-reactivity or endogenous detection activity (general IHC practice). An HPA negative result is not proof of absolute absence. |
| Broad, diffuse colour across tissue, including spaces outside cells. | This lacks the granular nuclear distribution reported for RBM15B (HPA tissue IHC; UniProt Q8NDT2). Consider nonspecific reagent binding, endogenous detection activity, or residual chromogen; inspect a matched no-primary control and review blocking and washes (general IHC practice). Do not score diffuse colour as RBM15B-positive nuclei. |
| No nuclear signal in colon glandular cells or kidney tubular cells. | Both populations showed medium staining in HPA tissue IHC (HPA tissue IHC). Check section quality, retrieval, antibody and detection steps against the assay's own controls (general IHC practice). A blank section alone cannot distinguish technical failure from variation in the sampled tissue. |
💡Expected RBM15B appearanceCall a section positive when the expected cells show predominantly medium, granular nuclear staining—such as colon glandular or kidney tubular cells (HPA tissue IHC)—with nucleolar exclusion consistent with UniProt Q8NDT2; diffuse cytoplasmic colour alone is suspect because HPA disregarded cytoplasmic positivity (HPA tissue IHC).
| Compartment used for scoring | Prioritise granular nuclei: HPA describes nuclear tissue staining, and UniProt places RBM15B in nucleoplasm, speckles and the nuclear envelope (HPA tissue IHC; UniProt Q8NDT2). The envelope annotation does not establish a separate IHC scoring pattern. |
| Choice of comparison cells | Colon glandular and kidney tubular cells were medium, while adipocytes and heart cardiomyocytes were not detected (HPA tissue IHC). Compare the specified cell populations, rather than treating an entire organ as uniformly positive or negative. |
| Antibody evidence | HPA lists HPA036645 as IHC Approved and its tissue result as pending external verification (HPA antibodies; HPA tissue IHC). HPA058136 has no listed IHC status; its Enhanced ICC status does not establish IHC performance (HPA antibodies). |
| Isoforms and epitope coverage | UniProt lists 2 RBM15B isoforms and an uninterrupted 1–890 protein chain (UniProt Q8NDT2). The supplied sources do not map antibody epitopes to isoforms, so an isoform-specific staining conclusion is unsupported. |
| IF/ICC Q&A: what localisation is expected? | HPA reports enhanced nucleoplasmic localisation in ICC-IF images from A-431, U-251MG and U2OS, with both listed antibodies rated ICC Enhanced (HPA subcellular; HPA antibodies). This supports a localisation comparison, not an IHC protocol or tissue-intensity prediction. |
| Situation | Likely cause | Next action |
|---|---|---|
| Only cytoplasmic colour is visible. | HPA observed cytoplasmic positivity but disregarded it; its supported tissue pattern is nuclear (HPA tissue IHC). | Review a no-primary control, detection reagents and wash conditions, then assess whether credible granular nuclei remain (general IHC practice; HPA tissue IHC). |
| Colon glandular cells appear completely negative. | HPA reported medium staining in this population, so a technical failure is possible (HPA tissue IHC; general IHC practice). | Check section integrity, retrieval, primary-antibody step and detection controls before interpreting the blank result (general IHC practice). |
| Colour is widespread and obscures nuclei. | Diffuse background does not fit HPA's granular nuclear pattern (HPA tissue IHC); nonspecific binding or endogenous detection activity may contribute (general IHC practice). | Compare a no-primary control and review blocking, detection chemistry, chromogen development and washes (general IHC practice). |
| Adipocytes stain strongly. | Adipocytes were not detected in HPA tissue IHC; cell identification, cross-reactivity or endogenous activity may explain the discrepancy (HPA tissue IHC; general IHC practice). | Verify the stained cell type and nuclear localisation, then compare with a known-positive population and matched controls (general IHC practice). |
| The apparent positive area changes when the counterstain is adjusted. | Counterstain can make weak chromogen harder to distinguish from nuclear detail (general IHC practice); HPA's expected pattern is granular and nuclear (HPA tissue IHC). | Review a lightly counterstained section and the detection control, and score chromogen confined to identifiable nuclei (general IHC practice). |
| An ICC-IF image looks nucleoplasmic, but tissue IHC does not. | HPA reports enhanced nucleoplasmic ICC-IF localisation, while tissue IHC is Approved pending external verification; these are separate assessments (HPA subcellular; HPA antibodies; HPA tissue IHC). | Evaluate tissue IHC with its own positive cells and controls; do not use the ICC rating to infer IHC sensitivity (HPA tissue IHC; HPA antibodies; general IHC practice). |
Experiment evidence
IHC & IF images showing RBM15B staining
A11484-1 · IHCIHC analysis of RBM15B using anti-RBM15B antibody (A11484-1). RBM15B was detected in a paraffin-embedded section of human breast cancer tiss
A11484-1 · IHCIHC analysis of RBM15B using anti-RBM15B antibody (A11484-1). RBM15B was detected in a paraffin-embedded section of human esophageal squamou
A11484-1 · IFIF analysis of RBM15B using anti-RBM15B antibody (A11484-1). RBM15B was detected in a paraffin-embedded section of human intestinal cancer t
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000259956-RBM15B/tissue/cerebral+cortex#imid_9548324
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000259956-RBM15B/tissue/colon#imid_9548250
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000259956-RBM15B/tissue/liver#imid_9548243
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000259956-RBM15B/tissue/kidney#imid_9548260
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036645; green = RBM15B, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036645; green = RBM15B, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for RBM15B IHC & IF
🧪Run colon first and score its glandular cells for RBM15B staining (HPA: Medium); use adipose tissue adipocytes as the negative comparator (HPA: Not detected). On the colon slide, treat non-glandular cells as an internal background reference only if they are unstained; the supplied HPA rows do not establish an internal negative cell population (HPA: colon glandular cells, Medium).
Positive control tissue: Caudate (Neuronal cells, HPA Medium)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RBM15B in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only slide, a concentration-matched nonimmune rabbit IgG isotype control, and an RBM15B knockout sample as a biological negative (caption: rabbit primary and HRP-conjugated anti-rabbit secondary; standard IHC control practice). Quench endogenous peroxidase and inspect luminal background when scoring colon glandular cells with DAB (HPA: colon glandular cells, Medium; caption: HRP/DAB detection).
⚠️Feasibility: The paraffin-section example uses heat retrieval in EDTA at pH 8.0, but it does not establish whether retrieval is required under other conditions (A11484-1 caption: EDTA retrieval). A target-specific fixation window or fixation effect is unreported, and the selected caption does not state the fixative (A11484-1 caption: fixative not stated). Frozen sections cannot be judged easier from the supplied evidence; ICC-IF images support assessing nucleoplasmic localisation, while colon luminal background needs review during chromogenic scoring (HPA: nucleoplasm; HPA: colon glandular cells, Medium; caption: DAB detection).
HPA tissue IHC evidence for RBM15B
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Pending external verification. Cytoplasmic positivity observed and disregarded.). 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 |
|---|---|---|---|---|
| Caudate | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Cerebellum | Cells in molecular layer | Medium | Protein (IHC) | HPA → |
| Cerebral cortex | Glial cells | Medium | Protein (IHC) | HPA → |
| Colon | Glandular cells | Medium | Protein (IHC) | HPA → |
| Duodenum | Glandular 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 → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced RBM15B IHC Tips
Troubleshoot RBM15B staining in paraffin sections by checking retrieval, nuclear localisation, assay controls and scoring consistency (datasheet A11484-1; HPA tissue IHC).
Which retrieval condition should I start with for RBM15B in paraffin sections?
Use heat-mediated retrieval in EDTA, pH 8.0, for the catalog antibody on paraffin sections (datasheet A11484-1). The selected tissue image used this condition before incubation with 2 μg/ml primary antibody overnight at 4°C (caption A11484-1). If nuclear staining is weak, check section processing and retrieval consistency before varying heating time across matched sections (standard IHC practice). Keep antibody concentration and detection constant during that comparison so a change in signal can be attributed to retrieval (standard IHC practice). Judge improvement by granular nuclear staining, rather than stronger cytoplasmic colour (HPA tissue IHC).
How should I investigate fixation as a cause of weak RBM15B staining?
Target-specific sensitivity to fixation is unknown: the selected RBM15B image identifies a paraffin section but does not state its fixative (caption A11484-1). Record the fixative and fixation duration for each specimen, then compare similarly processed sections using the documented EDTA, pH 8.0 retrieval condition (standard IHC practice; datasheet A11484-1). Include a previously stained control section in each run to distinguish specimen variation from a run-level failure (standard IHC practice). If a specimen is weak, assess tissue preservation and nuclear morphology before increasing primary antibody or retrieval intensity (standard IHC practice). Do not infer fixation sensitivity from the reported tissue staining pattern (HPA tissue IHC).
What RBM15B staining pattern should I accept as specific?
Look for predominantly nuclear staining with a granular pattern, consistent with the tissue profile (HPA tissue IHC). RBM15B is reported in the nucleoplasm, nuclear speckles and nuclear envelope, with granular nucleoplasmic staining that excludes nucleoli (UniProt Q8NDT2 subcellular location). The atlas describes its tissue IHC reliability as approved pending external verification and explicitly disregards cytoplasmic positivity (HPA tissue IHC). Compare nuclei in glandular cells of colon or tubular cells of kidney, both reported at medium staining, with adjacent background (HPA tissue IHC). Evaluate nuclear detail under the same counterstain and illumination settings across sections (standard IHC practice).
Could isoforms or epitope accessibility explain discordant nuclear staining?
RBM15B has 2 reported isoforms, while the supplied product caption does not identify the antibody epitope or establish isoform coverage (UniProt Q8NDT2 isoforms; caption A11484-1). Its three RNA recognition motifs occupy residues 139–219, 337–414 and 418–492, and its SPOC domain occupies 711–889 (UniProt Q8NDT2 domains). Reported phosphosites include serines 109 and 113, but their effect on this antibody's staining is unknown (UniProt Q8NDT2 modified residues). Check the antibody's documented immunogen before attributing discordance to an isoform or modification (standard IHC practice). Meanwhile, compare staining under the same retrieval and detection conditions (standard IHC practice).
How can IF help assess an ambiguous chromogenic RBM15B result?
Use IF/ICC as a complementary localisation check for an ambiguous IHC result; nucleoplasmic localisation is supported by cell images, including A-431, U-251MG and U2OS (HPA subcellular). For tissue multiplexing, pair RBM15B with a validated marker of the expected cell type, such as the glandular cells assessed in colon IHC (HPA tissue IHC; standard IF practice). Choose a fluorophore channel after measuring tissue autofluorescence, and include single-channel controls when separating overlapping signals (standard IF practice). RBM15B has no transmembrane segment and is nuclear, so permeabilisation must allow antibody access to the nucleus (UniProt Q8NDT2 topology and subcellular location; standard IF practice). Confirm the IF pattern against the granular nuclear IHC expectation (HPA tissue IHC).
What should I check when DAB obscures RBM15B nuclei?
First compare stained sections with a no-primary control to identify signal from the secondary reagent, chromogen or tissue rather than primary-antibody binding (standard IHC practice). The documented run used 10% goat serum blocking, a peroxidase-conjugated secondary for 30 minutes at 37°C, and DAB development (caption A11484-1). Check endogenous peroxidase blocking and shorten chromogen development if diffuse brown colour persists; these are general workflow adjustments (standard IHC practice). Review section edges and damaged areas separately because local background can distort nuclear assessment (standard IHC practice). Do not count cytoplasmic colour as RBM15B positivity (HPA tissue IHC).
How should I score RBM15B across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Score only interpretable nuclei within a defined cell population, because the reported tissue pattern is granular and nuclear (HPA tissue IHC; standard IHC practice). Record the percentage of positive nuclei and, if intensity grades are reproducible, calculate an H-score from 0–300 using percentages at intensity grades 0–3 (standard IHC practice). Normalise each percentage to the number of evaluable nuclei in that cell population, rather than to the whole section area (standard IHC practice). Keep retrieval, counterstain and scoring thresholds consistent between sections (standard IHC practice). Exclude folds, necrosis and edge staining before comparing samples (standard IHC practice).
How do I distinguish true RBM15B staining from artefact?
A plausible positive shows granular nuclear colour in intact cells, matching the reported tissue IHC pattern and nucleoplasmic localisation (HPA tissue IHC; UniProt Q8NDT2 subcellular location). Give greater weight to interpretable cells in a reported positive compartment, such as colon glandular cells or kidney tubular cells, which have medium atlas staining (HPA tissue IHC). Treat diffuse cytoplasmic colour cautiously because the atlas specifically disregards cytoplasmic positivity (HPA tissue IHC). Compare edge and necrotic regions with preserved interior tissue, and use a no-primary control to investigate endogenous enzyme or detection background (standard IHC practice). Resolve discrepancies with a matched positive control before assigning a biological difference (standard IHC practice).
Boster reagents
Best RBM15B / Putative RNA-binding protein 15B IHC Antibodies
A11484-1 has IHC images from human paraffin sections and IF images from human paraffin sections (catalog image captions); its listed reactivity is Human and Rat (catalog: reactivity).
Real IHC data
Anti-RBM15B Antibody ®
Cat # A11484-1
A11484-1 is listed for IHC and IF; its IHC captions show human breast, esophageal, gastric and liver cancer paraffin sections (catalog: applications; IHC image captions). Its IF captions show human intestinal and ovarian cancer paraffin sections (catalog: IF image captions).
Which to pick: For tissue IHC, choose A11484-1: its own captions document EDTA pH 8.0 retrieval and 2 μg/ml primary antibody in human paraffin sections; the fixative is unreported (catalog: IHC image captions). For IF, A11484-1 has paraffin-section examples at 5 μg/ml, while ICC validation is unreported (catalog: IF image captions; applications). For cross-species work, A11484-1 lists Human and Rat reactivity, but the supplied IHC and IF images show human samples only (catalog: reactivity; image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q8NDT2 (RB15B_HUMAN, Putative RNA-binding protein 15B).
- Human Protein Atlas. RBM15B tissue IHC expression (reliability: Approved).
- Human Protein Atlas. RBM15B subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. RBM15B antibody validation summary (2 antibodies).
- Identification and Characterization of Alcohol-related Hepatocellular Carcinoma Prognostic Subtypes based on an Integrative N6-methyladenosine methylation Model. International journal of biological sciences 2021 — PMC8416726.
- Diagnostic signature, subtype classification, and immune infiltration of key m6A regulators in osteomyelitis patients. Frontiers in genetics 2022 — PMC9760713.
- The predictive significance of a 5-m6A RNA methylation regulator signature in colorectal cancer. Heliyon 2023 — PMC10550633.
- PubMed PMID:16641997 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
- PubMed PMID:17974005 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for RBM15B (Putative RNA-binding protein 15B). Expected localisation Granular nuclear staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A11484-1). Validated IHC antibody: A11484-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Caudate (Neuronal cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A11484-1); 2. Fixation: Image fixative and duration unreported (datasheet A11484-1); 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 A11484-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A11484-1); 8. Primary antibody: Rabbit anti-RBM15B, 2-5 μg/ml (datasheet A11484-1); 9. Primary incubation: Overnight at 4 °C (datasheet A11484-1). Watch for: Fixation: Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Caveat: Cytoplasmic positivity was disregarded (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage is unspecified (UniProt); Regulation: Ubiquitously expressed (UniProt). Evidence: UniProt Q8NDT2, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet A11484-1, PMC8416726, PMC9760713. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/RBM15B/immunohistochemistry-ihc-guide.