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RPS21 / Small ribosomal subunit protein eS21 · IHC design guide
Design Immunohistochemistry for RPS21
Plan RPS21 paraffin-section IHC around its ubiquitous cytoplasmic staining pattern (HPA tissue IHC). Use breast glandular cells as a high-staining control (HPA tissue IHC) and start the IHC-validated antibody at 2–5 μg/ml (datasheet A10068-2).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
RPS21 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 | Ubiquitous cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining across diverse cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A10068-2) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue+2 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific) | |
| Caveat | Adipocytes and myocytes may lack detectable staining (HPA tissue IHC) | |
| Regulation | No target-specific regulation annotated (UniProt) | |
| Isoform / epitope | Single 1–83 chain; no isoforms annotated (UniProt) |
Section 1
Recommended RPS21 IHC & IF Protocols
The catalog antibody uses EDTA pH 8.0 heat retrieval (datasheet A10068-2). Two published RPS21 IHC studies provide prostate tissue staining conditions (PMC5634644; PMC6202544).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A10068-2) |
| Fixation | Image fixative and duration unreported (datasheet A10068-2); 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 A10068-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A10068-2) |
| Primary antibody | Rabbit anti-RPS21, 2-5 μg/ml (datasheet A10068-2) |
| Primary incubation | Overnight at 4 °C (datasheet A10068-2) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A10068-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | RPS21-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Ubiquitous cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 for the catalog antibody (datasheet A10068-2). Use pH 6 retrieval when reproducing the tissue array protocol (PMC5634644).
Section 2
What Is the Expected RPS21 Staining Pattern?
RPS21 is a 40S ribosomal protein detected on cytosolic polysomes and in ribosomes associated with rough endoplasmic reticulum (UniProt P63220). Expect predominantly cytoplasmic staining in many cell types, including breast glandular cells and cortical neurons (HPA tissue IHC: High). HPA describes the tissue pattern as ubiquitous cytoplasmic expression, but rates its reliability Approved, with medium agreement between staining and RNA data and external verification pending (HPA tissue IHC). RPS21 has no transmembrane segment (UniProt P63220 topology).
| Cytoplasmic chromogenic signal is strong in breast glandular cells or cortical neurons (HPA tissue IHC: High). | This fits the reported compartment and known positive cell types (UniProt P63220; HPA tissue IHC). Compare staining within the tissue: HPA also reports High signal in lymph-node germinal center cells and cerebellar Purkinje cells (HPA tissue IHC). Intensity alone does not establish antibody specificity (HPA antibody validation: IHC Approved; external verification pending). |
| Signal is predominantly nuclear, or outlines the plasma membrane while cytoplasm is unstained. | That distribution conflicts with the reported cytosolic and rough-ER association and the HPA cytoplasmic IHC profile (UniProt P63220; HPA tissue IHC). Check morphology, counterstain, detection controls, and a known positive tissue before interpreting it as RPS21; a single unusual compartment cannot establish a new localization. |
| Adipocytes, skeletal myocytes, or smooth-muscle cells stain strongly. | HPA reports RPS21 as Not detected in those cell types (HPA tissue IHC). Strong staining there raises concern for cross-reactivity or endogenous chromogenic activity (general IHC practice). Compare a matched negative-reagent control and inspect whether the color follows tissue structures rather than cell cytoplasm; HPA's Not detected call does not prove absolute protein absence. |
| Brown color spreads across stroma, tissue edges, or empty areas without clear cellular boundaries. | A diffuse deposit is hard to reconcile with the reported cytoplasmic pattern (HPA tissue IHC). Uneven detection or insufficient washing can create background (general IHC practice). Judge the result against a matched negative-reagent control and a positive tissue section, then score only signal that can be assigned to intact cells. |
| Breast glandular cells or lymph-node germinal center cells show no convincing signal (HPA tissue IHC: High). | A blank known positive section makes a negative study section uninterpretable (general IHC practice). Verify tissue preservation, retrieval, primary antibody application, and chromogen performance with run controls. HPA's IHC Approved rating has medium staining–RNA consistency and pending external verification, so investigate the assay before treating absence as biology (HPA tissue IHC). |
💡Expected RPS21 appearanceCall positive when cytoplasmic chromogen is clear in expected cells such as breast glandular cells or cortical neurons, where HPA reports High staining; isolated nuclear-only or strong adipocyte staining is suspect rather than a matching pattern (HPA tissue IHC; UniProt P63220).
| Compartment and epitope context | RPS21 is a 40S subunit component on cytosolic polysomes and rough-ER-associated ribosomes; it has no transmembrane segment (UniProt P63220). Expect cellular cytoplasmic staining, while routine chromogenic IHC may not resolve cytosol from ER-associated ribosomes (general IHC practice). |
| Tissue-dependent contrast | HPA reports High staining in breast glandular cells and lymph-node germinal center cells, Low staining in kidney tubule cells, and Not detected in adipocytes (HPA tissue IHC). Choose a reported High cell population as a run control and use the Low and Not detected calls as comparisons, not absolute expression limits. |
| Antibody evidence | The listed antibody HPA003371 is IHC Approved, while HPA describes medium consistency with RNA expression and pending external verification (HPA antibody validation; HPA tissue IHC). Treat the predicted pattern as a reference for evaluation, not proof that every colored cell is specific. |
| Maturation and isoform annotation | UniProt lists one chain spanning residues 1–83, no signal peptide or propeptide, and no annotated isoforms (UniProt P63220). The supplied record therefore gives no processing or alternate-isoform pattern to explain compartment shifts; these annotations do not specify retrieval conditions. |
| IF/ICC Q&A: where should signal appear? | HPA reports ER localization as approved and cytosol localization as supported in ICC-IF; its listed antibody has ICC Supported status (HPA subcellular; HPA antibody validation). Interpret IF/ICC localization on its dedicated guide page: those image-based compartment calls do not set a paraffin IHC protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known High tissue is blank while its morphology is intact (HPA tissue IHC: breast glandular cells High). | A failed staining step or weak detection is possible (general IHC practice); HPA staining levels do not establish RPS21 fixation sensitivity. | Check run controls, primary antibody addition, detection reagents, chromogen, and the locally validated retrieval workflow (general IHC practice). Repeat with a reported High cell population before scoring other sections negative (HPA tissue IHC). |
| All compartments and tissue structures appear brown, including areas without intact cells. | Diffuse background may arise from inadequate washing or excess detection signal (general IHC practice), rather than the HPA cytoplasmic pattern (HPA tissue IHC). | Inspect the negative-reagent control, wash and detection steps, and staining time; compare intact cells with adjacent acellular areas (general IHC practice). |
| Unexpected strong color appears in adipocytes or skeletal myocytes (HPA tissue IHC: Not detected). | Cross-reactivity or endogenous chromogenic activity is possible (general IHC practice). HPA's Not detected designation is an observation, not a universal absence claim (HPA tissue IHC). | Run a matched negative-reagent control, examine the intracellular distribution, and compare a reported High tissue in the same staining run (general IHC practice; HPA tissue IHC). |
| Only nuclei stain, with no corresponding cytoplasmic signal. | A nuclear-only result conflicts with cytosolic and rough-ER-associated localization (UniProt P63220) and HPA's cytoplasmic IHC profile (HPA tissue IHC). | Confirm nuclear boundaries with the counterstain, check control sections and detection artifacts, then repeat before assigning the nuclear color to RPS21 (general IHC practice). |
| Low-level cells look negative after the assay detects a reported High control. | Weak signal may fall below the scoring threshold; HPA reports Low staining in kidney tubule cells and prostate glandular cells (HPA tissue IHC). | Record cell-specific intensity and the assay's detection threshold; avoid converting a Low reference call into a required strong-positive control (HPA tissue IHC; general IHC practice). |
| Replicate sections give inconsistent cytoplasmic staining. | Variation in section handling or staining steps can affect chromogenic IHC (general IHC practice). HPA reports medium staining–RNA consistency and pending external verification, without target-specific fixation-effect data (HPA tissue IHC). | Compare matched sections and run controls, document retrieval and detection settings, and reproduce the result before drawing a biological conclusion (general IHC practice). |
Experiment evidence
IHC & IF images showing RPS21 staining
A10068-2 · IHCIHC analysis of RPS21 using anti-RPS21 antibody (A10068-2). RPS21 was detected in a paraffin-embedded section of human liver cancer tissue.
A10068-2 · IFIF analysis of RPS21 using anti-RPS21 antibody (A10068-2). RPS21 was detected in an immunocytochemical section of HELA cells. Enzyme antigen
A10068 · IHCImmunohistochemistry analysis of paraffin-embedded human breast carcinoma tissue, using RPS21 Antibody. The picture on the right is blocked
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000171858-RPS21/tissue/cerebral+cortex#imid_1249364
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000171858-RPS21/tissue/colon#imid_1249298
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000171858-RPS21/tissue/liver#imid_1249299
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000171858-RPS21/tissue/kidney#imid_1249300
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003371; green = RPS21, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003371; green = RPS21, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for RPS21 IHC & IF
🧪Run adrenal gland first: its glandular cells should stain strongly (HPA: High in adrenal gland glandular cells). Use adipose tissue as the negative comparator (HPA: Not detected in adipocytes); on the adrenal slide, cell-free areas should lack specific stain, but the supplied HPA row does not identify an internal negative cell type.
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RPS21 in A-431, U-251MG, U2OS, Sperm, with annotated localisation: Endoplasmic reticulum (approved), Cytosol (supported) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and concentration-matched rabbit IgG controls, plus a validated RPS21 knockout sample if available (selected caption: rabbit anti-RPS21 antibody; standard IHC practice). Check endogenous peroxidase background on the adrenal section before interpreting DAB staining (standard chromogenic IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A10068-2 paraffin-section caption does not state a fixative (selected caption: fixative unreported). The caption documents heat-mediated retrieval in EDTA at pH 8.0 for paraffin IHC, but does not establish that retrieval is required (selected caption: EDTA retrieval). The supplied evidence does not establish that frozen sections or IF are easier; assess adrenal-section background with the no-primary control (standard IHC practice).
HPA tissue IHC evidence for RPS21
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
Section 3
Advanced RPS21 IHC Tips
Troubleshoot RPS21 staining by checking retrieval, cytoplasmic localisation and cell type before comparing signal intensity across paraffin sections.
What retrieval should I try first when RPS21 staining is weak?
Start with heat-mediated retrieval in EDTA pH 8.0 for paraffin sections (datasheet A10068-2). The selected image used that retrieval, followed by 10% goat serum block and 2 μg/ml primary antibody overnight at 4°C (datasheet A10068-2). If signal remains weak, compare retrieval duration on adjacent sections while keeping antibody concentration and detection conditions constant (standard IHC practice). Judge improvement by stronger cytoplasmic staining in viable cells without increased tissue-edge staining or diffuse background; RPS21 is associated with cytosolic and rough-ER ribosomes (UniProt P63220).
Could fixation explain inconsistent RPS21 staining between paraffin blocks?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not report its fixative (datasheet A10068-2). Record each block’s fixative, fixation duration and processing history before comparing staining intensity, because these variables can alter antigen accessibility in IHC (standard IHC practice). Run blocks together with the same EDTA pH 8.0 retrieval and 2 μg/ml primary antibody conditions used for the selected image (datasheet A10068-2). If staining differs, compare viable regions and an internal reference cell population on each section before attributing the difference to RPS21 abundance (standard IHC practice).
Should convincing RPS21 signal look diffuse or organelle-associated?
Expect cytoplasmic signal because RPS21 is a component of the 40S small ribosomal subunit and occurs on cytosolic polysomes and rough-ER-associated ribosomes (UniProt P63220). The tissue IHC profile is described as ubiquitous cytoplasmic expression, while subcellular imaging places RPS21 in the ER and cytosol (HPA tissue IHC; HPA subcellular). Assess the distribution within viable cells alongside morphology and counterstain rather than requiring one uniform cytoplasmic texture (standard IHC practice). Predominantly nuclear-only staining or signal confined to extracellular deposits merits review of background and detection controls before interpretation (UniProt P63220 localisation; standard IHC practice).
How should epitope uncertainty affect interpretation of patchy staining?
The supplied record lists 0 alternative isoforms and one 1–83 chain for RPS21, so an isoform-specific staining explanation is unsupported (UniProt P63220). It reports N-acetylmethionine at residue 1 and N6-acetyllysine at residue 81, but no antibody epitope is supplied (UniProt P63220; datasheet A10068-2). Do not infer that either modification causes a staining difference without epitope mapping and direct experimental evidence (standard IHC practice). Compare adjacent sections under matched retrieval and primary-antibody conditions, then evaluate whether patchiness tracks viable cell populations, section edges or tissue damage (standard IHC practice).
How can I check RPS21 localisation by IF alongside the IHC result?
Use IF as a separate localisation check: RPS21 has supported cytosolic and approved ER localisation in subcellular imaging (HPA subcellular). Multiplex the RPS21 channel with a marker for the cell type being evaluated, then confirm that the signals occupy the same identified cells without requiring identical subcellular patterns (standard IF practice). Choose a fluorophore channel with low tissue autofluorescence, and include single-channel controls to assess bleed-through (standard IF practice). Because RPS21 lacks a transmembrane segment and is associated with cytosolic ribosomes, permeabilise sufficiently to expose the cytosolic side of membranes while preserving cell structure (UniProt P63220 topology and localisation; standard IF practice).
What should I change when the DAB background obscures cytoplasmic signal?
First compare the stained section with a no-primary control to distinguish antibody-dependent signal from detection background (standard IHC practice). The selected image used 10% goat serum block, 2 μg/ml primary antibody overnight at 4°C, a peroxidase-conjugated secondary for 30 minutes at 37°C, and DAB development (datasheet A10068-2). Include an endogenous-peroxidase block before HRP detection and monitor DAB development on matched sections; these are general chromogenic IHC controls (standard IHC practice). If background persists, assess whether it concentrates at section edges, damaged regions or tissue deposits before changing the primary-antibody conditions (standard IHC practice).
How should I score RPS21 across sections with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and viable tissue area before scoring, since HPA reports different staining levels among named cell types (HPA tissue IHC). For chromogenic IHC, record the percentage of positive cells and an intensity-based H-score within that population, using the same threshold and imaging conditions across sections (standard IHC practice). If counting positive cells, report density per mm² of viable tissue and normalise counts to the measured viable area (standard IHC practice). Exclude necrosis and section edges, and keep retrieval, 2 μg/ml primary-antibody concentration and DAB development consistent when comparing the selected workflow (datasheet A10068-2; standard IHC practice).
When is apparent RPS21 positivity more likely to be artefact?
Treat cytoplasmic staining in viable cells as anatomically plausible because RPS21 belongs to cytosolic and rough-ER-associated ribosomes (UniProt P63220). HPA reports high staining in Purkinje cells and no detected staining in skeletal-muscle myocytes, so identify the stained cell type before generalising from a tissue section (HPA tissue IHC). Review nuclear-only signal, tissue-edge gradients, necrotic regions and staining that persists in a no-primary control as possible artefacts (UniProt P63220 localisation; standard IHC practice). With HRP and DAB detection, check endogenous-peroxidase blocking and the no-primary control before calling deposit-associated brown colour RPS21-positive (datasheet A10068-2 detection; standard IHC practice).
Boster reagents
Best RPS21 / Small ribosomal subunit protein eS21 IHC Antibodies
Two anti-RPS21 antibodies have human paraffin-section IHC images (catalog IHC captions). A10068-2 also has an IF image of HeLa cells (catalog IF caption); both list Human, Mouse and Rat reactivity (catalog applications).
Real IHC data
Anti-RPS21 Antibody ®
Cat # A10068-2
Real IHC data
Anti-RPS21/Ribosomal Protein S21 Antibody
Cat # A10068
A10068-2 will render with IHC of a human liver cancer paraffin section and has IF data from HeLa cells (A10068-2 IHC and IF captions). A10068 will render with IHC of a human breast carcinoma paraffin section and lists IF/ICC among its applications, though no IF image is supplied (A10068 IHC caption; catalog applications and IF image list).
Which to pick: For tissue IHC, choose A10068-2 if its documented EDTA retrieval and 2 μg/ml overnight incubation fit your workflow (A10068-2 IHC caption); A10068 is another paraffin-section option, with peptide-blocked human breast carcinoma staining and a listed 1:100–1:300 IHC range (A10068 IHC caption; catalog dilution). For IF/ICC, choose A10068-2 because its HeLa-cell IF image documents staining at 5 μg/ml (A10068-2 IF caption); A10068 lists IF/ICC and is described as polyclonal, but has no IF image in this payload (A10068 catalog applications, dilution_raw and IF image list). Both list Human, Mouse and Rat reactivity, while the supplied IHC images show human samples only; neither IHC caption reports a fixative (catalog reactivity; A10068 and A10068-2 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P63220 (RS21_HUMAN, Small ribosomal subunit protein eS21).
- Human Protein Atlas. RPS21 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. RPS21 subcellular location (ICC-IF): Localized to the endoplasmic reticulum and cytosol..
- Human Protein Atlas. RPS21 antibody validation summary (1 antibodies).
- Expression of ribosomal proteins in normal and cancerous human prostate tissue. PloS one 2017 — PMC5634644.
- Identification of the key genes and pathways in prostate cancer. Oncology letters 2018 — PMC6202544.
- PubMed PMID:8332502 — UniProt-cited evidence.
- PubMed PMID:10900511 — UniProt-cited evidence.
- PubMed PMID:11875025 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for RPS21 (Small ribosomal subunit protein eS21). Expected localisation Ubiquitous cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A10068-2). Validated IHC antibody: A10068-2 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A10068-2); 2. Fixation: Image fixative and duration unreported (datasheet A10068-2); 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 A10068-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A10068-2); 8. Primary antibody: Rabbit anti-RPS21, 2-5 μg/ml (datasheet A10068-2); 9. Primary incubation: Overnight at 4 °C (datasheet A10068-2). Watch for: Fixation: Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific); Caveat: Adipocytes and myocytes may lack detectable staining (HPA tissue IHC); Isoform / epitope: Single 1–83 chain; no isoforms annotated (UniProt); Regulation: No target-specific regulation annotated (UniProt). Evidence: UniProt P63220, HPA tissue IHC · Approved, HPA ICC-IF · endoplasmic reticulum and cytosol., Datasheet A10068-2, PMC5634644, PMC6202544. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/RPS21/immunohistochemistry-ihc-guide.