RPS24 / Small ribosomal subunit protein eS24 · IHC design guide

Design Immunohistochemistry for RPS24

Plan RPS24 paraffin-section IHC around the cytoplasmic staining observed across several tissues (HPA tissue IHC). This guide covers fixation, staining controls and interpretation in light of the reported mismatch between antibody staining and RNA expression (HPA tissue IHC).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use

Immunohistochemistry protocol sheet for RPS24 (IHC for RPS24): expected localisation Cytoplasm (HPA tissue IHC); nucleus and nucleolus (UniProt), antibody A05730, validated IHC image, and IHC protocol steps — Printable **RPS24** IHC protocol sheet — expected localisation Cytoplasm (HPA tissue IHC); nucleus and nucleolus (UniProt), antibody A05730, controls and protocol steps. Open the full RPS24 IHC guide →

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RPS24 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	Cytoplasm (HPA tissue IHC); nucleus and nucleolus (UniProt)
	Staining pattern	Cytoplasmic staining in several tissues (HPA tissue IHC)
	Antigen retrieval	Citrate pH 6.0 HIER, heat-mediated (datasheet A05730)
	Positive control ⓘ	Breast+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 across samples (standard IHC practice; not target-specific)
	Caveat	Antibody staining has low consistency with RNA data (HPA tissue IHC)
	Regulation	Higher in tissues rich in proliferating cells (UniProt)
	Isoform / epitope	4 isoforms; epitope coverage is unknown (UniProt)

Section 1

Recommended RPS24 IHC & IF Protocols

The catalog antibody’s IHC-P protocol (datasheet A05730) is accompanied by published RPS24 IHC protocols for liver cancer and prostate tissue (PMC9820840; PMC5634644).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded rat brain tissue; fixative not specified (datasheet A05730)
Fixation	Image fixative and duration unreported (datasheet A05730); verify before use.
Sectioning	4–5 µm sections on charged slides (standard)
Deparaffinisation	Xylene, graded ethanol series to water (standard)
Antigen retrieval	Heat retrieval: Citrate pH 6.0 (datasheet A05730); 20 min, 95–100 °C (standard)
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-RPS24, 1:50-1:200 (datasheet A05730)
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	RPS24-positive staining in glandular cells of breast (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression in several tissues. No signal in the no-primary control.

💡Decision noteStart with heat-mediated citrate retrieval at pH 6.0 (datasheet A05730); the published prostate protocol also used pH 6 retrieval (PMC5634644).

Section 2

What Is the Expected RPS24 Staining Pattern?

RPS24 is a cytoplasmic ribosomal protein that is also reported in the nucleus and nucleolus (UniProt P62847: location and function). In paraffin-section IHC, expect mainly cytoplasmic staining in several tissues, including glandular cells in breast, duodenum, endometrium, and small intestine (HPA: tissue IHC). RPS24 has no transmembrane segment (UniProt P62847: topology). HPA rates its tissue IHC staining Approved but reports low consistency with RNA expression (HPA: tissue IHC reliability).

What am I looking at on my slide?
Cytoplasmic staining in breast or duodenal glandular cells.	This matches HPA's Medium staining in those cell types and its broader cytoplasmic IHC profile (HPA: tissue IHC). Judge the pattern in the named cells rather than expecting every cell in the section to stain equally; HPA reports low tissue RNA specificity and low consistency between staining and RNA data (HPA: tissue IHC).
Predominantly nuclear, nucleolar, or sharply membrane-bound chromogen, with little cytoplasmic staining.	UniProt includes nuclear and nucleolar RPS24, so nuclear signal alone does not prove an artifact (UniProt P62847: location). A pattern dominated by these compartments conflicts with HPA's cytoplasmic tissue IHC profile; a membrane outline also lacks support from RPS24 topology (HPA: tissue IHC; UniProt P62847: topology). Recheck controls before assigning either pattern to RPS24.
Strong staining in a cell population that HPA reports as Not detected.	For example, HPA reports Not detected in adipocytes and heart cardiomyocytes (HPA: tissue IHC). Unexpected staining may reflect antibody cross-reactivity or endogenous detection activity, both general IHC possibilities. A Not detected HPA result describes that assay's observation; it does not establish that the cell population can never contain RPS24.
Diffuse color over cells and surrounding tissue, with no clear intracellular pattern.	This is difficult to score as RPS24 because the reported tissue IHC pattern is cytoplasmic (HPA: tissue IHC). In general IHC practice, nonspecific antibody binding, incomplete blocking, or residual detection-reagent activity can produce background. Compare a no-primary control and review the staining distribution before increasing the assigned score.
No signal in breast or pancreatic glandular cells selected as a positive comparator.	HPA reports Medium staining in breast glandular cells and pancreatic exocrine glandular cells (HPA: tissue IHC). An absent signal in the selected cells makes the run difficult to interpret; check section identity, primary-antibody use, retrieval conditions, and detection controls as general IHC troubleshooting steps. HPA's Approved rating does not guarantee staining in every preparation (HPA: tissue IHC reliability).

💡Expected RPS24 appearanceCall a result positive when identifiable glandular cells show mainly cytoplasmic chromogen at a level consistent with HPA's Medium examples; isolated membrane outlines or diffuse section-wide color should not be scored as that pattern (HPA: tissue IHC; UniProt P62847: topology).

How each factor affects the staining
Compartment and application	HPA tissue IHC describes cytoplasmic expression, whereas HPA ICC-IF places signal in the endoplasmic reticulum and cytosol; UniProt also records nuclear and nucleolar localisation (HPA: tissue IHC and subcellular; UniProt P62847: location). Interpret paraffin-section chromogen against the tissue IHC pattern, since the ICC-IF result comes from a different application.
Choice of comparison tissue	HPA reports Medium staining in several glandular populations, Low staining in skin keratinocytes, and Not detected in adipocytes, bone-marrow hematopoietic cells, and heart cardiomyocytes (HPA: tissue IHC). These observed categories provide comparisons, with HPA's stated low staining-to-RNA consistency limiting predictions for an unlisted sample (HPA: tissue IHC reliability).
Antibody evidence	HPA003364 has Approved IHC status, while HPA073508 has no supplied IHC rating; both have Enhanced ICC status (HPA: antibody validation). ICC enhancement therefore cannot be treated as Enhanced IHC evidence. Use an antibody's IHC evidence when interpreting paraffin sections (HPA: antibody validation).
Isoforms and epitope coverage	UniProt lists four RPS24 isoforms and modified residues including N-acetylmethionine at position 1 and phosphothreonine at position 9 (UniProt P62847: isoforms and modified residues). The payload gives no antibody epitope, so isoform coverage or modification-dependent staining cannot be predicted; these annotations do not establish a retrieval or fixation effect.
Retrieval and detection conditions	Antigen retrieval and blocking are general paraffin-section IHC variables that can affect interpretability (general IHC practice). No supplied UniProt or HPA evidence establishes RPS24-specific fixation sensitivity or an optimal retrieval condition. Record the conditions used and interpret a failed run with its controls before attributing it to target biology.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
The expected cytoplasmic signal is absent in a selected Medium-staining glandular comparator.	A failed staining step or unsuitable run conditions are possible general IHC explanations; HPA's Medium category is an observed tissue result, not a guarantee for each section (HPA: tissue IHC).	Confirm the comparator cells are present, review primary-antibody and detection steps, and assess the run's controls before calling the test sample negative (general IHC practice).
Chromogen appears mainly in nuclei or along cell borders.	A nucleus-dominant result diverges from HPA's cytoplasmic tissue pattern, though UniProt also lists nuclear and nucleolar RPS24; a membrane outline is unsupported by the recorded topology (HPA: tissue IHC; UniProt P62847: location and topology).	Compare compartments in the positive comparator and inspect a no-primary control; report a reproducible discordant pattern separately from the expected IHC result (general IHC practice).
Adipocytes or cardiomyocytes stain more strongly than the chosen positive comparator.	HPA reports Not detected in adipocytes and cardiomyocytes, making cross-reactivity or endogenous detection activity possibilities to investigate (HPA: tissue IHC; general IHC practice).	Examine a no-primary control for detection-related color, then compare the suspect cells and comparator under the same staining conditions (general IHC practice).
Diffuse background obscures glandular cell boundaries.	Nonspecific binding or residual detection activity can obscure intracellular staining in chromogenic IHC (general IHC practice). The resulting diffuse pattern cannot be matched confidently to HPA's cytoplasmic profile (HPA: tissue IHC).	Check the no-primary control and review blocking, washing, and detection steps; score only cells whose staining can be localized reliably (general IHC practice).
Bone marrow appears negative despite a biological expectation of RPS24 expression.	UniProt describes higher levels in tissues with many proliferating cells, including bone marrow, while HPA reports Not detected in bone-marrow hematopoietic cells by tissue IHC (UniProt P62847: tissue specificity; HPA: tissue IHC).	Record the source-specific discrepancy. Use an HPA Medium-staining tissue as the staining comparator and avoid treating bone marrow as a guaranteed positive IHC control (HPA: tissue IHC).
Why does an ICC-IF image emphasize endoplasmic reticulum or cytosol when tissue IHC looks cytoplasmic?	HPA reports enhanced endoplasmic-reticulum and cytosol localisation for ICC-IF, while its paraffin tissue IHC summary says cytoplasmic expression (HPA: subcellular and tissue IHC).	Interpret each result within its application; use the cytoplasmic tissue IHC profile for this slide and the separate IF/ICC guide for fluorescence interpretation (HPA: tissue IHC and subcellular).

Source: UniProt P62847 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing RPS24 staining

Immunohistochemistry of paraffin-embedded rat brain using RPS24 Rabbit pAb at dilution of 1:200 .Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.

A05730 · IHCImmunohistochemistry of paraffin-embedded rat brain using RPS24 Rabbit pAb at dilution of 1:200 .Perform high pressure antigen retrieval wit

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000138326-RPS24/tissue/cerebral+cortex#imid_1258284

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000138326-RPS24/tissue/colon#imid_1258226

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000138326-RPS24/tissue/liver#imid_1258219

HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000138326-RPS24/tissue/pancreas#imid_1258170

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA003364; green = RPS24, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

Real IHC / IF data behind the staining-pattern expectations in this section.

Sample controls for RPS24 IHC & IF

🧪Run breast first: its glandular cells should stain at a medium level (HPA: breast glandular cells, Medium). Run adipose tissue as the negative; adipocytes within the breast section, when present, should show little or no staining (HPA: adipose tissue adipocytes, Not detected).

Positive control tissue: Breast (Glandular cells, HPA Medium)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RPS24 in A-431, U-251MG, U2OS, with annotated localisation: Endoplasmic reticulum (enhanced), Cytosol (enhanced) (HPA subcellular).

Technical controls: Include a no-primary (secondary-only) control, a concentration-matched rabbit polyclonal isotype control (A05730 caption: Rabbit pAb), and a biological specificity control using RPS24 knockout material or peptide competition if a suitable peptide is available. For chromogenic detection, block endogenous peroxidase; assess endogenous biotin if using avidin–biotin detection, particularly around adipose-rich regions of the breast section (HPA: breast glandular cells, Medium; adipose tissue adipocytes, Not detected).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported in the supplied evidence; the selected paraffin-section caption does not state its fixative (A05730 caption: fixative not stated). The caption uses high-pressure retrieval in 10 mM citrate, pH 6.0, at 1:200 for rat brain IHC, so retrieval conditions for the breast control require verification (A05730 caption). No matched evidence establishes that frozen sections or IF are easier; in breast sections, distinguish glandular staining from adipocyte-rich areas when assessing signal (HPA: breast glandular cells, Medium; adipose tissue adipocytes, Not detected).

HPA tissue IHC evidence for RPS24

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low 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
Breast	Glandular cells	Medium	Protein (IHC)	HPA →
Cerebellum	Purkinje cells	Medium	Protein (IHC)	HPA →
Duodenum	Glandular cells	Medium	Protein (IHC)	HPA →
Endometrium	Glandular cells	Medium	Protein (IHC)	HPA →
Pancreas	Exocrine 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 →
Bronchus	Respiratory epithelial cells	Not detected	Protein (IHC)	HPA →
Epididymis	Glandular cells	Not detected	Protein (IHC)	HPA →

Section 3

Advanced RPS24 IHC Tips

Troubleshoot RPS24 staining by checking retrieval, compartment, cell type, and controls before comparing signal across sections.

How should I retrieve RPS24 in paraffin sections with weak staining?

Start with heat-mediated retrieval in 10 mM citrate, pH 6.0, the condition reported for the catalog antibody in paraffin-embedded rat brain (datasheet A05730). Keep heating and cooling conditions identical across test and control sections, and begin antibody titration around the caption's 1:200 dilution (datasheet A05730; standard IHC practice). If staining remains weak, compare a longer citrate retrieval with a higher-pH buffer on adjacent sections, while monitoring tissue damage and nonspecific color (standard IHC practice). Judge improvement by cellular staining over matched background; cytoplasmic expression in several tissues provides a useful pattern to assess, although HPA reports low consistency with RNA expression (HPA tissue IHC).

Could fixation be masking RPS24 in my IHC sections?

The catalog image identifies paraffin-embedded rat brain but does not state its fixative, so RPS24-specific fixation sensitivity is unknown (datasheet A05730). For a controlled comparison, record fixative type and time, then stain adjacent sections with the same 10 mM citrate, pH 6.0 retrieval and 1:200 starting dilution (datasheet A05730; standard IHC practice). Compare preserved morphology, signal, and background rather than treating a darker section as proof of better antigen preservation (standard IHC practice). If archived sections differ in handling, include a shared reference section in each run; neither the protein's lack of a transmembrane segment nor its modified residues establishes fixation behavior (UniProt P62847; standard IHC practice).

Is nucleolar RPS24 staining credible when my tissue shows cytoplasm?

Cytoplasmic staining is compatible with the tissue IHC profile, while a nucleolar pool is compatible with RPS24's reported location and role in pre-rRNA processing (HPA tissue IHC; UniProt P62847). HPA cell imaging instead highlights endoplasmic reticulum and cytosol, so compartment patterns need assessment in the assay and specimen used here (HPA subcellular). Check cytoplasmic signal against a nuclear counterstain and examine nucleolar puncta at high magnification; avoid scoring diffuse chromogen over nuclei as nucleolar staining (standard IHC practice). Compare matched sections retrieved in 10 mM citrate, pH 6.0 and processed without primary antibody, using the same exposure to chromogen (datasheet A05730; standard IHC practice).

Can this stain distinguish RPS24 isoforms or a modified epitope?

RPS24 has 4 annotated isoforms and reported N-terminal acetylation and Thr-9 phosphorylation, but the supplied antibody evidence does not map its epitope (UniProt P62847; datasheet A05730). Consequently, chromogenic staining cannot be assigned to one isoform or modification from this evidence alone (UniProt P62847; datasheet A05730). Keep the catalog condition of 10 mM citrate, pH 6.0 and the reported 1:200 dilution as the starting point when comparing sections (datasheet A05730). If an isoform-specific conclusion is essential, require independently validated epitope information and an appropriate perturbation or orthogonal assay before interpreting differences in staining intensity (standard IHC practice).

How can I cross-check RPS24 localisation by multiplex IF?

Use IF as a separate validation experiment: pair RPS24 with a marker of the cell population being evaluated, such as a marker for Purkinje cells in cerebellum (HPA tissue IHC; standard IF practice). Select spectrally separated fluorophores and place the weaker signal in a channel with low tissue autofluorescence, using unstained and single-label controls to assess bleed-through (standard IF practice). Because the antibody epitope is unspecified, test permeabilisation conditions that provide access to intracellular cytosolic, endoplasmic-reticulum-associated, or nucleolar pools without assuming which side of a membrane contains the epitope (datasheet A05730; HPA subcellular; UniProt P62847). Evaluate colocalisation at the cell level rather than transferring the paraffin-section 1:200 dilution directly to IF (datasheet A05730; standard IF practice).

What should I check when RPS24 DAB staining looks widespread?

First compare a no-primary section and inspect whether color follows tissue edges, folds, debris, or endogenous pigment rather than intact cells (standard IHC practice). Check the peroxidase block and DAB development time, since both are general chromogenic workflow variables, and process controls alongside sections retrieved in 10 mM citrate, pH 6.0 (standard IHC practice; datasheet A05730). Titrate the antibody around the reported 1:200 condition and assess whether lower background preserves cellular signal (datasheet A05730; standard IHC practice). Widespread cytoplasmic staining can be plausible for RPS24, but HPA's low consistency between antibody staining and RNA expression makes morphology and controls especially important (HPA tissue IHC).

How should I score RPS24 staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and compartment before scoring, because HPA describes cytoplasmic tissue staining while UniProt also places RPS24 in the nucleolus (HPA tissue IHC; UniProt P62847). For each region, report the percentage of positive intact cells and an H-score using intensity grades 0–3, or count positive cells per mm² when cell density is the main question (standard IHC practice). Normalize counts to the number of evaluable cells or measured viable tissue area, and use identical thresholds and chromogen development across runs (standard IHC practice). Record retrieval and dilution, starting from 10 mM citrate, pH 6.0 and 1:200, so processing differences remain visible during comparison (datasheet A05730).

When is an apparent RPS24-positive cell likely an artefact?

Treat a well-defined cellular signal as more persuasive when its compartment fits cytoplasm or nucleolus, the reported RPS24 locations (HPA tissue IHC; UniProt P62847). Compare cell identity with the tissue pattern: HPA reports medium staining in cerebellar Purkinje cells but no detected staining in heart-muscle cardiomyocytes, with acknowledged low consistency against RNA expression (HPA tissue IHC). Distrust color restricted to cut edges, folds, necrotic regions, or a no-primary control; check endogenous peroxidase and pigment before calling those areas positive (standard IHC practice). Confirm uncertain calls on matched sections under the reported 10 mM citrate, pH 6.0 retrieval, keeping DAB development and scoring thresholds consistent (datasheet A05730; standard IHC practice).

Boster reagents

Best RPS24 / Small ribosomal subunit protein eS24 IHC Antibodies

A05730 has real IHC data from paraffin-embedded rat brain (catalog IHC image caption). Human, mouse, and rat reactivity is listed (catalog reactivity); no IF data are shown (catalog IF images).

Real IHC data Immunohistochemistry of paraffin-embedded rat brain using RPS24 Rabbit pAb at dilution of 1:200 .Perform high pressure antigen retrieval with 10 mM citrate buffer pH 6.0 before commencing with IHC staining protocol.

Anti-40S ribosomal protein S24 RPS24 Antibody

Cat # A05730

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A05730 is listed for IHC and has an image of paraffin-embedded rat brain stained at 1:200 after citrate retrieval (catalog applications; catalog IHC image caption). Human, mouse, and rat are listed as reactive species, while the image documents rat tissue only (catalog reactivity; catalog IHC image caption).

Which to pick: For tissue IHC, choose A05730: it is a rabbit polyclonal antibody listed for IHC at 1:50–1:200, with a paraffin-embedded rat brain example (catalog host and dilution; catalog IHC image caption). Its listed human, mouse, and rat reactivity makes it the cross-species candidate, though the supplied IHC image shows rat tissue only (catalog reactivity; catalog IHC image caption). No IF/ICC application or image is supplied for A05730, so an IF/ICC choice cannot be supported here; the IHC caption does not report the fixative (catalog applications; catalog IF images; catalog IHC image caption).

Each figure is that product's own IHC / IF validation image from its datasheet.

References

UniProt Consortium. UniProt entry P62847 (RS24_HUMAN, Small ribosomal subunit protein eS24).
Human Protein Atlas. RPS24 tissue IHC expression (reliability: Approved).
Human Protein Atlas. RPS24 subcellular location (ICC-IF): Localized to the endoplasmic reticulum and cytosol..
Human Protein Atlas. RPS24 antibody validation summary (2 antibodies).
RPS24 Is Associated with a Poor Prognosis and Immune Infiltration in Hepatocellular Carcinoma. International journal of molecular sciences 2023 — PMC9820840.
Development of Soft Tissue Sarcomas in Ribosomal Proteins L5 and S24 Heterozygous Mice. Journal of Cancer 2016 — PMC4679378.
Expression of ribosomal proteins in normal and cancerous human prostate tissue. PloS one 2017 — PMC5634644.
Single-cell multi-omics elucidates the role of RPS27-RPS24 fusion gene in osteosarcoma chemoresistance and metabolic regulation. Cell death discovery 2025 — PMC12032165.
PubMed PMID:2210388 — UniProt-cited evidence.
PubMed PMID:8647458 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.