RRM2 / Ribonucleoside-diphosphate reductase subunit M2 · IHC design guide

Design Immunohistochemistry for RRM2

Plan RRM2 staining in paraffin sections using its observed intestinal and lymphoid tissue pattern (HPA tissue IHC). This guide covers fixation, controls and interpretation of staining that may vary with cell-cycle phase (UniProt).

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

Immunohistochemistry protocol sheet for RRM2 (IHC for RRM2): expected localisation Membranous and cytoplasmic tissue staining (HPA tissue IHC), antibody PB9817, validated IHC image, and IHC protocol steps — Printable **RRM2** IHC protocol sheet — expected localisation Membranous and cytoplasmic tissue staining (HPA tissue IHC), antibody PB9817, controls and protocol steps. Open the full RRM2 IHC guide →

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RRM2 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	Membranous and cytoplasmic tissue staining (HPA tissue IHC)
	Staining pattern	Enterocytes and germinal centres: membranous/cytoplasmic (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet PB9817)
	Positive control ⓘ	Appendix+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 conditions consistent across sections. (standard IHC practice; not target-specific)
	Caveat	Cell-cycle phase may shift cytoplasmic versus nuclear signal (UniProt)
	Regulation	Expression regulation is unreported (UniProt)
	Isoform / epitope	2 isoforms; epitope differences are unspecified (UniProt)

Section 1

Recommended RRM2 IHC & IF Protocols

The catalog antibody’s IHC-P protocol is followed by 3 published RRM2 IHC protocols, each using the conditions reported for its own tissue and antibody.

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human mammary cancer tissue; fixative not specified (datasheet PB9817)
Fixation	Image fixative and duration unreported (datasheet PB9817); 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 PB9817); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet PB9817)
Primary antibody	Rabbit anti-RRM2, 0.5-1μg/ml (datasheet PB9817)
Primary incubation	Overnight at 4 °C (datasheet PB9817)
Detection	Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9817)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	RRM2-positive staining in enterocytes of appendix (HPA tissue IHC: Medium). HPA tissue profile: Membranous and cytoplasmic expression mainly in lymphoid germinal centra and intestinal tract. No signal in the no-primary control.

💡Decision noteStart with the catalog antibody’s heat-mediated EDTA retrieval at pH 8.0 (datasheet PB9817); adjust retrieval for other antibodies using their published conditions.

Section 2

What Is the Expected RRM2 Staining Pattern?

RRM2 staining is expected mainly in the cytoplasm of intestinal enterocytes and cells in lymphoid germinal centers (HPA tissue IHC). HPA rates its tissue IHC reliability Enhanced, with medium consistency between staining and RNA expression (HPA tissue IHC). Cytoplasmic localization, with possible nuclear localization in G2, fits a protein with no transmembrane segment (UniProt P31350).

What am I looking at on my slide?
Enterocytes show moderate cytoplasmic staining, sometimes with membranous accentuation.	This fits the reported medium enterocyte staining across the sampled intestinal tissues and HPA’s cytoplasmic and membranous tissue profile (HPA tissue IHC). Judge the signal in identified cells, rather than treating every unstained neighbor as a failed reaction (general IHC practice).
Staining is concentrated in a lymphoid germinal center; nearby cells stain less.	This fits the reported emphasis on lymphoid germinal centers, although lymph node germinal center cells are listed at a low level (HPA tissue IHC). Do not require every lymphoid cell to stain equally (HPA tissue IHC).
A sharp membrane-only outline dominates, without convincing intracellular signal.	Review this as a compartment mismatch: HPA reports cytoplasmic and membranous tissue staining, while cytosol is the supported main ICC-IF location and RRM2 lacks a transmembrane segment (HPA tissue IHC; HPA ICC-IF; UniProt P31350). Membranous accentuation alone is not grounds to reject a slide (HPA tissue IHC).
Cells listed as not detected stain as strongly as the expected positive cells.	Consider cross-reactivity or endogenous detection activity, particularly if the signal also appears in reagent controls (general IHC practice). HPA lists adipocytes in adipose tissue and hematopoietic cells in bone marrow as not detected in its sampled tissue IHC (HPA tissue IHC).
A known-positive intestinal section has no discernible enterocyte signal.	Treat the run as inconclusive until a control section performs as expected (general IHC practice). HPA reports medium staining in enterocytes of appendix, colon, duodenum, rectum and small intestine; that reference pattern supports a positive-control choice but does not guarantee every specimen will stain (HPA tissue IHC).

💡Expected RRM2 appearanceCall a positive result when identified intestinal enterocytes show medium cytoplasmic staining, with possible membranous accentuation; widespread uniform staining of cells listed as not detected is a suspect positive pattern (HPA tissue IHC; general IHC practice).

How each factor affects the staining
Cell identity and tissue context	HPA reports medium staining in enterocytes of appendix, colon, duodenum, rectum and small intestine, and a profile centered on lymphoid germinal centers and the intestinal tract (HPA tissue IHC). Compare like cell types before judging intensity (general IHC practice).
Compartment and cell cycle	RRM2 is cytoplasmic in S phase and may localize to the nucleus in G2 (UniProt P31350). A nuclear component can therefore be plausible; interpret it alongside morphology and the cytoplasmic signal, without inferring cell-cycle phase from RRM2 staining alone (UniProt P31350; general IHC practice).
Topology and processing	UniProt lists no transmembrane segment, signal peptide or propeptide, and annotates a chain spanning residues 1–389 (UniProt P31350). These annotations give no basis to expect a secreted or shed staining pattern (UniProt P31350).
Antibody evidence and RNA comparison	The listed IHC antibody, HPA056994, has Enhanced validation, while overall staining-to-RNA consistency is medium (HPA antibodies; HPA tissue IHC). Bone marrow has tissue-enhanced RNA specificity yet its sampled hematopoietic cells are listed as not detected by IHC; RNA enrichment alone is not a protein-positive control (HPA tissue IHC).
Isoforms and phosphorylation	UniProt annotates 2 isoforms and phosphoserine 20 and phosphothreonine 33 (UniProt P31350). The supplied evidence does not locate the IHC antibody’s epitope or establish isoform- or phosphorylation-specific staining; do not assign a compartment pattern to either modification (UniProt P31350).
IF/ICC Q&A: where should RRM2 appear?	Mainly in the cytosol, with additional supported nucleoplasmic localization (HPA ICC-IF). This answers localization for the separate IF/ICC application; use the tissue IHC observations to assess paraffin sections (HPA tissue IHC; HPA ICC-IF).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
No staining in the intestinal positive-control section.	The staining run may have failed, or the chosen section may lack assessable enterocytes (general IHC practice). HPA reports medium enterocyte staining in several intestinal tissues, not guaranteed positivity in every specimen (HPA tissue IHC).	Confirm enterocytes on the counterstained section, inspect the same-run detection controls, and repeat with a suitable known-positive section if needed (general IHC practice). Do not score the test section negative while its positive control fails (general IHC practice).
Strong signal appears in cells expected to be negative.	Cross-reactivity or endogenous detection activity is possible (general IHC practice). HPA lists adipocytes in adipose tissue and several other sampled cell types as not detected (HPA tissue IHC).	Check morphology and compare a primary-antibody omission control; address endogenous detection activity according to the chromogenic system if the control also stains (general IHC practice). Interpret the HPA negatives at the listed cell-type level (HPA tissue IHC).
Diffuse color covers cells and surrounding tissue.	Non-specific binding, residual detection reagents or excessive chromogen development can obscure localization (general IHC practice). This appearance does not establish an RRM2 compartment (UniProt P31350; general IHC practice).	Inspect the reagent control, washing and development time; adjust blocking or reagent concentration within the validated IHC workflow as indicated by the controls (general IHC practice). Reassess cell-specific staining after background clears (general IHC practice).
Only a crisp cell-surface rim is visible.	The result may reflect non-specific membrane-associated signal, although HPA does describe membranous and cytoplasmic tissue staining (HPA tissue IHC; general IHC practice). RRM2 has no annotated transmembrane segment (UniProt P31350).	Compare the rim with intracellular staining in the same cells and with the reagent control; retain membranous accentuation only when the overall cell and tissue pattern fits the HPA observation (HPA tissue IHC; general IHC practice).
Nuclear staining appears in a subset of otherwise plausible positive cells.	Nuclear localization can occur in G2, and HPA ICC-IF supports additional nucleoplasmic localization (UniProt P31350; HPA ICC-IF). Nuclear staining alone cannot establish G2 identity (general IHC practice).	Review cytoplasmic signal, cell identity, morphology and controls before classifying the nuclear component as specific (HPA tissue IHC; general IHC practice). Avoid using nuclear intensity alone as an RRM2-positive call (UniProt P31350; general IHC practice).
Bone marrow is unstained despite its tissue-enhanced RNA designation.	HPA lists tissue-enhanced RNA specificity in bone marrow but reports hematopoietic cells as not detected in its sampled IHC (HPA tissue IHC). The two readouts need not yield the same positive-control choice (HPA tissue IHC).	Assess run performance with a section containing the HPA-reported medium-staining intestinal enterocytes, then report the bone-marrow result at the observed cell-type level (HPA tissue IHC; general IHC practice).

Source: UniProt P31350 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing RRM2 staining

IHC analysis of RRM2 using anti-RRM2 antibody (PB9817). RRM2 was detected in a paraffin-embedded section of human mammary cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 μg/ml rabbit anti-RRM2 Antibody (PB9817) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1022) with DAB as the chromogen.

PB9817 · IHCIHC analysis of RRM2 using anti-RRM2 antibody (PB9817). RRM2 was detected in a paraffin-embedded section of human mammary cancer tissue. Hea

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000171848-RRM2/tissue/cerebral+cortex#imid_15787235

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000171848-RRM2/tissue/colon#imid_15787161

HPA · RectumHuman Protein Atlas IHC, Rectum. Source: https://www.proteinatlas.org/ENSG00000171848-RRM2/tissue/rectum#imid_15787137

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000171848-RRM2/tissue/liver#imid_15787162

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA056994; green = RRM2, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for RRM2 IHC & IF

🧪Run colon first: enterocytes should show medium RRM2 staining (HPA: Colon, Enterocytes, Medium). Use adipose tissue as the negative comparator because adipocytes are not detected (HPA: Adipose tissue, Adipocytes, Not detected); on the colon slide, treat cells without enterocyte staining as background references, since HPA does not identify a specific internal negative cell type (HPA: Colon row).

Positive control tissue: Appendix (Enterocytes, HPA Medium)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RRM2 in A-431, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Cytosol (supported) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only control and a host-matched rabbit IgG isotype control matched to the catalog antibody’s clonality (caption: rabbit anti-RRM2; standard IHC practice). Use RRM2 knockout material as a biological negative where available, and block endogenous peroxidase and biotin when using the caption’s biotin-based, DAB detection system (caption: biotinylated secondary, SABC, DAB; standard IHC practice).

⚠️Feasibility: A target-specific fixation window and fixation effect are unreported, and the selected PB9817 paraffin-section caption does not state a fixative (caption: fixative not stated). The caption uses heat retrieval in EDTA, pH 8.0, but does not establish that retrieval is required; the supplied evidence also does not establish whether frozen sections or IF are easier (caption: heat-mediated EDTA retrieval; supplied application evidence). In colon, check for background from endogenous peroxidase or biotin when interpreting enterocyte staining with the caption’s SABC/DAB method (HPA: Colon, Enterocytes, Medium; caption: SABC/DAB; standard IHC practice).

HPA tissue IHC evidence for RRM2

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.

Positive expression · recommended positive controls

Tissue	Cell type	Level	Evidence	Source
Appendix	Enterocytes	Medium	Protein (IHC)	HPA →
Colon	Enterocytes	Medium	Protein (IHC)	HPA →
Duodenum	Enterocytes	Medium	Protein (IHC)	HPA →
Rectum	Enterocytes	Medium	Protein (IHC)	HPA →
Small intestine	Enterocytes	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 →
Bronchus	Respiratory epithelial cells	Not detected	Protein (IHC)	HPA →

Section 3

Advanced RRM2 IHC Tips

Use the matched paraffin-section method as the starting point, then interpret RRM2 staining against compartment, cell type, and section-level controls (datasheet PB9817; UniProt P31350; HPA tissue IHC).

What retrieval should I start with if RRM2 staining is weak in paraffin sections?

Use heat-mediated EDTA at pH 8.0 for antigen retrieval (datasheet PB9817). The matched paraffin-section image used 1 μg/ml primary antibody overnight at 4°C, so check those conditions before attributing weak signal to retrieval (PB9817 tissue-IHC caption). Run a known positive section, such as intestinal enterocytes, beside the test section and compare staining within the same cellular compartment (HPA tissue IHC; standard IHC practice). If signal remains weak, adjust heating duration on serial sections while keeping antibody concentration and detection constant, and reject conditions that increase diffuse background or tissue damage (standard IHC practice).

How should I investigate weak RRM2 staining across differently fixed specimens?

The matched image identifies a paraffin-embedded section but does not report its fixative, so RRM2-specific fixation sensitivity is unknown (PB9817 tissue-IHC caption). Record fixative, fixation duration, tissue thickness, and processing history for each specimen before comparing chromogenic intensity (standard IHC practice). Stain serial sections in one run with the same EDTA pH 8.0 retrieval and detection settings, including an established positive control (datasheet PB9817; standard IHC practice). If a specimen loses both RRM2 and control-antigen signal, investigate preservation and processing; do not infer a particular RRM2 fixation effect from its localisation, phosphorylation sites, or tissue staining pattern (standard IHC practice; UniProt P31350; HPA tissue IHC).

Should I score nuclear, cytoplasmic, or membranous RRM2 staining?

Score cytoplasmic and nuclear staining separately because RRM2 is reported in the cytoplasm and nucleus, with cytoplasmic localisation in S phase and possible nuclear localisation in G2 (UniProt P31350). Supported cytosolic and additional nucleoplasmic localisation provides an independent compartment check (HPA subcellular). HPA tissue IHC describes membranous and cytoplasmic staining, but RRM2 has no transmembrane segment, so inspect apparently membranous signal against cell boundaries and background controls before calling it specific (HPA tissue IHC; UniProt P31350 topology; standard IHC practice). Compare matched cells in positive and negative sections, and retain separate compartment scores so a bright edge pattern does not mask convincing intracellular staining (standard IHC practice).

Can this antibody distinguish RRM2 isoforms or phosphorylation-dependent staining?

RRM2 has 2 annotated isoforms and reported phosphorylation at Ser-20 and Thr-33 (UniProt P31350). The supplied tissue image establishes paraffin-section staining with PB9817 but gives no epitope map or isoform-selectivity evidence (PB9817 tissue-IHC caption). Treat its chromogenic signal as RRM2-family target staining without assigning an isoform or phosphorylation state unless the antibody’s epitope and selectivity are established independently (standard IHC interpretation). When samples differ, compare the same cellular compartment under identical retrieval and detection conditions; a change in staining alone cannot establish altered phosphorylation, epitope exposure, or isoform abundance (standard IHC practice; UniProt P31350).

How can I check RRM2 localisation by multiplex IF alongside the IHC result?

For the separate IF/ICC workflow, pair RRM2 with a marker identifying the expected cell population, such as enterocytes in an intestinal section, and inspect overlap at single-cell resolution (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and place the weaker signal in a channel with low measured tissue autofluorescence; include single-stain and no-primary controls (standard IF practice). RRM2 is cytosolic with additional nucleoplasmic localisation and has no transmembrane segment, so permeabilise the plasma membrane to reach intracellular epitopes after fixation, while recognising that this antibody’s epitope is unspecified (HPA subcellular; UniProt P31350 topology; standard IF practice). Compare compartments with the chromogenic result without treating IF intensity as an IHC score (standard assay interpretation).

What controls help distinguish diffuse DAB background from RRM2 signal?

Run a no-primary control and inspect the section for diffuse precipitate, edge staining, and staining of structures outside the expected cells (standard IHC practice). The matched method uses a biotinylated secondary, avidin-biotin detection, and DAB; assess secondary-only and endogenous-biotin contributions if background follows that detection route (PB9817 tissue-IHC caption; standard IHC practice). Include a peroxidase block and verify it with a no-primary section, since endogenous enzyme activity can create chromogenic signal unrelated to RRM2 (standard IHC practice). Titrate the primary around the documented 1 μg/ml condition only after confirming retrieval and controls, and preserve a positive section to detect lost sensitivity (PB9817 tissue-IHC caption; standard IHC practice).

How should I quantify RRM2 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and compartment before scoring, then report the percentage of positive cells and an intensity-weighted H-score separately for cytoplasm and nucleus (UniProt P31350 localisation; standard IHC scoring practice). For spatial comparisons, count positive cells per mm² within a prespecified viable region and normalise to all eligible cells or the measured viable area, as appropriate (standard IHC quantification practice). Apply the same retrieval, 1 μg/ml starting concentration, detection, imaging, and scoring thresholds across the batch (datasheet PB9817; standard IHC practice). Record tissue region and cell identity because HPA reports enterocyte staining and heterogeneous tissue patterns; pooled whole-section intensity can obscure those differences (HPA tissue IHC; standard IHC practice).

When is a strong RRM2-positive region likely to be artefactual?

Favour intracellular signal in plausible cells: RRM2 is cytoplasmic and may enter the nucleus with cell-cycle phase, while HPA reports enterocyte staining in intestinal tissues (UniProt P31350; HPA tissue IHC). Question isolated membranous outlines because RRM2 lacks a transmembrane segment, even though membranous staining appears in the HPA tissue profile (UniProt P31350 topology; HPA tissue IHC). Exclude section-edge concentration, necrotic regions, and signal reproduced in no-primary controls before interpreting a strong DAB focus (standard IHC practice). Compare the focus with a positive control and matched viable cells; endogenous peroxidase or biotin-route background should be resolved before assigning RRM2 positivity (PB9817 tissue-IHC caption; standard IHC practice).

Boster reagents

Best RRM2 / Ribonucleoside-diphosphate reductase subunit M2 IHC Antibodies

PB9817 has a human paraffin-section IHC image (PB9817 image caption). M01978 lists human IF/ICC applications, with no IF image supplied (M01978 applications; M01978 image payload).

Real IHC data

Anti-RRM2 Antibody ®

Cat # PB9817

View product →

The rendered PB9817 card shows RRM2 staining in a paraffin-embedded human mammary cancer section (PB9817 image caption). PB9817 lists IHC and human, mouse, and rat reactivity; its pictured IHC result is from human tissue (PB9817 applications/reactivity; PB9817 image caption).

Which to pick: Choose PB9817 for paraffin-section IHC: its image caption reports EDTA retrieval at pH 8.0 and 1 μg/ml primary antibody in human mammary cancer tissue; the fixative is unreported (PB9817 image caption). Choose M01978 for human IF/ICC based on its listed applications and rabbit monoclonal clone AEAE-18; no IF image is supplied (M01978 catalog payload). For cross-species work, PB9817 lists human, mouse, and rat reactivity, while its IHC image documents human tissue only (PB9817 reactivity; PB9817 image caption).

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

References

UniProt Consortium. UniProt entry P31350 (RIR2_HUMAN, Ribonucleoside-diphosphate reductase subunit M2).
Human Protein Atlas. RRM2 tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. RRM2 subcellular location (ICC-IF): Mainly localized to the cytosol. In addition localized to the nucleoplasm..
Human Protein Atlas. RRM2 antibody validation summary (1 antibodies).
Ribonucleotide reductase subunit M2 predicts survival in subgroups of patients with non-small cell lung carcinoma: effects of gender and smoking status. PloS one 2015 — PMC4441434.
Human equilibrative nucleoside transporter-1 (hENT1) and ribonucleotide reductase regulatory subunit M1 (RRM1) expression; do they have survival impact to pancreatic cancer?. Annals of hepato-biliary-pancreatic surgery 2020 — PMC7271117.
High expression of ribonucleotide reductase subunit M2 correlates with poor prognosis of hepatocellular carcinoma. Gut and liver 2014 — PMC4215454.
A Novel Predictive Model Based on Glutathione Metabolism Genes RRM2 and G6PD in Hepatocellular Carcinoma. Journal of hepatocellular carcinoma 2026 — PMC13282963.
PubMed PMID:1627826 — UniProt-cited evidence.
PubMed PMID:11978970 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.