PGM3 / Phosphoacetylglucosamine mutase · IHC design guide

Design Immunohistochemistry for PGM3

Plan PGM3 chromogenic IHC using the observed cytoplasmic and nuclear tissue pattern (HPA tissue IHC). Compare strongly stained prostate glandular cells with an appropriate negative control, and interpret lung macrophage staining in light of the differing tissue reports (HPA tissue IHC; UniProt).

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

Immunohistochemistry protocol sheet for PGM3 (IHC for PGM3): expected localisation Cytoplasmic and nuclear staining (HPA tissue IHC), antibody A02052-1, validated IHC image, and IHC protocol steps — Printable **PGM3** IHC protocol sheet — expected localisation Cytoplasmic and nuclear staining (HPA tissue IHC), antibody A02052-1, controls and protocol steps. Open the full PGM3 IHC guide →

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PGM3 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	Cytoplasmic and nuclear staining (HPA tissue IHC)
	Staining pattern	High in prostate glandular cells; cytoplasmic and nuclear overall (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet A02052-1)
	Positive control ⓘ	Cerebellum+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	Lung macrophages stain despite the reported absence in lung (HPA tissue IHC; UniProt)
	Regulation	Specific expression regulation unreported (UniProt)
	Isoform / epitope	Three isoforms; epitope differences unreported (UniProt)

Section 1

Recommended PGM3 IHC & IF Protocols

The catalog antibody protocol uses EDTA pH 8.0 heat retrieval (datasheet A02052-1). One published brain tumor IHC protocol provides a citrate retrieval comparison (PMC12007565).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human testicular seminoma tissue; fixative not specified (datasheet A02052-1)
Fixation	Image fixative and duration unreported (datasheet A02052-1); 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 A02052-1); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet A02052-1)
Primary antibody	Rabbit anti-PGM3, 2-5 μg/ml (datasheet A02052-1)
Primary incubation	Overnight at 4 °C (datasheet A02052-1)
Detection	HRP-conjugated secondary, DAB chromogen (datasheet A02052-1)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	PGM3-positive staining in processes in granular layer of cerebellum (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and nuclear expression in several tissues, most abundant in prostate and placenta. No signal in the no-primary control.

💡Decision noteStart with EDTA pH 8.0 heat retrieval (datasheet A02052-1); consider citrate pH 6.0 if optimizing against the published brain tumor protocol (PMC12007565).

Section 2

What Is the Expected PGM3 Staining Pattern?

PGM3 staining should be assessed in both cytoplasm and nucleus, with the clearest tissue examples in prostate glandular cells, duodenal Brunner glands, small intestinal Paneth cells and cerebellar granular-layer processes (HPA tissue IHC). PGM3 has no transmembrane segment (UniProt O95394 topology). HPA rates its tissue IHC profile Enhanced, while reporting medium agreement with RNA data and pending external verification (HPA tissue IHC).

What am I looking at on my slide?
Cytoplasmic and nuclear staining appears in prostate glandular cells, with a tissue pattern that follows the gland boundaries (HPA tissue IHC).	This fits HPA's reported cytoplasmic and nuclear expression and high staining in prostate glandular cells (HPA tissue IHC). Judge the relevant cells and compartments together; staining intensity alone does not establish specificity (general IHC practice).
A slide shows only sharp membrane outlines, without convincing cytoplasmic or nuclear staining.	Membrane-only staining does not match the reported tissue pattern or the approved nucleoplasm and cytosol locations (HPA tissue IHC; HPA subcellular). Treat it as a possible artefact and review controls before calling the tissue positive (general IHC practice).
A supposedly negative cell population stains strongly, while the expected positive cells do not.	For example, adipocytes in adipose tissue are reported as not detected (HPA tissue IHC). Staining there may reflect cross-reactivity or endogenous detection activity; compare a matched control and the expected cell population (general IHC practice).
Brown reaction product spreads evenly across cells, stroma and blank areas of the section.	This diffuse background cannot be assigned to the cell-specific PGM3 pattern reported by HPA (HPA tissue IHC). Review blocking, detection reagent exposure and wash stringency, and check a control lacking primary antibody (general IHC practice).
A prostate section shows no signal in glandular cells, even though the tissue is intact.	Prostate glandular cells are a reported high staining population (HPA tissue IHC). First check section quality, antibody and detection controls, and the documented retrieval conditions; absence of staining alone does not prove that PGM3 is absent (general IHC practice).

💡Expected PGM3 appearanceCall the result positive when cytoplasmic and nuclear staining marks the appropriate cells, such as high staining in prostate glandular cells; membrane-only staining or uniform background is suspect (HPA tissue IHC; HPA subcellular; general IHC practice).

How each factor affects the staining
Cell population and tissue	Expression is cell dependent: Paneth cells and Brunner glands stain high, whereas liver hepatocytes stain low and adipose adipocytes are not detected (HPA tissue IHC). Score the named cells, rather than assigning one intensity to an entire organ (general IHC practice).
Lung interpretation	UniProt describes PGM3 as found in many tissues except lung (UniProt O95394 tissue specificity), while HPA reports medium staining in lung macrophages (HPA tissue IHC). Record the stained cell type and source; the two statements should not be treated as identical measurements.
Antibody evidence	Two listed antibodies have Enhanced IHC validation and one has Supported IHC validation (HPA antibodies). HPA's overall tissue profile still has medium agreement with RNA data and awaits external verification (HPA tissue IHC); interpret unexpected staining with appropriate controls (general IHC practice).
Isoforms and epitope coverage	UniProt lists 3 PGM3 isoforms (UniProt O95394 isoforms). The supplied records do not identify the IHC antibody epitope or establish which isoforms it detects; do not infer isoform-specific staining from a positive section.
Fixation sensitivity	Target-specific effects of fixation or antigen retrieval on PGM3 staining are unreported in the supplied UniProt and HPA records. Document the conditions used when comparing sections (general IHC practice).
IF/ICC Q&A: where should signal appear?	In IF/ICC, look for nucleoplasm and cytosol signal; both locations are approved, with images listed for A-431, U-251MG and U2OS (HPA subcellular). This localization answer does not establish an IF/ICC protocol or transfer IHC antibody validation to IF/ICC (HPA antibodies).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
The expected prostate glandular-cell signal is absent (HPA tissue IHC).	An assay or section problem is possible; HPA's high prostate observation does not identify a particular technical failure (HPA tissue IHC).	Check tissue integrity and positive and detection controls; review the antibody conditions and retrieval record before interpreting the negative result (general IHC practice).
Only nuclei or only cytoplasm stain in an otherwise positive tissue.	The result incompletely matches the reported cytoplasmic and nuclear tissue pattern (HPA tissue IHC); relative signal can vary by cell and assay (general IHC practice).	Compare the named positive cells with a matched control, then review counterstain and chromogen development before calling the compartment restriction biological (general IHC practice).
Membrane rims dominate the staining.	A membrane-only pattern conflicts with approved nucleoplasm and cytosol localization and with the absence of a transmembrane segment (HPA subcellular; UniProt O95394 topology).	Inspect background and detection controls and reassess localization in the expected cell population before scoring the membrane signal as PGM3 (general IHC practice).
Brown signal appears broadly in tissue and in the no-primary control.	Signal in a control lacking primary antibody suggests endogenous detection activity or nonspecific detection reagent staining (general IHC practice).	Review the detection system's endogenous activity block, blocking steps and washes; compare the corrected control with the cell-specific HPA pattern (general IHC practice; HPA tissue IHC).
Adipocytes stain strongly, but nearby cells provide no convincing expected pattern.	Adipose adipocytes are reported as not detected; strong staining there raises concern for nonspecific signal (HPA tissue IHC).	Compare a matched no-primary control and a known-positive tissue, then reassess antibody conditions before accepting the adipocyte signal (general IHC practice; HPA tissue IHC).
A lung section gives an apparently conflicting result.	UniProt's tissue statement excludes lung, whereas HPA reports medium lung macrophage staining (UniProt O95394 tissue specificity; HPA tissue IHC).	Record whether macrophages specifically stain and report which source supports the interpretation; use matched controls to investigate unexpected staining (HPA tissue IHC; general IHC practice).

Source: UniProt O95394 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing PGM3 staining

IHC analysis of PGM3 using anti-PGM3 antibody (A02052-1). PGM3 was detected in a paraffin-embedded section of human testicular seminoma 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 2 μg/ml rabbit anti-PGM3 Antibody (A02052-1) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.

A02052-1 · IHCIHC analysis of PGM3 using anti-PGM3 antibody (A02052-1). PGM3 was detected in a paraffin-embedded section of human testicular seminoma tiss

IF analysis of PGM3 using anti-PGM3 antibody (A02052-1) and anti-Beta Tubulin antibody (M01857-3). PGM3 was detected in immunocytochemical section of HELA cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL rabbit anti-PGM3 Antibody (A02052-1) and mouse anti-Beta Tubulin antibody (M01857-3) overnight at 4°C. Cy3 Conjugated Goat Anti-Rabbit IgG (BA1032) and DyLight®488 Conjugated Goat Anti-Mouse IgG (BA1126) were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

A02052-1 · IFIF analysis of PGM3 using anti-PGM3 antibody (A02052-1) and anti-Beta Tubulin antibody (M01857-3). PGM3 was detected in immunocytochemical s

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000013375-PGM3/tissue/cerebral+cortex#imid_7725493

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000013375-PGM3/tissue/colon#imid_7725425

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000013375-PGM3/tissue/liver#imid_7725418

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000013375-PGM3/tissue/kidney#imid_7725427

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA029760; green = PGM3, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for PGM3 IHC & IF

🧪Run small intestine first: Paneth cells should stain strongly (HPA: High in Paneth cells). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the positive slide, nearby non-Paneth epithelial cells should be assessed for background-level staining, although their PGM3 negativity is not established by the supplied HPA rows.

Positive control tissue: Cerebellum (Processes in granular layer, HPA High)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show PGM3 in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (approved), Cytosol (approved) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only control, a host-matched rabbit isotype control, and PGM3 knockout material or a peptide-block control (caption: rabbit primary antibody; standard IHC practice). Quench endogenous peroxidase in the intestinal section, and check endogenous biotin if using avidin–biotin detection (standard IHC practice).

⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A02052-1 paraffin-section caption does not state a fixative (selected-SKU caption: fixative unreported). Paraffin IHC has a documented starting condition—heat retrieval in EDTA at pH 8.0—but retrieval dependence has not been established (selected-SKU caption: EDTA retrieval). There is no supplied comparison showing that frozen sections or IF are easier; intestinal background from endogenous peroxidase warrants checking when scoring Paneth-cell staining (standard IHC practice).

HPA tissue IHC evidence for PGM3

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

Positive expression · recommended positive controls

Tissue	Cell type	Level	Evidence	Source
Cerebellum	Processes in granular layer	High	Protein (IHC)	HPA →
Duodenum	Glands of Brunner	High	Protein (IHC)	HPA →
Prostate	Glandular cells	High	Protein (IHC)	HPA →
Small intestine	Paneth cells	High	Protein (IHC)	HPA →
Lung	Macrophages	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 →
Breast	Adipocytes	Not detected	Protein (IHC)	HPA →
Bronchus	Respiratory epithelial cells	Not detected	Protein (IHC)	HPA →
Caudate	Glial cells	Not detected	Protein (IHC)	HPA →

Section 3

Advanced PGM3 IHC Tips

Troubleshoot PGM3 staining by checking retrieval, cell type, and subcellular pattern before interpreting chromogenic signal.

How should I retrieve PGM3 antigen when paraffin sections stain weakly?

Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet A02052-1). The selected paraffin-section example used 2 μg/ml catalog antibody overnight at 4°C, so compare weak runs against those documented incubation conditions before changing retrieval (datasheet A02052-1). If staining remains weak, test a different retrieval duration on adjacent sections while keeping antibody concentration, detection, and development consistent (standard IHC practice). Include a known staining compartment and a no-primary control in each comparison, because stronger DAB alone cannot distinguish recovered PGM3 signal from increased background (HPA: cytoplasmic and nuclear tissue expression; standard IHC practice).

Can fixation explain inconsistent PGM3 staining across paraffin blocks?

The selected paraffin-section caption does not report a fixative, so PGM3-specific fixation sensitivity is unknown (datasheet A02052-1). Record the fixative, fixation interval, and processing history for each block, then compare sections processed alike before attributing differences to biology (standard IHC practice). Keep retrieval at EDTA pH 8.0 and the antibody incubation at 2 μg/ml overnight at 4°C when benchmarking against the documented example (datasheet A02052-1). If blocks still differ, use matched internal control cells and a no-primary section to separate variable antigen preservation from nonspecific chromogen deposition (standard IHC practice).

Should PGM3 stain nuclei, cytoplasm, or both in tissue sections?

Assess both nuclear and cytoplasmic DAB signal: tissue IHC reports expression in both compartments, while cell imaging places PGM3 in the nucleoplasm and cytosol (HPA: tissue IHC profile; HPA: subcellular localisation). PGM3 has no annotated transmembrane segment, so a crisp membrane-only pattern warrants scrutiny rather than immediate acceptance as its expected distribution (UniProt O95394 topology). Evaluate staining within identified cells, since prostate glandular cells are reported high and lung macrophages medium despite different surrounding tissue patterns (HPA: prostate glandular cells High; HPA: lung macrophages Medium). Compare compartment scores with no-primary sections and tissue morphology before assigning a nuclear-to-cytoplasmic shift (standard IHC practice).

Could PGM3 isoforms or modified residues alter tissue staining?

PGM3 has 3 annotated isoforms and modified residues including phosphothreonine 62 and phosphoserine 64 (UniProt O95394 isoforms and modified residues). The supplied caption gives no antibody epitope or isoform coverage, so this assay cannot assign a staining difference to one isoform or modification (datasheet A02052-1: epitope not supplied). Compare serial sections using the same EDTA pH 8.0 retrieval and 2 μg/ml antibody conditions before investigating epitope accessibility (datasheet A02052-1; standard IHC practice). If isoform-specific interpretation matters, obtain epitope mapping or orthogonal isoform evidence and keep the chromogenic result labeled as total antibody-reactive PGM3 until specificity is established (standard IHC practice).

How can IF help resolve an ambiguous PGM3 IHC pattern?

Use the companion IF/ICC assay to examine whether signal occupies nucleoplasm and cytosol, the approved cellular locations (HPA: subcellular localisation). For multiplexing, pair PGM3 with a validated marker of the cell type being assessed, such as the glandular population when investigating prostate IHC, and choose fluorophores in channels with low measured tissue autofluorescence (HPA: prostate glandular cells High; standard IF practice). Because PGM3 lacks an annotated transmembrane segment, permeabilise cells when testing access to its cytosolic or nucleoplasmic signal, then verify the paired marker survives that treatment (UniProt O95394 topology; HPA: subcellular localisation; standard IF practice). Compare single-label and no-primary controls before using IF colocalisation to explain a DAB pattern (standard IF practice).

What should I change when PGM3 DAB staining appears diffuse?

Inspect a no-primary section and shorten chromogen development if diffuse color also appears without the catalog antibody (standard IHC practice). The documented tissue example used 10% goat serum block, 2 μg/ml primary overnight at 4°C, and peroxidase-conjugated secondary for 30 minutes at 37°C (datasheet A02052-1). Apply a peroxidase block, adequate washes, and a matched isotype or other specificity control as appropriate to the detection system; these are general troubleshooting steps, not evidence of a PGM3-specific requirement (standard IHC practice). Judge improvements against expected nuclear and cytoplasmic cell staining rather than overall tissue darkness (HPA: tissue IHC profile; standard IHC practice).

How should I score heterogeneous PGM3 staining in paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and nuclear or cytoplasmic compartment before scoring, because PGM3 can appear in both (HPA: tissue IHC profile). For each compartment, report the percentage of positive cells and an intensity-based H-score using a fixed threshold and the same DAB development settings across slides (standard IHC quantification practice). If counting discrete positive cells, report density per mm² of viable tissue and normalise glandular-cell comparisons to the number or area of eligible glandular cells (standard IHC quantification practice). Record the region sampled and exclude folds, edges, and necrotic areas consistently; report compartment scores separately so a shift in tissue composition does not masquerade as changed PGM3 expression (standard IHC quantification practice).

When is a strong PGM3 stain likely to be artefactual?

A credible result identifies stained cells and shows plausible nuclear or cytoplasmic signal, matching reported tissue and cellular distributions (HPA: tissue IHC profile; HPA: subcellular localisation). Scrutinise membrane-only staining because PGM3 has no annotated transmembrane segment, and check whether a supposed positive belongs to the expected population rather than adjacent unstained cells (UniProt O95394 topology; standard IHC interpretation practice). Signal confined to section edges or necrotic areas, or present on a no-primary slide, supports processing or detection artefact (standard IHC practice). Use peroxidase blocking and a no-primary control to assess endogenous enzyme activity before interpreting DAB deposits as PGM3 (standard IHC practice).

Boster reagents

Best PGM3 / Phosphoacetylglucosamine mutase IHC Antibodies

A02052-1 has documented IHC on human paraffin sections and IF on HeLa cells and rat paraffin sections (catalog image captions). Human, mouse, and rat reactivity is listed (catalog: A02052-1).

Real IHC data

Anti-PGM3 Antibody ®

Cat # A02052-1

View product →

A02052-1 will render with IHC data from a human testicular seminoma paraffin section (IHC image caption: A02052-1). Its IF images show HeLa cells and a rat brain paraffin section; the catalog lists ICC and IF applications and human, mouse, and rat reactivity (IF image captions and catalog: A02052-1).

Which to pick: For tissue IHC, choose A02052-1: its caption documents EDTA retrieval at pH 8.0 and 2 μg/ml antibody on a human paraffin section; the fixative is unreported (IHC image caption: A02052-1). For IF/ICC, the same SKU has HeLa cell and rat brain section images at 5 μg/ml (IF image captions: A02052-1). For cross-species work, A02052-1 lists human, mouse, and rat reactivity, although the supplied IHC image documents human tissue only (catalog and IHC image caption: A02052-1).

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

References

UniProt Consortium. UniProt entry O95394 (AGM1_HUMAN, Phosphoacetylglucosamine mutase).
Human Protein Atlas. PGM3 tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. PGM3 subcellular location (ICC-IF): Localized to the nucleoplasm and cytosol..
Human Protein Atlas. PGM3 antibody validation summary (3 antibodies).
PSMD11 stabilizes PGM3 by antagonizing Parkin to promote bladder cancer progression through energy metabolism reprogramming. Cell death & disease 2026 — PMC13184242.
Targeting PGM3 abolishes SREBP-1 activation-hexosamine synthesis feedback regulation to effectively suppress brain tumor growth. Science advances 2025 — PMC12007565.
PubMed PMID:10721701 — UniProt-cited evidence.
PubMed PMID:11004509 — UniProt-cited evidence.
PubMed PMID:14702039 — UniProt-cited evidence.