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APIP / Methylthioribulose-1-phosphate dehydratase · IHC design guide
Design Immunohistochemistry for APIP
Plan APIP paraffin IHC with colon glandular cells as a positive reference and cerebellar granular layer cells as a negative reference (HPA tissue IHC). The catalog antibody lists an IHC dilution of 1:50–1:200 (datasheet), and tissue staining can be nuclear and cytoplasmic at variable levels (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
APIP 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 | Nuclear and cytoplasmic tissue staining (HPA tissue IHC) | |
| Staining pattern | Variable nuclear and cytoplasmic staining; high in glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Cerebellum+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | Isoform 1 is ubiquitous; isoform 2 is lower (UniProt) | |
| Isoform / epitope | 2 isoforms; epitope coverage needs verification (UniProt) |
Section 1
Recommended APIP IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet) is accompanied by one published APIP protocol for HCC tissue sections (PMC12541992).
| Sample | Paraffin-embedded human colon tissue; fixative not specified (datasheet A09029) |
| Fixation | Image fixative and duration unreported (datasheet A09029); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| 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-APIP, 1:50-1:200 (datasheet A09029) |
| 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 | APIP-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Nuclear and cytoplasmic expression at variable levels in several different tissue types. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval rule); the published protocol uses high-pressure citrate retrieval (PMC12541992).
Section 2
What Is the Expected APIP Staining Pattern?
APIP should stain glandular cells in several tissues, including colon and stomach, and exocrine glandular cells in pancreas (HPA tissue IHC: High). Nuclear and cytoplasmic staining is reported across tissues (HPA tissue IHC: Approved; medium consistency), while UniProt lists cytoplasmic localisation and no transmembrane segment (UniProt Q96GX9). Interpret intensity by cell type and compartment, since HPA reports variable tissue expression (HPA tissue IHC).
| Glandular cells show nuclear and cytoplasmic chromogen, especially in colon or stomach (HPA tissue IHC: High). | This fits the reported tissue pattern (HPA tissue IHC: nuclear and cytoplasmic). Score the glandular cells rather than treating every cell in the section as an expected positive; HPA assigns staining levels to specified cell types (HPA tissue IHC). |
| The signal is mainly nuclear, with less obvious cytoplasmic staining. | Do not reject it solely for nuclear enrichment: tissue IHC reports both compartments (HPA tissue IHC), and ICC-IF places APIP mainly in the nucleoplasm with additional cytosol (HPA subcellular: approved). UniProt lists cytoplasm (UniProt Q96GX9), so record the compartment difference. |
| Staining is confined to cell borders or extracellular material. | That pattern does not match reported nuclear and cytoplasmic staining (HPA tissue IHC) or the lack of a transmembrane segment (UniProt Q96GX9 topology). Treat it as suspect and compare with controls before attributing it to APIP (general IHC practice). |
| Strong deposit appears in a cell type reported as not detected, such as ovarian stroma cells (HPA tissue IHC). | Investigate cross-reactivity or endogenous detection activity (general IHC practice). The HPA call applies to ovarian stroma cells, not every cell in an ovary section (HPA tissue IHC: Not detected); a tissue-wide negative conclusion would exceed that evidence. |
| Haze covers the section, or known-positive colon glandular cells show no signal (HPA tissue IHC: High). | Diffuse haze prevents compartment scoring and suggests background (general IHC practice). Absent signal in a specified high-staining cell type raises an assay-performance question; HPA's medium staining–RNA consistency does not guarantee every specimen will stain (HPA tissue IHC). |
💡Expected APIP appearanceA convincing positive shows nuclear and cytoplasmic staining in glandular cells, with high staining reported in colon and stomach and in pancreatic exocrine glandular cells (HPA tissue IHC: High); border-only or diffuse section-wide deposit is suspect (UniProt Q96GX9 topology; general IHC practice).
| Compartment evidence | Tissue IHC reports nuclear and cytoplasmic expression (HPA tissue IHC); ICC-IF favours nucleoplasm plus cytosol (HPA subcellular: approved), whereas UniProt lists cytoplasm (UniProt Q96GX9). Preserve that distinction when scoring nuclear signal. |
| Cell type and tissue choice | Colon glandular and pancreatic exocrine glandular cells are high-staining comparators (HPA tissue IHC: High). Cerebellar granular-layer cells and ovarian stroma cells are reported not detected (HPA tissue IHC); these calls are cell-specific. |
| Antibody evidence | Both listed antibodies have Approved IHC status (HPA antibodies: HPA021188, HPA024131). The tissue profile is Approved with medium staining–RNA consistency (HPA tissue IHC); these ratings support comparison but do not establish specimen-level certainty. |
| Isoforms and processing | Isoform 1 is ubiquitous; isoform 2 is lower and detected in listed organs including pancreas (UniProt Q96GX9). No signal peptide, propeptide, or cleaved chain is listed (UniProt Q96GX9); the supplied record does not map antibody epitopes to isoforms. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in colon glandular cells reported High (HPA tissue IHC). | The expected comparator failed; the record does not identify an APIP-specific fixation or retrieval failure (HPA tissue IHC; UniProt Q96GX9). | Check the IHC-validated antibody's documented IHC-P conditions, detection reagents, and a positive control; adjust retrieval only as a general IHC optimisation step (general IHC practice). |
| Strong staining appears only at membranes or outside cells. | This conflicts with nuclear and cytoplasmic staining (HPA tissue IHC) and a protein without a transmembrane segment (UniProt Q96GX9 topology). | Inspect morphology, compare a no-primary control, and reassess the detection conditions before scoring this deposit as APIP (general IHC practice). |
| The entire section has diffuse chromogen. | Background can obscure cell boundaries and compartments (general IHC practice); it cannot be validated by HPA's cell-specific high-staining calls (HPA tissue IHC). | Review blocking, washes, antibody dilution, and chromogen development using appropriate controls (general IHC practice). |
| A reported not-detected cell type stains strongly, such as cervix glandular cells (HPA tissue IHC). | Cross-reactivity or endogenous detection activity is possible (general IHC practice); the HPA result is specific to those glandular cells (HPA tissue IHC: Not detected). | Compare matched cell types and no-primary controls; check whether deposit follows cell morphology before calling it APIP (general IHC practice). |
| Staining is nuclear, while the UniProt location says cytoplasm (UniProt Q96GX9). | The sources differ in emphasis: HPA tissue IHC includes nuclear signal, and HPA ICC-IF places APIP mainly in the nucleoplasm (HPA subcellular: approved). | Record nuclear and cytoplasmic scores separately; assess cell type and controls before classifying nuclear staining as artefact (HPA tissue IHC; general IHC practice). |
| Does IF/ICC need the same interpretation as paraffin-section IHC? | HPA ICC-IF reports nucleoplasm and cytosol (HPA subcellular: approved), while HPA021188 is IHC Approved but ICC Uncertain (HPA antibodies). | Use the separate IF/ICC guide for that application; compare its localisation evidence without treating the IHC approval as ICC validation (HPA antibodies; HPA subcellular). |
Experiment evidence
IHC & IF images showing APIP staining
A09029 · IHCImmunohistochemistry of paraffin-embedded human colon using APIP Rabbit pAb at dilution of 1:100 .Perform microwave antigen retrieval with 1
A09029 · IHCImmunohistochemistry of paraffin-embedded mouse liver using APIP Rabbit pAb at dilution of 1:100 .Perform microwave antigen retrieval with 1
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000149089-APIP/tissue/cerebral+cortex#imid_7919832
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000149089-APIP/tissue/colon#imid_7917246
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000149089-APIP/tissue/liver#imid_7917247
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000149089-APIP/tissue/kidney#imid_7917256
HPA · ICC-IF PC-3Human Protein Atlas immunofluorescence, PC-3, antibody HPA021188; green = APIP, blue = nucleus, red = microtubules.
HPA · ICC-IF PC-3Human Protein Atlas immunofluorescence, PC-3, antibody HPA021188; green = APIP, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for APIP IHC & IF
🧪Run colon first: its glandular cells should stain strongly (HPA: High in colon glandular cells; selected A09029 tissue-IHC caption: human colon). Run cervix glandular cells as a negative comparison (HPA: Not detected in cervix glandular cells); on the colon slide, assess cells outside the glandular compartment as internal background comparators without assuming they are APIP-negative (HPA: colon result specifies glandular cells only).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Cerebellum (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show APIP in PC-3, Rh30, U2OS, with annotated localisation: Nucleoplasm (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), matched rabbit polyclonal IgG isotype, and APIP knockout tissue or a peptide-block control if the immunizing peptide is available (selected A09029 caption: rabbit pAb; standard IHC practice). For chromogenic colon IHC, quench endogenous peroxidase and check inflammatory cells for residual signal (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported in the supplied evidence; the exact selected-SKU colon caption does not state a fixative, so its paraffin section does not establish fixation (selected A09029 tissue-IHC caption). That caption specifies microwave antigen retrieval in 10 mM PBS, pH 7.2, followed by antibody at 1:100 (selected A09029 tissue-IHC caption). Whether frozen sections or IF are easier is unreported; in colon chromogenic IHC, endogenous peroxidase in inflammatory cells can complicate scoring (standard IHC practice). The selected A09029 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image A09029).
HPA tissue IHC evidence for APIP
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Pancreas | Exocrine glandular cells | High | Protein (IHC) | HPA → |
| Prostate | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced APIP IHC Tips
Troubleshoot APIP staining in paraffin sections by checking retrieval, cell type, subcellular pattern, and controls before comparing samples (IHC workflow guidance).
Which retrieval conditions should I try first for APIP in paraffin sections?
Start with citrate buffer at pH 6.0 for heat-induced retrieval at 95–98 °C for 20 min (page retrieval rule). If staining is weak, compare adjacent sections with microwave retrieval in 10 mM PBS, pH 7.2, the condition reported for catalog antibody A09029 in paraffin-embedded human colon (A09029 tissue-IHC caption). Keep antibody dilution, detection, and development time constant so the retrieval comparison is interpretable (standard IHC practice). Inspect glandular-cell staining alongside morphology and background: colon glandular cells are reported as high for APIP, but the caption does not state a fixative (HPA tissue IHC; A09029 tissue-IHC caption).
How should I troubleshoot inconsistent APIP staining across paraffin blocks?
Target-specific sensitivity of APIP to fixation is unknown because no fixation comparison is supplied, and the A09029 paraffin-section caption does not state a fixative (evidence supplied; A09029 tissue-IHC caption). Record each block’s fixative, fixation duration, processing history, and section age before comparing staining intensity (standard IHC practice). Run sections from the blocks together with the same pH 6.0 retrieval, antibody dilution, and chromogen development time to limit technical variation (page retrieval rule; standard IHC practice). If one block remains weak, compare its morphology and internal glandular cells with a separately processed positive control, without attributing the difference to a specific fixation effect (HPA tissue IHC; standard IHC practice).
Should APIP staining appear in nuclei, cytoplasm, or both?
Evaluate nuclear and cytoplasmic staining separately: tissue IHC reports variable staining in both compartments, while UniProt annotates APIP as cytoplasmic (HPA tissue IHC; UniProt Q96GX9 subcellular). Cell-based IF also reports nucleoplasm as the main location and cytosol as an additional location, so nuclear signal alone is not grounds for rejection (HPA subcellular). Confirm that the pattern follows intact cells rather than section edges, folds, or damaged regions, and compare it with a no-primary control (standard IHC practice). APIP has no annotated transmembrane segment, so a sharply membrane-restricted pattern warrants a specificity check before scoring (UniProt Q96GX9 topology; standard IHC practice).
Can this stain distinguish APIP isoforms or reveal epitope masking?
APIP has 2 annotated isoforms, but the supplied evidence does not map the catalog antibody’s epitope or establish isoform selectivity (UniProt Q96GX9 isoforms; evidence supplied). Score the stain as APIP immunoreactivity rather than an isoform-specific readout unless independent antibody documentation or a selective control establishes that distinction (standard IHC practice). UniProt records phosphoserine at residue 87; an effect on binding is only a possibility if an epitope overlaps or depends on that site (UniProt Q96GX9 modified residues; standard immunostaining interpretation). When signal varies, compare the stated pH 6.0 retrieval with the A09029 caption’s pH 7.2 condition on adjacent sections before invoking epitope masking (page retrieval rule; A09029 tissue-IHC caption).
How can I check an ambiguous IHC pattern with multiplex IF?
Use IF as a complementary localisation check, with an independently validated glandular-cell marker in tissue where glandular APIP staining is reported (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and consider a far-red channel when tissue autofluorescence obscures shorter wavelengths; include single-label and no-primary controls (standard IF practice). APIP has no transmembrane segment and is reported in cytoplasmic, nucleoplasmic, and cytosolic compartments, so intracellular access requires permeabilisation after fixation (UniProt Q96GX9 topology and subcellular; HPA subcellular; standard IF practice). A mild starting comparison such as 0.1% detergent for 5 min can assess access, but its effect on the chosen antibody must be checked empirically (standard IF practice).
What should I check when APIP chromogenic staining looks diffuse?
First compare the slide with a no-primary control and inspect whether color follows cells or accumulates at folds, edges, and damaged areas (standard IHC practice). For peroxidase-based detection, a 3% hydrogen peroxide block for about 10 min is a general way to assess endogenous peroxidase contribution, subject to the detection system’s instructions (standard IHC practice). Titrate primary antibody and chromogen development on adjacent sections while keeping retrieval constant; A09029 was illustrated at 1:100 in paraffin-embedded colon, which is an example rather than a universal optimum (A09029 tissue-IHC caption; standard IHC practice). Check cell-specific contrast because colon glandular cells are reported as high, whereas cervical glandular cells are reported as not detected (HPA tissue IHC).
How should I quantify APIP IHC across samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scored cell population and compartment before analysis, since APIP tissue staining can be nuclear and cytoplasmic at variable levels (HPA tissue IHC). For intensity scoring, record the percentage of cells at 0, 1, 2, and 3 intensity and calculate an H-score from 0–300; retain nuclear and cytoplasmic scores separately (standard IHC scoring practice). Alternatively report percent-positive cells, or positive-cell density per mm² when cell abundance is the question, using the same threshold across slides (standard IHC scoring practice). Normalise counts to the eligible intact tissue area or target-cell count, and compare batches against the same reference section and detection settings (standard IHC practice).
How can I distinguish true APIP signal from staining artefact?
Look for reproducible staining within intact cells, with patterns evaluated against reported nuclear and cytoplasmic tissue staining and cytoplasmic UniProt annotation (HPA tissue IHC; UniProt Q96GX9 subcellular; standard IHC practice). Colon glandular-cell positivity supports a control, while an exclusively membrane-rim pattern is unexpected for a protein with no annotated transmembrane segment (HPA tissue IHC; UniProt Q96GX9 topology). Exclude staining concentrated at edges, folds, or necrotic regions, and compare with a no-primary section to identify detection background (standard IHC practice). If peroxidase detection is used, check whether residual endogenous enzyme reproduces the signal before interpreting DAB as APIP-specific (standard IHC practice).
Boster reagents
Best APIP / Methylthioribulose-1-phosphate dehydratase IHC Antibodies
The IHC-validated anti-APIP antibody A09029 has paraffin-section images from human colon and mouse liver (catalog IHC captions); IF/ICC data are unreported (catalog applications and images).
Real IHC data
Anti-APIP Antibody
Cat # A09029
A09029 is listed for IHC and reacts with human, mouse and rat samples (catalog applications and reactivity). Its IHC images show paraffin-embedded human colon and mouse liver at 1:100 (A09029 IHC captions).
Which to pick: Choose A09029 for tissue IHC in paraffin sections; its captions specify microwave retrieval in 10 mM PBS, pH 7.2, but do not report the fixative (A09029 IHC captions). For cross-species planning, the catalog lists human, mouse and rat reactivity, while the pictured IHC examples cover human and mouse only (catalog reactivity; A09029 IHC captions). No IF/ICC choice is supported by this payload because A09029 lists IHC and WB applications and has no IF images (catalog applications and images).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q96GX9 (MTNB_HUMAN, Methylthioribulose-1-phosphate dehydratase).
- Human Protein Atlas. APIP tissue IHC expression (reliability: Approved).
- Human Protein Atlas. APIP subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol..
- Human Protein Atlas. APIP antibody validation summary (2 antibodies).
- Clinical pathological significance and biological functions of APIP in hepatocellular carcinoma. Cancer cell international 2025 — PMC12541992.
- PubMed PMID:10810093 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16554811 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for APIP (Methylthioribulose-1-phosphate dehydratase). Expected localisation Nuclear and cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A09029 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Cerebellum. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colon tissue; fixative not specified (datasheet A09029); 2. Fixation: Image fixative and duration unreported (datasheet A09029); 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-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-APIP, 1:50-1:200 (datasheet A09029); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Antibody staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope coverage needs verification (UniProt); Regulation: Isoform 1 is ubiquitous; isoform 2 is lower (UniProt). Evidence: UniProt Q96GX9, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A09029, PMC12541992. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/APIP/immunohistochemistry-ihc-guide.