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PIK3R3 / Phosphatidylinositol 3-kinase regulatory subunit gamma · IHC design guide
Design Immunohistochemistry for PIK3R3
Plan chromogenic IHC for PIK3R3 in paraffin sections using cytoplasmic staining as the expected tissue pattern (HPA tissue IHC). Compare positive and negative cell populations reported in tissue IHC, and interpret staining cautiously because the HPA antibody may recognize proteins from more than one gene (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
PIK3R3 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 | General cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in breast glandular cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Breast+4 more · see all | |
| Negative control | Esophagus+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep formalin fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | HPA antibody may recognize more than one gene (HPA tissue IHC) | |
| Regulation | Expression regulation unreported (UniProt) | |
| Isoform / epitope | 3 isoforms; verify epitope coverage (UniProt) |
Section 1
Recommended PIK3R3 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet) is accompanied by published PIK3R3 staining methods for colorectal tissue (PMC7735421), mouse skin (PMC5546695), and liver cancer tissue (PMC10358214).
| Sample | Paraffin-embedded Human pituitary tumor tissue; fixative not specified (datasheet M06707) |
| Fixation | Image fixative and duration unreported (datasheet M06707); 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 monoclonal (clone DHC-16) anti-PIK3R3, 1:50 recommended; image 1:200 (datasheet M06707) |
| 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 | PIK3R3-positive staining in glandular cells of breast (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. 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); one published protocol uses boiling citrate for 10 min (PMC7735421).
Section 2
What Is the Expected PIK3R3 Staining Pattern?
PIK3R3 is expected to show general cytoplasmic staining in tissue IHC, including strong staining in selected neuronal, epithelial, endocrine, and immune cell populations (HPA: tissue IHC). It has no transmembrane segment, so a membrane-only pattern is unexpected (UniProt Q92569 topology). Treat the pattern as provisional: HPA rates tissue IHC Approved but cautions that the antibody targets proteins from more than one gene and awaits external verification (HPA: tissue IHC reliability).
| Clear cytoplasmic staining in cerebral cortex or hippocampal neurons, bronchial respiratory epithelium, or pancreatic endocrine cells. | These cell populations are reported High by HPA and fit its general cytoplasmic IHC profile (HPA: tissue IHC). Compare staining within the identified cell population, rather than treating every cell in the section as an expected positive (general IHC practice). |
| Signal is restricted to cell borders or another unexpected compartment, with little cytoplasmic staining. | A border-only pattern conflicts with HPA's general cytoplasmic tissue profile and the absence of a transmembrane segment (HPA: tissue IHC; UniProt Q92569 topology). Check morphology and detection artefacts before assigning this signal to PIK3R3 (general IHC practice). |
| Strong staining appears in an unexpected cell population, such as cardiomyocytes or esophageal squamous epithelium. | HPA reports these cell populations as Not detected (HPA: heart muscle and esophagus tissue IHC). Investigate antibody cross-reactivity and endogenous detection activity; HPA specifically cautions that its antibody targets proteins from more than one gene (HPA: tissue IHC reliability; general IHC practice). |
| Brown signal spreads across extracellular space, tissue edges, or many cell types without clear cellular boundaries. | This does not provide a convincing cell-resolved cytoplasmic pattern (HPA: tissue IHC profile; general IHC practice). Background from detection reagents, inadequate blocking, or excessive antibody concentration can obscure interpretation (general IHC practice). |
| No convincing staining appears in an expected-positive cell population. | HPA reports High staining in breast glandular cells, kidney glomerular cells, skin keratinocytes, and lymph-node germinal-center cells (HPA: tissue IHC). First verify that the relevant cells are present and assess assay controls; an absent signal alone does not establish absent PIK3R3 (general IHC practice). |
💡Expected PIK3R3 appearanceCall a result consistent with the reference pattern when identifiable HPA High cell populations show clear cytoplasmic chromogen above local background; widespread extracellular staining or isolated cell-border signal is suspect (HPA: tissue IHC; UniProt Q92569 topology; general IHC practice).
| Cell population and tissue selection | HPA reports High staining in selected cells of breast, bronchus, cerebral cortex, hippocampus, kidney, lymph node, pancreas, and skin, while several other populations are Low or Not detected (HPA: tissue IHC). Score the named cell type, since a whole-tissue label can hide that distinction (general IHC practice). |
| Antibody evidence | HPA005751 is Approved for IHC, but HPA cautions that the tissue antibody targets proteins from more than one gene and awaits external verification (HPA: antibody status and tissue IHC reliability). A matching pattern supports interpretation without proving that every stained cell contains PIK3R3. |
| Topology and isoforms | PIK3R3 has no transmembrane segment and has 3 reported isoforms (UniProt Q92569 topology and isoforms). The supplied record gives no antibody epitope or isoform coverage, so neither membrane-only staining nor isoform-specific staining can be established from these facts. |
| IF/ICC Q: Where should fluorescence localize? | A: HPA reports mainly nucleoplasm and cytosol in ICC-IF, with additional locations in sperm structures (HPA: subcellular ICC-IF). Its subcellular summary cautions that the antibodies target proteins from multiple genes; this observation is context for IF interpretation, not an IHC compartment requirement (HPA: subcellular ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| An expected-positive cell population has no chromogen. | The relevant cells may be absent from the section, or the IHC run may have failed (general IHC practice). HPA High is an observed pattern, not a guarantee for every specimen (HPA: tissue IHC). | Confirm cell identity and tissue preservation, then inspect the positive control and detection controls; optimize the assay using established antibody guidance (general IHC practice). |
| Signal is diffuse across the section. | Nonspecific binding or detection background can mask cell boundaries (general IHC practice). HPA instead describes general cytoplasmic expression (HPA: tissue IHC). | Compare a negative reagent control, review blocking and washes, and adjust antibody concentration within a validated range (general IHC practice). |
| Unexpected cells stain strongly. | Cross-reactivity is plausible because HPA cautions that its antibody targets proteins from more than one gene; endogenous detection activity is another possible IHC source (HPA: tissue IHC reliability; general IHC practice). | Check a reagent control and compare the cell type with HPA's named positive and Not detected populations; seek independent target validation before assigning PIK3R3 (HPA: tissue IHC; general IHC practice). |
| Staining appears only along membranes or tissue edges. | That pattern conflicts with general cytoplasmic tissue staining and PIK3R3's lack of a transmembrane segment (HPA: tissue IHC; UniProt Q92569 topology). Edge artefacts can occur in IHC (general IHC practice). | Examine intact interior cells, compare controls, and repeat staining if the compartment remains ambiguous (general IHC practice). |
| The candidate negative tissue shows some staining. | HPA's Not detected result applies to its specified cell population and assay; another population or assay background may account for the observed signal (HPA: tissue IHC; general IHC practice). | Identify the stained cells and compare them with the HPA cell-level entry; inspect reagent controls before using the section as a negative comparator (HPA: tissue IHC; general IHC practice). |
| IF/ICC shows nucleoplasmic signal that seems inconsistent with tissue IHC. | HPA reports nucleoplasm and cytosol in ICC-IF but a general cytoplasmic tissue IHC profile, both with antibody specificity cautions (HPA: subcellular ICC-IF and tissue IHC). | Interpret each observation within its assay and cell context; do not score nucleoplasmic fluorescence as proof of a nuclear IHC pattern (HPA: subcellular ICC-IF and tissue IHC; general IHC practice). |
Experiment evidence
IHC & IF images showing PIK3R3 staining
M06707 · IHCImmunohistochemical analysis of paraffin-embedded Human pituitary tumor, using the Antibody at 1:200 dilution.
M06707 · IHCImmunohistochemical analysis of paraffin-embedded Human lung adenocarcinoma, using the Antibody at 1:200 dilution.
M06707 · IFImmunofluorescent analysis of C6 cells, using PI 3 Kinase p55 gamma Antibody .
A06707-1 · IHCFormalin-fixed and paraffin-embedded human hepatocarcinoma tissue reacted with PIK3R3 antibody (C-term), which was peroxidase-conjugated to
A06707-1 · IFFluorescent image of U251 cells stained with PIK3R3 Antibody (C-term). PIK3R3 Antibody (C-term) was diluted at 1:100 dilution. An Alexa Fluo
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000117461-PIK3R3/tissue/cerebral+cortex#imid_2251536
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000117461-PIK3R3/tissue/colon#imid_2251478
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000117461-PIK3R3/tissue/liver#imid_2251479
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000117461-PIK3R3/tissue/kidney#imid_2251472
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA005751; green = PIK3R3, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA005751; green = PIK3R3, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for PIK3R3 IHC & IF
🧪Run cerebral cortex first: neuronal cells must show PIK3R3 staining (HPA: High in cerebral cortex neuronal cells). Use esophagus squamous epithelium as the negative tissue (HPA: Not detected in esophageal squamous epithelial cells); on the cortex slide, assess neighboring nonneuronal cells for background staining, without assuming they are PIK3R3-negative (HPA: neuronal-cell result only).
Positive control tissue: Breast (Glandular cells, HPA High)
Negative control tissue: Esophagus (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show PIK3R3 in A-431, U-251MG, U2OS, ASC52telo, hTERT-RPE1 (serum starved), Sperm, with annotated localisation: Nucleoplasm (approved), Cytosol (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), host-species- and clonality-matched isotype, and PIK3R3-knockout biological controls (standard IHC practice). Quench endogenous peroxidase for chromogenic detection, and check for lipofuscin autofluorescence if using cerebral cortex for IF (standard IHC/IF practice).
⚠️Feasibility: A target-specific fixation window and antigen-retrieval dependence are unreported in the supplied evidence; compare retrieval conditions during paraffin-section optimization (standard IHC practice). The selected M06707 paraffin-section caption reports a 1:200 dilution but leaves the fixative unreported (selected M06707 caption). The evidence does not establish that frozen sections or IF are easier; cortical lipofuscin can complicate IF interpretation (standard IF practice).
HPA tissue IHC evidence for PIK3R3
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Caution, targets protein from more than one gene. 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 |
|---|---|---|---|---|
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
| Hippocampus | Neuronal cells | High | Protein (IHC) | HPA → |
| Kidney | Cells in glomeruli | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced PIK3R3 IHC Tips
Troubleshoot PIK3R3 chromogenic IHC in paraffin sections using the catalog image, expression evidence, and matched controls.
What retrieval should I try first if PIK3R3 staining is weak?
Start with citrate pH 6.0 heat-induced epitope retrieval at 95–98 °C for 20 min (page retrieval setting). The M06707 paraffin-section image reports a 1:200 antibody dilution, but its caption does not report retrieval conditions or fixative (M06707 caption). Keep heating, cooling, section thickness, and detection consistent while comparing runs, and inspect both signal and tissue integrity (standard IHC practice). If staining remains weak, test alkaline EDTA retrieval as a separately validated fallback and check for increased background (standard IHC practice). Score cytoplasmic staining in the same cell population across conditions (HPA: general cytoplasmic expression; standard IHC practice).
Can I infer the best fixative for PIK3R3 from the published staining pattern?
No target-specific fixation sensitivity is established by the supplied evidence: the M06707 image shows paraffin-embedded human pituitary tumor at 1:200, but its caption does not state a fixative (M06707 caption). Record fixative identity, fixation time, processing history, and block age for each specimen before comparing staining (standard IHC practice). If your laboratory uses neutral buffered formalin, follow its validated schedule and compare differently processed samples with a consistently processed control (standard IHC practice). Judge morphology and staining together, since uneven processing can alter apparent intensity (standard IHC practice). Do not attribute an intensity difference specifically to PIK3R3 fixation sensitivity without a controlled comparison.
Should nuclear PIK3R3 staining be scored as positive in paraffin sections?
Start by assessing cytoplasmic staining, the pattern described for tissue IHC (HPA: general cytoplasmic expression). Nuclear signal needs separate validation: HPA reports nucleoplasm and cytosol in subcellular imaging, but cautions that those antibodies target proteins from multiple genes (HPA: subcellular summary). UniProt supplies no annotated subcellular location, although PIK3R3 has no transmembrane segment (UniProt Q92569: subcellular record and topology). Compare nuclear and cytoplasmic staining separately across matched sections, and check omission controls and a second specificity method before calling nuclear staining PIK3R3 (standard IHC practice). Record the scored compartment explicitly so apparent differences do not reflect inconsistent scoring.
How could isoforms or epitope placement complicate a PIK3R3 IHC result?
PIK3R3 has 3 reported isoforms, while the supplied M06707 caption does not identify the antibody epitope or establish isoform coverage (UniProt Q92569: isoforms; M06707 caption). Its SH2 domains span residues 65–160 and 358–452, and phosphotyrosine is reported at residue 341 (UniProt Q92569: domains and modified residues). Do not assume that staining distinguishes isoforms or phosphorylation states without epitope and specificity evidence. If samples disagree, compare an independently validated antibody recognizing a documented region and use an orthogonal expression measure on matched material (standard IHC practice). Keep retrieval and detection conditions constant during that comparison (standard IHC practice).
How should I check PIK3R3 localisation with multiplex IF?
Treat IF/ICC as a separate validation: the M06707 caption documents paraffin-section chromogenic IHC at 1:200, not an IF dilution or fixation method (M06707 caption). In cerebral cortex, pair PIK3R3 with a neuronal marker to identify the cell population reported as highly stained in tissue IHC (HPA: high in cerebral-cortex neuronal cells). Choose a far-red fluorophore when shorter-wavelength tissue autofluorescence interferes, and include single-channel and no-primary controls (standard IF practice). PIK3R3 lacks a transmembrane segment, so evaluate permeabilisation for access to intracellular epitopes rather than assuming a surface epitope (UniProt Q92569: topology; standard IF practice). Interpret nuclear overlap cautiously because HPA's subcellular antibodies target multiple genes (HPA: subcellular caution).
What should I adjust when PIK3R3 chromogenic staining looks diffuse?
First inspect a no-primary control and adjacent tissue for diffuse color, edge accentuation, pigment, or precipitate (standard IHC practice). For peroxidase-based detection, verify the peroxidase block, then shorten DAB development or adjust antibody concentration one variable at a time (standard IHC practice). The M06707 image reports 1:200 in paraffin-embedded human pituitary tumor, but does not establish that dilution for other tissues or processing conditions (M06707 caption). Include an appropriate protein block and matched wash conditions when troubleshooting nonspecific binding (standard IHC practice). Compare candidate signal with the reported general cytoplasmic tissue pattern while remembering HPA's cross-gene antibody caution (HPA: tissue profile and reliability description).
How can I quantify PIK3R3 staining across paraffin tissue samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and scored compartment before analysis; tissue IHC is described as generally cytoplasmic (HPA: tissue profile; standard IHC practice). For intensity, use an H-score based on percentages of cells at intensity grades 0–3, yielding 0–300, or report percentage-positive cells at a prespecified threshold (standard IHC practice). For sparse populations, report positive-cell density per mm² of viable tissue and normalize to the corresponding cell population or analyzed viable area (standard IHC practice). Apply one threshold, counterstain setting, and exclusion rule to all samples (standard IHC practice). Report nuclear staining separately if investigated, given the qualified subcellular evidence (HPA: subcellular caution).
How do I distinguish credible PIK3R3 positivity from staining artefact?
Look for reproducible staining in the expected cell type and compartment: HPA describes general cytoplasmic tissue expression and high staining in cerebral-cortex neuronal cells (HPA: tissue profile and cerebral cortex). HPA also reports no detection in heart-muscle cardiomyocytes, although its tissue-IHC reliability note cautions that antibodies may target proteins from multiple genes (HPA: heart muscle and reliability description). Treat isolated strong nuclear color, tissue-edge signal, necrotic debris, and color persisting without primary antibody as findings requiring investigation (standard IHC practice; HPA: general cytoplasmic expression). Check endogenous peroxidase and compare independently validated antibody or orthogonal expression evidence before assigning disputed staining to PIK3R3 (standard IHC practice).
Boster reagents
Best PIK3R3 / Phosphatidylinositol 3-kinase regulatory subunit gamma IHC Antibodies
The catalog shows human tissue IHC and cell IF images for PIK3R3 (catalog image captions). M06707 lists Human, Mouse and Rat reactivity; A06707-1 lists Human (catalog reactivity).
Real IHC data
Anti-PI 3 Kinase p55 gamma PIK3R3 Rabbit Monoclonal Antibody
Cat # M06707
Real IHC data
Anti-PIK3R3 Antibody (C-term)
Cat # A06707-1
M06707 has IHC images of paraffin-embedded human pituitary tumor and lung adenocarcinoma, plus an IF image of C6 cells (M06707 image captions). A06707-1 has a DAB IHC image of formalin-fixed, paraffin-embedded human hepatocarcinoma and an IF image of U251 cells (A06707-1 image captions).
Which to pick: For tissue IHC, A06707-1 documents paraffin-section, paraffin-embedded tissue (A06707-1 IHC caption); M06707 documents paraffin sections, but its caption does not report the fixative (M06707 IHC caption). For IF/ICC, M06707 lists both applications and shows C6 IF, while A06707-1 lists IF and shows U251 IF (catalog applications; IF captions). Choose M06707 for listed Human, Mouse and Rat reactivity and a monoclonal format; A06707-1 is Human-reactive and polyclonal (catalog reactivity; catalog descriptions). The selected M06707 tissue-IHC caption documents paraffin sections, but does not specify the fixative (selected-SKU IHC image M06707).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q92569 (P55G_HUMAN, Phosphatidylinositol 3-kinase regulatory subunit gamma).
- Human Protein Atlas. PIK3R3 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. PIK3R3 subcellular location (ICC-IF): Mainly localized to the nucleoplasm and cytosol. In addition localized to the connecting piece, mid piece, principal piece and end piece. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. PIK3R3 antibody validation summary (1 antibodies).
- CircRNA_0000392 promotes colorectal cancer progression through the miR-193a-5p/PIK3R3/AKT axis. Journal of experimental & clinical cancer research : CR 2020 — PMC7735421.
- Cutaneous transcriptome analysis in NIH hairless mice. PloS one 2017 — PMC5546695.
- Predictive microRNAs for lymph node metastasis in endoscopically resectable submucosal colorectal cancer. Oncotarget 2016 — PMC5078061.
- PIK3R3 is upregulated in liver cancer and activates Akt signaling to control cancer growth by regulation of CDKN1C and SMC1A. Cancer medicine 2023 — PMC10358214.
- PubMed PMID:9524259 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
- PubMed PMID:16710414 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for PIK3R3 (Phosphatidylinositol 3-kinase regulatory subunit gamma). Expected localisation General cytoplasmic staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: M06707 at 1:50 recommended; image 1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Breast (Glandular cells); negative Esophagus. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded Human pituitary tumor tissue; fixative not specified (datasheet M06707); 2. Fixation: Image fixative and duration unreported (datasheet M06707); 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 monoclonal (clone DHC-16) anti-PIK3R3, 1:50 recommended; image 1:200 (datasheet M06707); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep formalin fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: HPA antibody may recognize more than one gene (HPA tissue IHC); Isoform / epitope: 3 isoforms; verify epitope coverage (UniProt); Regulation: Expression regulation unreported (UniProt). Evidence: UniProt Q92569, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm and cytosol. In addition…, Datasheet M06707, PMC7735421, PMC5546695. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/PIK3R3/immunohistochemistry-ihc-guide.