This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
AKT2 / RAC-beta serine/threonine-protein kinase · IHC design guide
Design Immunohistochemistry for AKT2
Plan AKT2 chromogenic IHC on paraffin sections using the catalog antibody’s IHC protocol (datasheet PB9043). Interpret cytoplasmic staining (HPA tissue IHC) in light of uncertain staining reliability (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
AKT2 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 in tissue (HPA tissue IHC); nuclear or membrane localisation is possible (UniProt) | |
| Staining pattern | General cytoplasmic staining across tissues (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet PB9043) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Heart muscle |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining has medium RNA concordance (HPA tissue IHC) | |
| Regulation | Myoblast differentiation shifts nuclear localisation (UniProt) | |
| Isoform / epitope | Two isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended AKT2 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: PB9043) is followed by published AKT2 staining protocols from three articles (PMC8353139; PMC9224635; PMC3271997).
| Sample | Paraffin-embedded human lung cancer tissues; fixative not specified (datasheet PB9043) |
| Fixation | Image fixative and duration unreported (datasheet PB9043); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet PB9043) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB9043) |
| Primary antibody | Rabbit anti-AKT2, 0.5-1μg/ml (datasheet PB9043) |
| Primary incubation | Overnight at 4 °C (datasheet PB9043) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9043) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | AKT2-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General cytoplasmic expression. No signal in the no-primary control. |
💡Decision noteStart with heat retrieval in citrate buffer at pH 6 (datasheet: PB9043); a published AKT2 protocol also used citrate at pH 6 (PMC3271997).
Section 2
What Is the Expected AKT2 Staining Pattern?
AKT2 is expected mainly in the cytoplasm on tissue IHC, with context-dependent nuclear and membrane localisation (UniProt P31751; HPA: general cytoplasmic expression). High staining is reported in selected glandular, ciliated, glial, Purkinje and endothelial cells (HPA: tissue IHC). Treat these as reference patterns: HPA rates the tissue IHC profile Uncertain because antibody staining and RNA expression have medium consistency (HPA: tissue IHC).
| Cytoplasmic staining in adrenal or breast glandular cells, with identifiable cell boundaries and a clean surrounding field. | This fits the reported High glandular-cell staining and general cytoplasmic profile (HPA: tissue IHC). Compare signal within the named cells; a uniformly dark section gives less confidence in the pattern (general IHC practice). |
| Nuclear staining accompanies cytoplasmic staining, or a small subset of cells shows membrane-associated signal. | Neither compartment alone proves an artefact: UniProt lists nucleus and cell membrane and describes phosphoinositide-dependent membrane recruitment (UniProt P31751). Judge whether the distribution is cell-associated and reproducible; widespread extracellular deposit is inconsistent with those localisations (general IHC practice). |
| Strong staining appears in a cell population other than the one listed for that tissue. | Check the named cell population before scoring the section positive (HPA: tissue IHC). Signal in unrelated structures can reflect cross-reactivity or endogenous detection activity (general IHC practice); HPA's Uncertain IHC reliability limits claims of AKT2 specificity (HPA: tissue IHC). |
| Brown colour spreads across stroma, lumina or the entire section without a readable cellular pattern. | That is background rather than evidence for the reported cytoplasmic distribution (HPA: tissue IHC; general IHC practice). Review blocking, washes and detection controls before interpreting intensity; a tissue-wide haze can obscure genuine positive cells (general IHC practice). |
| No staining appears in adrenal glandular cells or another supplied High reference population. | First suspect an assay or section problem and inspect controls (HPA: High in adrenal glandular cells; general IHC practice). Absence in one section does not overturn the tissue profile, especially given its Uncertain IHC reliability (HPA: tissue IHC). |
💡Expected AKT2 appearanceCall positive a readable, predominantly cytoplasmic signal in an HPA-listed cell population—potentially High in breast or adrenal glandular cells—while treating diffuse extracellular colour or staining unrelated to identifiable cells as suspect (HPA: tissue IHC; general IHC practice).
| Which localisation should guide IHC scoring? | Use the HPA general cytoplasmic tissue profile as the starting point (HPA: tissue IHC). UniProt also lists nucleus, cell membrane and early endosome, so compartment alone cannot validate antibody specificity (UniProt P31751; general IHC practice). |
| How strong is the tissue IHC evidence? | HPA labels the overall tissue IHC profile Uncertain; the listed IHC antibody records CAB004204 and CAB080295 are also Uncertain (HPA: tissue IHC; HPA: antibodies). Interpret a match as supportive, then assess controls and cellular morphology (general IHC practice). |
| Do membrane or nuclear patterns require context? | Membrane recruitment depends on binding of the PH domain to phosphoinositides, while localisation in primary myoblasts differs with differentiation (UniProt P31751). Those observations explain possible variation; they do not predict a fixed membrane-to-cytoplasm ratio in paraffin tissue (UniProt P31751). |
| Can the ICC-IF pattern be used as an IHC-P scoring rule? | No. ICC-IF shows mainly vesicles and cytosol, with additional nucleoplasm; its localisation evidence is from a separate assay (HPA: subcellular ICC-IF). Use tissue IHC observations and tissue morphology for chromogenic section scoring (HPA: tissue IHC; general IHC practice). |
| Do isoforms or protein processing establish a distinct stain? | UniProt lists 2 isoforms and a chain spanning residues 1–481, with no signal peptide or propeptide (UniProt P31751). Without an antibody epitope in the supplied evidence, assign neither isoform-specific staining nor a cleavage-related pattern (UniProt P31751). |
| Situation | Likely cause | Next action |
|---|---|---|
| Reference tissue is blank, including HPA-listed glandular cells. | The section, primary-antibody step or detection sequence may have failed (general IHC practice); HPA reports High staining in adrenal and breast glandular cells (HPA: tissue IHC). | Check a processed positive-control section, reagent order and detection controls; then optimise retrieval and antibody concentration as general IHC variables (general IHC practice). Do not infer AKT2-specific fixation sensitivity from this result. |
| Signal is weak but confined to the expected cells. | A low-intensity run can be difficult to distinguish from a real tissue difference (general IHC practice). HPA reports different staining levels across named cell populations (HPA: tissue IHC). | Compare the same cell type across a positive control and test section, then adjust detection or primary concentration with a matched negative control (general IHC practice). Record intensity separately from the fraction of positive cells. |
| Background covers stroma or lumina. | Nonspecific detection, inadequate blocking or insufficient washes can obscure cellular staining (general IHC practice). HPA describes a general cytoplasmic pattern (HPA: tissue IHC). | Inspect the no-primary control, blocking step and wash conditions; reduce detection or primary concentration if the control supports that change (general IHC practice). Score only signal attributable to intact cells. |
| Only broad nuclear staining is visible. | Nuclear AKT2 is possible, but a nuclear-only tissue pattern lacks the reported general cytoplasmic IHC pattern (UniProt P31751; HPA: tissue IHC). | Check morphology and a negative detection control, then compare another HPA-listed positive cell population (general IHC practice; HPA: tissue IHC). Treat the result as unresolved unless controls support specific staining. |
| Unexpected cells or vessel contents stain strongly. | Cross-reactivity or endogenous detection activity may mimic a positive result (general IHC practice). HPA lists specific positive cell populations, including cerebral-cortex endothelial cells (HPA: tissue IHC). | Identify the stained cell type on the counterstain and inspect an appropriate detection control (general IHC practice). Do not assign AKT2 positivity solely from colour intensity, especially with Uncertain IHC reliability (HPA: tissue IHC). |
| Cardiomyocytes appear positive in heart muscle. | HPA reports cardiomyocytes as Not detected, but its tissue IHC profile has Uncertain reliability (HPA: tissue IHC). A discrepant result needs verification. | Confirm cardiomyocyte identity, inspect background and detection controls, and compare with a listed positive tissue processed in the same run (general IHC practice; HPA: tissue IHC). Report the discrepancy rather than forcing a negative score. |
Experiment evidence
IHC & IF images showing AKT2 staining
PB9043 · IHCIHC analysis of AKT2 using anti-AKT2 antibody (PB9043). AKT2 was detected in paraffin-embedded section of human lung cancer tissues. Heat me
PB9043 · IHCIHC analysis of AKT2 using anti-AKT2 antibody (PB9043). AKT2 was detected in frozen section of rat brain tissues. The tissue section was blo
M00725-1 · IFIF analysis of AKT2 using anti-AKT2 antibody (M00725-1). AKT2 was detected in immunocytochemical section of MCF7 cells. Enzyme antigen retri
M00725-3 · IFImmunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (human cervical epithelial adenocarcinoma cell
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000105221-AKT2/tissue/cerebral+cortex#imid_1368892
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000105221-AKT2/tissue/colon#imid_1368818
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000105221-AKT2/tissue/liver#imid_1368819
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000105221-AKT2/tissue/kidney#imid_1368804
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA064521; green = AKT2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA064521; green = AKT2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for AKT2 IHC & IF
🧪Run breast first and expect staining in glandular cells (HPA: High in breast glandular cells). Use heart muscle cardiomyocytes as the negative tissue (HPA: Not detected in cardiomyocytes); treat cells outside the breast glands as internal negatives only if their AKT2-negative status is independently established (UniProt P31751: widely expressed).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Heart muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show AKT2 in A-549, RT-4, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Vesicles (enhanced), Cytosol (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, a rabbit IgG isotype control matched to the primary antibody’s clonality, and AKT2 knockout material or a validated peptide-block control (PB9043 tissue-IHC caption: rabbit primary antibody). For SABC/DAB detection, block endogenous peroxidase and check endogenous biotin background on the breast section (PB9043 tissue-IHC caption: SABC/DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected PB9043 paraffin-section caption does not state the fixative (PB9043 tissue-IHC caption). The caption uses citrate buffer at pH 6 for 20 minutes, but does not establish that retrieval is required; the supplied evidence does not establish that frozen sections or IF are easier (PB9043 tissue-IHC caption). In breast, score glandular cells against adjacent background while checking for endogenous peroxidase or biotin signal from the SABC/DAB method (HPA: High in breast glandular cells; PB9043 tissue-IHC caption: SABC/DAB).
HPA tissue IHC evidence for AKT2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — 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 → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (cell body) | High | Protein (IHC) | HPA → |
| Caudate | Glial cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced AKT2 IHC Tips
Troubleshoot AKT2 staining in paraffin sections by checking retrieval, compartment pattern, antibody specificity and cell level scoring before interpreting signal.
What retrieval conditions should I try when AKT2 staining is weak?
Start with heat mediated retrieval in citrate buffer at pH 6 for 20 minutes (datasheet PB9043). The catalog antibody detected AKT2 in a paraffin section of human lung cancer tissue under those conditions (datasheet PB9043). Keep the antibody concentration at the reported 1 μg/ml during the initial comparison, so a retrieval change remains interpretable (datasheet PB9043; standard IHC practice). If staining stays weak, check heating consistency, section adhesion and whether the detection reagents work on a control section (standard IHC practice). Record retrieval temperature and cooling time alongside the staining result, because these affect epitope exposure in paraffin sections (standard IHC practice).
Could fixation explain weak or uneven AKT2 staining?
Target specific AKT2 sensitivity to fixation is unknown: the PB9043 tissue caption identifies a paraffin section but does not state its fixative (datasheet PB9043). If sections differ in staining, compare documented fixation conditions and processing histories before attributing the difference to AKT2 expression (standard IHC practice). Keep citrate retrieval at pH 6 for 20 minutes and the reported antibody concentration at 1 μg/ml while assessing that variation (datasheet PB9043; standard IHC practice). Examine morphology and staining across each section, since uneven processing can complicate chromogenic interpretation (standard IHC practice). Neither tissue staining patterns nor AKT2 sequence features establish a fixation effect for this antibody (HPA tissue IHC; UniProt P31751; datasheet PB9043).
Should nuclear or membrane AKT2 staining count as positive?
Cytoplasmic staining is a useful starting pattern in tissue: the HPA describes general cytoplasmic expression, while rating its tissue IHC reliability uncertain (HPA tissue IHC). AKT2 can also localize to the nucleus, cell membrane and early endosomes; its PH domain mediates lipid dependent membrane recruitment (UniProt P31751 localisation). Judge these patterns in intact cells against matched controls and morphology, rather than counting every brown deposit as intracellular signal (standard IHC practice). A nuclear pattern can be biologically plausible, but the reported shift toward nuclear localization concerns differentiated primary myoblasts specifically (UniProt P31751 localisation). Do not extrapolate that cell state pattern to every paraffin tissue (UniProt P31751 localisation; standard IHC practice).
How do I assess whether staining distinguishes AKT2 isoforms or related kinases?
AKT2 has 2 reported isoforms and is closely related to AKT1 and AKT3 (UniProt P31751 isoforms; UniProt P31751 function). The supplied PB9043 caption does not identify its epitope or establish which isoforms it recognizes (datasheet PB9043). Before assigning an isoform, obtain epitope and cross reactivity information for the catalog antibody and compare that region with the relevant sequences (standard IHC practice). The PH domain spans residues 5–108 and the kinase domain spans 152–409, so an epitope's location could matter for interpretation (UniProt P31751 domains). Treat chromogenic staining as AKT2 associated until antibody specificity and the isoform question are supported independently (standard IHC practice).
How should I plan IF alongside the AKT2 tissue IHC result?
For a separate IF experiment, multiplex AKT2 with a marker of the expected cell type so cell identity can be checked independently of its signal (standard IF practice). The HPA reports AKT2 mainly in vesicles and cytosol, with additional nucleoplasmic localization in ICC/IF images (HPA subcellular). Choose a fluorophore and imaging channel that separate specific signal from the specimen's autofluorescence, using an unstained specimen to assess that background (standard IF practice). AKT2 has no transmembrane segment; plan permeabilisation to access intracellular cytosolic, vesicular or nuclear epitopes, then verify the pattern for the antibody used (UniProt P31751 topology; HPA subcellular; standard IF practice). The PB9043 paraffin IHC caption does not establish IF performance (datasheet PB9043).
What should I change when DAB appears throughout the section?
First distinguish diffuse cellular staining from deposits over empty spaces, folds, damaged edges or necrotic areas (standard IHC practice). The reported PB9043 tissue method used 10% goat serum blocking, a biotinylated goat anti-rabbit secondary, a streptavidin biotin complex and DAB (datasheet PB9043). Include a no primary control and assess endogenous peroxidase and biotin contributions before changing the AKT2 antibody concentration (standard IHC practice). If background persists, review wash stringency, blocking and DAB development time while retaining a positively stained control section (standard IHC practice). Because AKT2 is widely expressed, genuine cytoplasmic staining may occur in several cell populations (UniProt P31751 tissue specificity; HPA tissue IHC).
How should I score AKT2 staining across specimens? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, since AKT2 may appear in cytoplasm, nucleus or membrane (UniProt P31751 localisation; standard IHC practice). For chromogenic sections, report an H-score or the percentage of positive cells within that defined population, and record intensity separately when useful (standard IHC practice). For spatial comparisons, count positive cells per mm² of viable, evaluable tissue and normalize to the sampled area or total eligible cells (standard IHC practice). Apply one threshold, exposure and exclusion rule across specimens, and document how edges, folds and necrosis were handled (standard IHC practice). Treat HPA tissue patterns as context rather than a scoring standard because their AKT2 IHC reliability is uncertain (HPA tissue IHC).
When is an unexpected AKT2 positive region convincing?
Check whether DAB lies within intact cells and a plausible AKT2 compartment: cytoplasm, nucleus or a localized membrane pattern can fit the record (UniProt P31751 localisation; standard IHC practice). Identify the stained cell type before comparing it with reference patterns; HPA reports high signal in several cell types but rates tissue IHC reliability uncertain (HPA tissue IHC). Be cautious with signal confined to section edges, folds, necrosis or extracellular debris, which can reflect processing artefact (standard IHC practice). A no primary control helps reveal detection background, including endogenous enzyme activity (standard IHC practice). Interpret a convincing pattern alongside morphology and independent evidence, without equating total AKT2 staining with kinase activity (UniProt P31751 function; standard IHC practice).
Boster reagents
Best AKT2 / RAC-beta serine/threonine-protein kinase IHC Antibodies
Anti-AKT2 antibodies have image data for human paraffin-section IHC, rat frozen-section IHC, and human-cell IF in MCF7 and HeLa (catalog image captions).
Real IHC data
Anti-AKT2 Antibody ®
Cat # PB9043
Real IF data
Anti-AKT2 Antibody ® (monoclonal, 10C6)
Cat # M00725-1
Real IF data
Anti-AKT2 Antibody
Cat # M00725-3
PB9043 has IHC images from paraffin-embedded human lung and intestinal cancer tissue and frozen rat brain (PB9043 image captions). M00725-1 has IF data in MCF7 cells, while M00725-3 has IF data in HeLa cells (M00725-1 and M00725-3 image captions).
Which to pick: For paraffin-section tissue IHC, choose PB9043: its rabbit antibody was used at 1 μg/ml with citrate retrieval at pH 6; the fixative is unreported (PB9043 image captions). For human-cell IF/ICC, choose mouse monoclonal M00725-1 at 2 μg/ml in MCF7; M00725-3 is an IF option shown in HeLa at 1:25 (catalog applications; M00725-1 and M00725-3 image captions). For cross-species work, PB9043 lists human, mouse and rat reactivity with mouse/rat frozen-section IHC, while M00725-3 lists human and rat reactivity for IF (PB9043 datasheet; M00725-3 catalog).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P31751 (AKT2_HUMAN, RAC-beta serine/threonine-protein kinase).
- Human Protein Atlas. AKT2 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. AKT2 subcellular location (ICC-IF): Mainly localized to vesicles and cytosol. In addition localized to the nucleoplasm..
- Human Protein Atlas. AKT2 antibody validation summary (4 antibodies).
- Potential of AKT2 expression as a predictor of lymph-node metastasis in invasive breast carcinoma of no special type. Journal of pathology and translational medicine 2021 — PMC8353139.
- Vav1 Selectively Down-Regulates Akt2 through miR-29b in Certain Breast Tumors with Triple Negative Phenotype. Journal of personalized medicine 2022 — PMC9224635.
- Akt2 regulates metastatic potential in neuroblastoma. PloS one 2013 — PMC3582607.
- Human Papillomaviruses, p16INK4a and Akt expression in basal cell carcinoma. Journal of experimental & clinical cancer research : CR 2011 — PMC3271997.
- PubMed PMID:1801921 — UniProt-cited evidence.
- PubMed PMID:1409633 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for AKT2 (RAC-beta serine/threonine-protein kinase). Expected localisation Cytoplasmic in tissue (HPA tissue IHC); nuclear or membrane localisation is possible (UniProt). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet PB9043). Validated IHC antibody: PB9043 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Heart muscle. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissues; fixative not specified (datasheet PB9043); 2. Fixation: Image fixative and duration unreported (datasheet PB9043); 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 retrieval: Citrate pH 6 , 20 min (datasheet PB9043); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB9043); 8. Primary antibody: Rabbit anti-AKT2, 0.5-1μg/ml (datasheet PB9043); 9. Primary incubation: Overnight at 4 °C (datasheet PB9043). Watch for: Fixation: Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Caveat: Antibody staining has medium RNA concordance (HPA tissue IHC); Isoform / epitope: Two isoforms; epitope coverage is unspecified (UniProt); Regulation: Myoblast differentiation shifts nuclear localisation (UniProt). Evidence: UniProt P31751, HPA tissue IHC · Uncertain, HPA ICC-IF · Mainly localized to vesicles and cytosol. In addition localized to…, Datasheet PB9043, PMC8353139, PMC9224635. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/AKT2/immunohistochemistry-ihc-guide.