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AK2 / Adenylate kinase 2, mitochondrial · IHC design guide
Design Immunohistochemistry for AK2
Plan paraffin-section AK2 IHC around cytoplasmic staining (HPA tissue IHC) and its mitochondrial intermembrane-space location (UniProt). Use kidney tubules or glandular cells as positive references (HPA tissue IHC), and optimize the catalog antibody within 0.5–1 µg/mL (datasheet PB10034).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
AK2 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 stain (HPA tissue IHC); mitochondrial intermembrane space (UniProt) | |
| Staining pattern | Cytoplasmic; strong in glandular and kidney tubule cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet PB10034) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Caudate+3 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image PB10034) | |
| Caveat | Glial and smooth muscle cells: not detected (HPA tissue IHC) | |
| Regulation | No expression regulator specified (UniProt) | |
| Isoform / epitope | Six isoforms; N-terminal processing may alter epitopes (UniProt) |
Section 1
Recommended AK2 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet PB10034) is accompanied by a published human glioma tissue microarray protocol (PMC10290226).
| Sample | Paraffin-embedded rat liver tissue; fixative not specified (datasheet PB10034) |
| Fixation | Image fixative and duration unreported (datasheet PB10034); 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 PB10034); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB10034) |
| Primary antibody | Rabbit anti-AK2, 0.5-1μg/ml (datasheet PB10034) |
| Primary incubation | Overnight at 4 °C (datasheet PB10034) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB10034) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | AK2-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet PB10034); the glioma article reports antigen retrieval without specifying its conditions (PMC10290226).
Section 2
What Is the Expected AK2 Staining Pattern?
AK2 localizes to the mitochondrial intermembrane space and has no transmembrane segment (UniProt P54819: localization and topology). In paraffin-section IHC, expect cytoplasmic staining in many tissues, including strong staining in kidney tubule cells and colon glandular cells (HPA: tissue IHC). HPA rates the tissue pattern Approved, with medium consistency between antibody staining and RNA expression; interpret individual cell populations accordingly (HPA: tissue IHC reliability).
| Cytoplasmic staining is strong in kidney tubule cells or colon glandular cells (HPA: High in both cell populations). | This fits the observed IHC pattern (HPA: cytoplasmic expression in most tissues). A chromogenic section shows a cellular pattern; it cannot by itself resolve AK2 within the mitochondrial intermembrane space (UniProt P54819: localization; standard IHC practice). |
| A predominantly nuclear or cell-surface pattern replaces cytoplasmic staining (HPA: tissue IHC profile). | Treat this as discordant with the reported pattern, since AK2 is mitochondrial and lacks a transmembrane segment (UniProt P54819: localization and topology). Review morphology, controls and antibody specificity before interpreting it as AK2 (standard IHC practice). |
| Strong staining appears in caudate glial cells or smooth muscle cells (HPA: Not detected in those populations). | These cell-specific results conflict with HPA observations and may reflect cross-reactivity or detection background (HPA: tissue IHC; standard IHC practice). Check the stained cell identity and compare the negative reagent control before assigning AK2 positivity (standard IHC practice). |
| Broad, uniform color obscures cell boundaries or appears outside cells (standard IHC practice). | This is background rather than the interpretable cytoplasmic pattern reported for AK2 (HPA: tissue IHC; standard IHC practice). Compare a negative reagent control and inspect blocking, washes and chromogen development (standard IHC practice). |
| Kidney tubule cells show no signal (HPA: High in kidney tubule cells). | An absent result in this known-positive cell population warrants a technical check, though tissue processing and antibody performance cannot be diagnosed from HPA alone (HPA: tissue IHC; standard IHC practice). Verify that the positive control stains and that the relevant tubules are present (standard IHC practice). |
💡Expected AK2 appearanceCall AK2 positive when kidney tubule or colon glandular cells show clear, predominantly cytoplasmic staining at the reported High level; isolated nuclear, cell-surface or structureless color is discordant (HPA: tissue IHC; UniProt P54819: localization; standard IHC practice).
| Compartment and optical resolution | AK2 is in the mitochondrial intermembrane space (UniProt P54819: localization), while HPA describes tissue IHC as cytoplasmic (HPA: tissue IHC). Interpret chromogenic IHC at the compartment level; mitochondrial detail may be unresolved in a section (standard IHC practice). |
| Cell-specific reference levels | HPA reports High staining in kidney tubule, lung type II alveolar and pancreatic exocrine glandular cells, but Not detected in caudate glial and smooth muscle cells (HPA: tissue IHC). Judge each named population rather than treating an entire tissue as uniformly positive or negative (standard IHC practice). |
| Evidence strength | The tissue IHC reliability is Approved with medium consistency against RNA expression (HPA: tissue IHC reliability). HPA lists HPA018479 as IHC Approved; that status supports the reported pattern but does not make every unexpected stain target-specific (HPA: antibodies; standard IHC practice). |
| Processing and isoforms | UniProt lists six isoforms and chains spanning residues 1–239 or 2–239, including an N-terminally processed form (UniProt P54819: isoforms and processing). Without an antibody epitope in the supplied evidence, their effects on IHC recognition cannot be predicted (UniProt P54819: record scope). |
| Blood-cell interpretation | AK2 is reported in nucleated blood-cell populations and thrombocytes but not erythrocytes, which lack mitochondria (UniProt P54819: tissue specificity). HPA also reports Not detected in spleen red-pulp cells; distinguish that specific IHC observation from a claim about every splenic cell (HPA: tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive kidney tubules are unstained (HPA: High in kidney tubule cells). | The run or control may have failed; HPA provides no AK2-specific fixation sensitivity or retrieval requirement (HPA: tissue IHC; standard IHC practice). | Confirm tubule identity and a stained positive-control section; then review antibody application, retrieval and detection steps against the validated IHC workflow (standard IHC practice). |
| Only nuclei appear positive, without convincing cytoplasm (HPA: cytoplasmic tissue profile). | The compartment conflicts with mitochondrial AK2 localization (UniProt P54819: localization). | Recheck counterstain and cell boundaries, compare controls, and require reproducible cytoplasmic staining before scoring AK2 (standard IHC practice). |
| Caudate glial or smooth muscle cells stain strongly (HPA: Not detected in those cells). | Possible nonspecific binding or detection activity; the stain alone cannot identify which mechanism applies (HPA: tissue IHC; standard IHC practice). | Verify the cell type, inspect a negative reagent control and compare a known-positive cell population on the same run (standard IHC practice). |
| Diffuse color covers both tissue and empty spaces (standard IHC practice). | Background from the staining or detection workflow may obscure the reported cytoplasmic pattern (HPA: tissue IHC; standard IHC practice). | Review blocking, washing and chromogen development, then score only color with clear cellular localization (standard IHC practice). |
| Spleen red pulp appears negative while other nucleated blood cells stain (HPA: red-pulp cells Not detected; UniProt P54819: blood-cell expression). | The sources describe different cell populations and evidence types; a negative red-pulp observation does not negate expression in all nucleated blood cells (HPA: tissue IHC; UniProt P54819: tissue specificity). | Identify the specific cells being scored and report the red-pulp observation separately from other blood-cell populations (standard IHC practice). |
| Can IF/ICC confirm mitochondrial localization? | HPA reports approved mitochondrial localization by ICC-IF and lists CAB079913 as ICC Approved (HPA: subcellular; HPA: antibodies). | Use that observation to interpret IF/ICC images in its separate guide; keep this IHC call based on the tissue IHC pattern (HPA: tissue IHC and subcellular). |
Experiment evidence
IHC & IF images showing AK2 staining
PB10034 · IHCIHC analysis of AK2 using anti-AK2 antibody (PB10034). AK2 was detected in a paraffin-embedded section of rat liver tissue. Heat mediated an
PB10034 · IHCIHC analysis of AK2 using anti-AK2 antibody (PB10034). AK2 was detected in a paraffin-embedded section of mouse kidney tissue. Heat mediated
PB10034 · IFIF analysis of AK2 using anti-AK2 antibody (PB10034). AK2 was detected in immunocytochemical section of U20S cells. Enzyme antigen retrieval
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000004455-AK2/tissue/cerebral+cortex#imid_4999532
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000004455-AK2/tissue/colon#imid_4999458
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000004455-AK2/tissue/liver#imid_4999451
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000004455-AK2/tissue/pancreas#imid_4999410
HPA · ICC-IF U2OSHuman Protein Atlas immunofluorescence, U2OS, antibody CAB079913; green = AK2, blue = nucleus, red = microtubules.
HPA · ICC-IF U2OSHuman Protein Atlas immunofluorescence, U2OS, antibody CAB079913; green = AK2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for AK2 IHC & IF
🧪Run colon first and score glandular cells for AK2 staining (HPA: High in colon glandular cells). Use caudate glial cells as the negative tissue comparator (HPA: Not detected in caudate glial cells); intravascular erythrocytes on the colon slide should remain unstained (UniProt P54819: AK2 absent from erythrocytes).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show AK2 in U2OS, with annotated localisation: Mitochondria (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and rabbit immunoglobulin isotype controls matched to the catalog antibody’s class and clonality, plus an AK2-knockout biological negative (caption: rabbit primary antibody; standard IHC practice). Quench endogenous peroxidase and check for endogenous biotin background when using the caption’s SABC–DAB detection scheme (caption: SABC–DAB; standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the PB10034 paraffin-section caption does not state a fixative (caption: fixative not stated). The rat-liver IHC caption uses EDTA retrieval at pH 8.0, but it does not establish that AK2 staining depends on retrieval (caption: EDTA pH 8.0). The supplied evidence does not establish whether frozen sections or IF are easier; assess colonic mucin-related background when scoring glandular staining (HPA: High in colon glandular cells; standard IHC practice).
HPA tissue IHC evidence for AK2
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 → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
| Spleen | Cells in red pulp | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced AK2 IHC Tips
Troubleshoot AK2 staining by checking retrieval, mitochondrial localisation, cell identity, and controls before comparing chromogenic IHC scores across sections.
What retrieval should I start with for AK2 in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 for PB10034 (datasheet PB10034). The rat liver demonstration used a paraffin section, followed by 10% goat serum block and 1 μg/ml primary antibody overnight at 4°C; its fixative was not reported (datasheet PB10034). If signal is weak, compare a second retrieval condition on serial sections while holding antibody concentration, detection, and development time constant (standard IHC practice). Include a no-primary control to separate retrieval-related background from specific staining (standard IHC practice). Score the same cell population across conditions because AK2 expression varies among tissue compartments (HPA: tissue IHC profile).
Could fixation explain weak or uneven AK2 staining?
Target-specific sensitivity of AK2 to fixation is unknown from the supplied evidence; the PB10034 liver caption does not identify a fixative (datasheet PB10034). Record the fixative, fixation duration, processing history, and section age for each specimen before comparing staining (standard IHC practice). For uneven signal, compare similarly processed sections with the same EDTA pH 8.0 retrieval and 1 μg/ml primary concentration used in the catalog demonstration (datasheet PB10034; standard IHC practice). Inspect folds, detached tissue, and poorly preserved areas before changing antibody conditions, since these can distort chromogenic staining (standard IHC practice). Do not infer a fixation effect from cell-specific expression patterns (HPA: tissue IHC profile).
Should AK2 appear as diffuse cytoplasm or punctate mitochondrial staining?
AK2 resides in the mitochondrial intermembrane space (UniProt P54819: subcellular location). At routine chromogenic IHC resolution, that distribution may appear granular or broadly cytoplasmic, consistent with the reported cytoplasmic tissue profile (HPA: tissue IHC profile; standard IHC practice). Compare signal within identifiable cells and use a mitochondrial reference stain on an adjacent section when compartment assignment matters (standard IHC practice). Predominantly nuclear staining without convincing cytoplasmic signal deserves antibody and detection-control review because nuclei are outside the reported AK2 location (UniProt P54819: subcellular location; standard IHC practice). Avoid interpreting intensity across unlike cell populations as a localisation change (HPA: tissue IHC profile).
How could AK2 isoforms or epitope access affect my IHC result?
AK2 has 6 annotated isoforms and an N-terminally processed chain beginning at residue 2 (UniProt P54819: isoforms and processing). Its listed modifications include N-terminal acetylation and phosphorylation or succinylation at specified residues, but their effects on PB10034 binding are not established here (UniProt P54819: modified residues; datasheet PB10034). Check the antibody's disclosed immunogen or epitope before claiming that a staining pattern represents every isoform (standard IHC practice). Because AK2 has no annotated transmembrane segment, membrane topology alone cannot identify which sequence this antibody detects (UniProt P54819: topology; standard IHC practice). Validate an unexpected pattern with an independent AK2 reagent or orthogonal assay (standard IHC practice).
How should I assess AK2 by multiplex IF alongside cell identity?
For a separate IF experiment, pair AK2 with a validated marker of the expected cell type, such as a renal tubular cell marker when examining kidney; HPA reports high AK2 staining in kidney tubule cells (HPA: kidney cells in tubules; standard IF practice). Select spectrally separated fluorophores and test unstained tissue for autofluorescence before assigning a weak AK2 channel (standard IF practice). AK2 is in the mitochondrial intermembrane space, so intracellular staining requires permeabilisation that permits antibody access across mitochondrial membranes; the epitope's precise accessibility is unknown (UniProt P54819: subcellular location; standard IF practice). HPA reports approved mitochondrial localisation and U2OS ICC/IF images, which support a localisation comparison rather than an IF protocol for PB10034 (HPA: subcellular).
What controls help distinguish AK2 signal from chromogenic background?
Run no-primary and secondary-only controls alongside the AK2 section to reveal detection-system staining (standard IHC practice). Quench endogenous peroxidase before DAB development and assess tissue without primary antibody, because DAB signal can arise from endogenous enzyme activity (standard IHC practice). The PB10034 demonstration used a biotinylated secondary and streptavidin-biotin detection, so investigate endogenous biotin if background persists with that workflow (datasheet PB10034; standard IHC practice). Compare a 10% goat serum block and 1 μg/ml primary concentration with carefully controlled development time, as used in the rat liver demonstration (datasheet PB10034). Interpret diffuse haze against cell-level localisation and negative controls (UniProt P54819: subcellular location; standard IHC practice).
How should I quantify AK2 staining across IHC sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and tissue region before scoring, because HPA reports high staining in kidney tubule cells but no detection in caudate glial cells (HPA: kidney and caudate tissue IHC). For cellular staining, record percent positive cells and intensity grades, then calculate an H-score from 0–300 using the same thresholds across batches (standard IHC practice). For spatial comparisons, report positive-cell density per mm² of viable tissue or a specified compartment (standard IHC practice). Normalise to the number of eligible cells or viable compartment area, and keep retrieval, imaging, counterstain, and DAB development consistent (standard IHC practice). Exclude folds and necrotic regions by preset criteria (standard IHC practice).
When is an apparent AK2-positive cell likely to be an artefact?
A convincing AK2 result should fit a cytoplasmic mitochondrial pattern in an identifiable cell and exceed matched no-primary background (UniProt P54819: subcellular location; HPA: tissue IHC profile; standard IHC practice). Strong isolated nuclear staining is discordant with the reported mitochondrial location and merits a control check (UniProt P54819: subcellular location; standard IHC practice). Be cautious with edge-only deposits, necrotic tissue, and staining that persists after primary omission, which can reflect section or detection artefacts (standard IHC practice). Investigate endogenous peroxidase when DAB appears without primary antibody, and endogenous biotin when using the catalog demonstration's streptavidin-biotin system (datasheet PB10034; standard IHC practice). Interpret weak cells against tissue-specific reference patterns rather than declaring all cytoplasmic staining positive (HPA: tissue IHC profile; standard IHC practice).
Boster reagents
Best AK2 / Adenylate kinase 2, mitochondrial IHC Antibodies
PB10034 has IHC images from paraffin sections of rat liver, mouse kidney and human lung cancer tissue, plus a cell IF image (PB10034 image captions).
Real IHC data
Anti-AK2 Antibody ®
Cat # PB10034
PB10034 is the SKU that will render; its IHC captions document staining in paraffin sections from rat, mouse and human tissue (PB10034 IHC image captions). Its IF caption documents staining in U20S cells, and the catalog lists human, mouse and rat reactivity (PB10034 IF image caption; catalog reactivity).
Which to pick: For tissue IHC, pick PB10034: its own captions document EDTA retrieval at pH 8.0 and DAB detection in paraffin sections; the fixative is unreported (PB10034 IHC image captions). For IF/ICC, PB10034 has a cell IF image; M04253-1 lists IF/ICC but has no supplied IF image (PB10034 IF image caption; M04253-1 catalog applications and image alts). For work across human, mouse and rat, both list those species; PB10034 has IHC images across all three, while M04253-1 is a rabbit monoclonal with no supplied IHC image (catalog reactivity; PB10034 IHC image captions; M04253-1 catalog host, clone and image alts).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P54819 (KAD2_HUMAN, Adenylate kinase 2, mitochondrial).
- Human Protein Atlas. AK2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. AK2 subcellular location (ICC-IF): Localized to the mitochondria..
- Human Protein Atlas. AK2 antibody validation summary (2 antibodies).
- Adenylate kinase 2 is a biomarker related to the prognosis of glioma and the immune microenvironment. Journal of clinical laboratory analysis 2023 — PMC10290226.
- AK2 is an AMP-sensing negative regulator of BRAF in tumorigenesis. Cell death & disease 2022 — PMC9117275.
- Aurora kinase A (AURKA) expression in colorectal cancer liver metastasis is associated with poor prognosis. British journal of cancer 2013 — PMC3817339.
- The expression of AURKA is androgen regulated in castration-resistant prostate cancer. Scientific reports 2017 — PMC5740165.
- PubMed PMID:8843353 — UniProt-cited evidence.
- PubMed PMID:9504408 — UniProt-cited evidence.
- PubMed PMID:9434148 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for AK2 (Adenylate kinase 2, mitochondrial). Expected localisation Cytoplasmic stain (HPA tissue IHC); mitochondrial intermembrane space (UniProt). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet PB10034). Validated IHC antibody: PB10034 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Caudate. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded rat liver tissue; fixative not specified (datasheet PB10034); 2. Fixation: Image fixative and duration unreported (datasheet PB10034); 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: EDTA pH 8.0 (datasheet PB10034); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB10034); 8. Primary antibody: Rabbit anti-AK2, 0.5-1μg/ml (datasheet PB10034); 9. Primary incubation: Overnight at 4 °C (datasheet PB10034). Watch for: Fixation: Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image PB10034); Caveat: Glial and smooth muscle cells: not detected (HPA tissue IHC); Isoform / epitope: Six isoforms; N-terminal processing may alter epitopes (UniProt); Regulation: No expression regulator specified (UniProt). Evidence: UniProt P54819, HPA tissue IHC · Approved, HPA ICC-IF · mitochondria., Datasheet PB10034, PMC10290226. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/AK2/immunohistochemistry-ihc-guide.