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EIF2AK2 / Interferon-induced, double-stranded RNA-activated protein kinase · IHC design guide
Design Immunohistochemistry for EIF2AK2
Plan EIF2AK2 chromogenic IHC using its general cytoplasmic tissue pattern (HPA tissue IHC) and interferon-induced expression (UniProt). Score staining by cell type, intensity and compartment, allowing for elevated nuclear localisation in some cancers (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
EIF2AK2 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 tissue staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining across tissue cell types (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A01384-1) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific) | |
| Caveat | Nuclear localisation may rise in some cancers (UniProt) | |
| Regulation | Interferon-induced expression (UniProt) | |
| Isoform / epitope | Two isoforms; no extracellular domain; epitope coverage unknown (UniProt) |
Section 1
Recommended EIF2AK2 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A01384-1) is paired with one published EIF2AK2 paraffin-section IHC protocol (PMC10561166).
| Sample | Formaldehyde-fixed, paraffin-embedded rat lung tissue (datasheet A01384-1) |
| Fixation | Image formalin-fixed; duration unreported (datasheet A01384-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6 (datasheet A01384-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% serum (datasheet A01384-1) |
| Primary antibody | Rabbit anti-EIF2AK2, 2.5-5 μg/mL (datasheet A01384-1) |
| 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 | EIF2AK2-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 noteTry citrate pH 6 heat retrieval first (datasheet A01384-1); the published protocol also uses sodium citrate, but does not state its pH (PMC10561166).
Section 2
What Is the Expected EIF2AK2 Staining Pattern?
EIF2AK2 is found in the cytoplasm, nucleus and perinuclear region, and has no transmembrane segment (UniProt P19525). In paraffin sections, expect predominantly cytoplasmic staining in cell populations such as bone marrow hematopoietic cells and breast glandular cells (HPA: general cytoplasmic expression; High in both populations). HPA rates its tissue IHC profile Approved, with medium consistency between staining and RNA expression (HPA: reliability description).
| Cytoplasmic chromogen in bone marrow hematopoietic cells or breast glandular cells. | This fits the reported EIF2AK2 tissue pattern; HPA scores both cell populations High (HPA: tissue IHC). Judge staining within the named cells, rather than treating all cells in the section as equally positive. HPA's Approved profile has medium staining–RNA consistency, so it supports an expected pattern without proving every stained cell is specific (HPA: reliability description). |
| Strong nuclear staining dominates the section, with little cytoplasmic signal. | A broadly nuclear-only pattern departs from HPA's general cytoplasmic IHC profile (HPA: tissue IHC). Nuclear EIF2AK2 is biologically possible, and UniProt reports elevated nuclear localization in specified cancers and other disease samples (UniProt P19525: subcellular location). Review tissue context and controls before calling nuclear signal artefactual; an unexplained nuclear pattern warrants antibody or detection checks. |
| Adipocytes stain strongly, while expected positive cells stain weakly. | HPA reports EIF2AK2 as Not detected in adipocytes, whereas bone marrow hematopoietic cells and breast glandular cells are High (HPA: tissue IHC). Strong adipocyte staining is therefore discordant with this reference pattern. Consider nonspecific antibody binding or endogenous detection activity, and compare with a control section processed without primary antibody (general IHC practice). |
| Brown signal covers extracellular spaces, tissue edges and multiple cell types uniformly. | Uniform diffuse color makes cellular localization difficult to assess against HPA's general cytoplasmic pattern (HPA: tissue IHC). It can reflect background from detection chemistry, inadequate washing or nonspecific binding (general IHC practice). Check the no-primary control and assess whether any convincing cytoplasmic signal remains confined to cells after background is reduced. |
| No staining appears in bone marrow hematopoietic cells on a run. | That result conflicts with HPA's High staining call for those cells (HPA: tissue IHC). First check whether the control section, antigen retrieval and detection reagents worked, then verify that the sampled area contains the expected cells (general IHC practice). A blank slide alone cannot establish that EIF2AK2 is absent from the specimen. |
💡Expected EIF2AK2 appearanceCall the IHC result consistent with EIF2AK2 when identifiable high-expressing cells show predominantly cytoplasmic staining (HPA: High in bone marrow hematopoietic and breast glandular cells; general cytoplasmic expression); strong adipocyte or uniform extracellular color is discordant (HPA: adipocytes Not detected; general cytoplasmic expression).
| Cell population and tissue | HPA scores several glandular, respiratory epithelial and neuronal populations High, but adipocytes Not detected and several muscle or stromal populations Low (HPA: tissue IHC). Compare like cell types when judging intensity; a weak population is a poor sole positive control. |
| Compartment and disease context | HPA describes general cytoplasmic IHC expression and enhanced cytosolic ICC-IF localization (HPA: tissue IHC; subcellular). UniProt also lists nucleus and perinuclear region, with elevated nuclear localization in specified disease samples (UniProt P19525: subcellular location). Interpret nuclear signal in its specimen context. |
| Antibody validation | HPA lists IHC as Approved for HPA019795, HPA063893 and CAB003845; it lists ICC as Supported for the latter two (HPA: antibody validation). These application ratings help select a comparison reagent, but do not identify the epitope or establish that a different antibody will reproduce the pattern. |
| Isoforms and modified residues | UniProt lists two isoforms and multiple modified residues, including phosphorylation sites (UniProt P19525: isoforms; modified residues). Without the antibody's epitope and modification sensitivity, these facts do not predict a staining difference or justify a phospho-specific interpretation. |
| IF/ICC Q&A: What localization should fluorescence show? | Expect cytosolic signal in the HPA ICC-IF reference, which calls the cytosol the enhanced main location (HPA: subcellular). UniProt also permits nuclear and perinuclear localization (UniProt P19525: subcellular location); assess fluorescence with an appropriate nuclear counterstain and controls (general IF practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected positive cells are blank. | The run may have failed, or the sampled field may lack the expected cells (general IHC practice). | Confirm identifiable bone marrow hematopoietic cells or another HPA High population, then check the run's positive control, retrieval and detection steps (HPA: tissue IHC; general IHC practice). |
| Adipocytes show strong chromogen. | This conflicts with HPA's Not detected call and may reflect nonspecific binding or endogenous detection activity (HPA: tissue IHC; general IHC practice). | Inspect a no-primary control, check any detection-specific endogenous-activity block, and reassess staining in the same cell population (general IHC practice). |
| Most nuclei stain strongly across unrelated cell types. | The distribution differs from HPA's general cytoplasmic IHC profile, although UniProt allows nuclear localization in particular disease contexts (HPA: tissue IHC; UniProt P19525: subcellular location). | Review the specimen context, compare cytoplasmic signal and controls, and confirm a persistent nuclear pattern with another IHC-approved antibody if available (HPA: antibody validation; general IHC practice). |
| Diffuse color obscures cellular borders. | Background from nonspecific binding, detection chemistry or insufficient washing can obscure localization (general IHC practice). | Compare a no-primary control; adjust blocking, antibody concentration or washing according to the antibody and detection system (general IHC practice). Reassess whether the remaining signal matches HPA's cytoplasmic pattern (HPA: tissue IHC). |
| A low-staining cell type is used as the only positive control. | HPA calls alveolar cells, follicle cells, cardiomyocytes, smooth muscle cells, myocytes and fibroblasts Low (HPA: tissue IHC). Weak staining in one of these populations may be difficult to distinguish from a failed run. | Include a section containing a named HPA High population, such as bone marrow hematopoietic cells, as the positive reference (HPA: tissue IHC; general IHC practice). |
| ICC-IF looks cytosolic but the IHC section appears compartmentally different. | HPA's ICC-IF reference emphasizes cytosol, while UniProt also lists nucleus and perinuclear region; the records do not establish that every specimen must match exactly (HPA: subcellular; UniProt P19525: subcellular location). | Score each preparation against its own controls and cell identity, and investigate a reproducible discordance before assigning it biological meaning (general IHC/IF practice). |
Experiment evidence
IHC & IF images showing EIF2AK2 staining
A01384-1 · IHCImmunohistochemistry Validation of PKR in Rat Lung Immunohistochemical analysis of paraffin-embedded rat lung tissue using anti-PKR antibody
A01384-1 · IHCImmunohistochemistry Validation of PKR in Mouse Lung Immunohistochemical analysis of paraffin-embedded mouse lung tissue using anti-PKR anti
A01384-1 · IFImmunofluorescence Validation of PKR in Mouse Lung Immunofluorescent analysis of 4% paraformaldehyde-fixed mouse lung tissue labeling PKR wi
M01384 · IHCImmunohistochemical analysis of paraffin-embedded human kidney, using PKR Antibody.
M01384 · IFImmunofluorescent analysis of Hela cells, using PKR Antibody.
P01384 · IHCImmunohistochemical analysis of paraffin-embedded human colon, using Phospho-PKR (T446) Antibody.
PB9803 · IFIF analysis of PKR/EIF2AK2 using anti-PKR/EIF2AK2 antibody (PB9803). PKR/EIF2AK2 was detected in an immunocytochemical section of A549 cells
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000055332-EIF2AK2/tissue/cerebral+cortex#imid_21218923
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000055332-EIF2AK2/tissue/colon#imid_21218865
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000055332-EIF2AK2/tissue/liver#imid_21218858
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000055332-EIF2AK2/tissue/kidney#imid_21218851
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB003845; green = EIF2AK2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB003845; green = EIF2AK2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for EIF2AK2 IHC & IF
🧪Run bone marrow first: hematopoietic cells should stain (HPA: High in bone marrow hematopoietic cells). Use adipose tissue as the negative comparator; adipocytes within the bone marrow slide, if present, should show background-level staining (HPA: adipose adipocytes Not detected).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show EIF2AK2 in A-431, U-251MG, U2OS, SK-MEL-30, with annotated localisation: Cytosol (enhanced) (HPA subcellular).
Technical controls: Include no-primary (secondary-only) and primary-host-matched irrelevant IgG or isotype controls, plus EIF2AK2-knockout tissue as a biological negative (IHC practice). Quench endogenous peroxidase in bone marrow sections before chromogenic detection (IHC practice).
⚠️Feasibility: A target-specific fixation window is unreported, and the selected-SKU fixative is unreported (selected caption: fixative field not stated). The paraffin-section caption uses heat-mediated citrate retrieval at pH 6, but does not establish that retrieval is required (selected caption: citrate pH 6). The supplied evidence does not establish whether frozen sections or IF are easier; bone marrow hematopoietic cells may require careful assessment of endogenous peroxidase background (HPA: High in bone marrow hematopoietic cells; IHC practice).
HPA tissue IHC evidence for EIF2AK2
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 → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced EIF2AK2 IHC Tips
Troubleshoot chromogenic IHC for EIF2AK2 in paraffin sections by checking retrieval, compartment-specific staining and appropriate controls before scoring.
What retrieval should I try first when EIF2AK2 staining is weak?
Start with heat-mediated citrate buffer at pH 6 for antigen retrieval in paraffin sections (datasheet A01384-1). The selected antibody’s rat lung IHC example used this retrieval with 2.5 µg/ml primary antibody incubated overnight at 4°C (datasheet A01384-1). If staining remains weak, compare retrieval duration on adjacent sections while keeping antibody concentration and detection conditions fixed, and inspect tissue morphology for heat damage (standard IHC practice). Include a section processed without primary antibody and a consistently stained reference section in each comparison, so stronger signal is assessed alongside background (standard IHC practice).
Could fixation explain weak or uneven EIF2AK2 staining?
Target-specific fixation sensitivity for EIF2AK2 is unknown from the supplied evidence, so a weak section alone cannot establish that fixation masked its epitope. Record the fixative, fixation duration, tissue thickness and time before fixation for every specimen, then compare sections with similar processing histories (standard IHC practice). In paraffin IHC, variable fixation can change antigen accessibility and tissue morphology; assess retrieval and antibody conditions using adjacent sections before interpreting a staining difference biologically (standard IHC practice). Check a reference section and a no-primary control in the same run, and exclude damaged or poorly preserved regions from scoring (standard IHC practice).
Which EIF2AK2 staining pattern is plausible in paraffin IHC?
Expect predominantly cytoplasmic staining: tissue IHC reports general cytoplasmic expression, and cell imaging places EIF2AK2 in the cytosol (HPA tissue IHC; HPA subcellular). Perinuclear and nuclear localisation are also described, with increased nuclear localisation reported in several disease samples, so nuclear signal requires context rather than automatic rejection (UniProt P19525 subcellular). Score cytoplasmic and nuclear staining separately, using hematoxylin to identify nuclei and reviewing intact cells away from section edges (standard IHC practice). If staining appears confined to extracellular material or tissue tears, compare a no-primary control and inspect morphology before calling it EIF2AK2 (standard IHC practice).
Can the antibody distinguish EIF2AK2 isoforms or phosphorylation states in IHC?
EIF2AK2 has 2 listed isoforms and contains RNA-binding domains at residues 9–77 and 100–167, plus a kinase domain at 267–538 (UniProt P19525 domains and isoforms). Its record also lists phosphorylation sites, including residues 83 and 88–90 (UniProt P19525 modified residues). The supplied antibody evidence does not identify an epitope or establish isoform or phosphorylation-state selectivity, so chromogenic staining should be reported as EIF2AK2 immunoreactivity (datasheet A01384-1; UniProt P19525 isoforms). For isoform-specific or activation claims, obtain epitope information and validate specificity with suitable controls before interpreting stained cells (standard IHC practice).
How should I assess EIF2AK2 by multiplex IF alongside this IHC guide?
For a separate IF/ICC experiment, pair EIF2AK2 with a marker that identifies the cell population being evaluated, and compare channels in individual cells (standard IF practice). Choose a fluorophore and imaging channel that separate the EIF2AK2 signal from tissue autofluorescence, with single-stain and no-primary controls to assess bleed-through and background (standard IF practice). EIF2AK2 has no transmembrane segment and is reported in the cytosol, so use permeabilisation suitable for access to an intracellular epitope after fixation (UniProt P19525 topology; HPA subcellular; standard IF practice). Verify the antibody’s IF/ICC suitability separately; the supplied product example establishes paraffin IHC conditions (datasheet A01384-1).
How can I reduce diffuse brown staining without losing EIF2AK2 signal?
First compare stained sections with a no-primary control to identify signal arising from the detection system rather than the primary antibody (standard IHC practice). The selected example used 10% serum for 1 hour at room temperature and an HRP secondary at 1:250; these are starting conditions from that experiment (datasheet A01384-1). Check peroxidase blocking, washing and DAB development, then change one condition at a time while examining both background and cell-associated signal (standard chromogenic IHC practice). Because EIF2AK2 is broadly cytoplasmic in tissue IHC, diffuse staining alone is insufficient to judge specificity without morphology and controls (HPA tissue IHC; standard IHC practice).
How should I quantify EIF2AK2 staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, and apply the same region-selection rules to every section (standard IHC practice). For chromogenic IHC, report an H-score from intensity and percentage of positive cells, or report percentage positive cells and positive-cell density per mm² when cell counts suit the study question (standard IHC practice). Normalise positive counts to the number of evaluable cells in the selected population, or density to the measured viable tissue area; exclude folds and necrosis consistently (standard IHC practice). Keep cytoplasmic and nuclear scores separate because both compartments are reported for EIF2AK2 (UniProt P19525 subcellular; HPA tissue IHC).
When should EIF2AK2 staining be treated as artefact rather than a positive result?
Judge signal against intact cell morphology and expected cytoplasmic or possible nuclear localisation, rather than calling every brown deposit positive (HPA tissue IHC; UniProt P19525 subcellular; standard IHC practice). Staining restricted to edges, folds, necrotic regions or extracellular debris warrants review with a no-primary control and adjacent viable tissue (standard IHC practice). Signal in an unexpected cell population needs cell identification before assignment: tissue IHC reports high staining in bone-marrow hematopoietic cells but low staining in lung alveolar cells (HPA tissue IHC). If no-primary sections also stain, investigate endogenous peroxidase or detection background before interpreting the pattern as EIF2AK2 (standard chromogenic IHC practice).
Boster reagents
Best EIF2AK2 / Interferon-induced, double-stranded RNA-activated protein kinase IHC Antibodies
The catalog shows EIF2AK2 staining in paraffin-embedded rat lung, mouse lung, human kidney and human colon, plus IF in mouse lung and human cells (catalog image captions).
Real IHC data
Anti-PKR EIF2AK2 Antibody
Cat # A01384-1
Real IHC data
Anti-PKR EIF2AK2 Rabbit Monoclonal Antibody
Cat # M01384
Real IHC data
Anti-Phospho-PKR (T446) EIF2AK2 Rabbit Monoclonal Antibody
Cat # P01384
Real IF data
Anti-PKR/EIF2AK2 Antibody ®
Cat # PB9803
A01384-1 has IHC images from rat and mouse lung and IF images from mouse lung (catalog image captions); M01384 has a paraffin-embedded human kidney IHC image (catalog image caption). P01384 has a paraffin-embedded human colon phospho-PKR (T446) IHC image (catalog image caption); PB9803 has an A549 cell IF image and lists ICC and IF applications (catalog image caption; catalog applications).
Which to pick: For tissue IHC, choose M01384 for total PKR in human paraffin sections or P01384 for phospho-PKR (T446) in human paraffin sections; both are rabbit monoclonals, and neither IHC caption reports a fixative (catalog antibody details; catalog IHC image captions). For IF/ICC in human cells, PB9803 lists both applications and has an A549 IF image; its product record gives 5 μg/mL for IF, while the image caption reports 2 μg/mL, so optimize the working concentration (catalog applications; catalog dilution; catalog IF image caption). For IHC across species, A01384-1 lists human, mouse and rat reactivity, with IHC images in mouse and rat lung; those captions specify formaldehyde fixation and citrate-buffer heat retrieval, while human IHC validation is unreported (catalog reactivity; catalog IHC image captions; catalog validation statement).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P19525 (E2AK2_HUMAN, Interferon-induced, double-stranded RNA-activated protein kinase).
- Human Protein Atlas. EIF2AK2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. EIF2AK2 subcellular location (ICC-IF): Localized to the cytosol..
- Human Protein Atlas. EIF2AK2 antibody validation summary (3 antibodies).
- Eukaryotic translation initiation factor 2α kinase 2 in pancreatic cancer: An approach towards managing clinical prognosis and molecular immunological characterization. Oncology letters 2023 — PMC10561166.
- Mutation in Prkra results in cerebellar abnormality and reduced eIF2α phosphorylation in a model of DYT-PRKRA. Disease models & mechanisms 2024 — PMC11625895.
- A frameshift mutation in the murine Prkra gene causes dystonia and exhibits abnormal cerebellar development and reduced eIF2α phosphorylation. bioRxiv : the preprint server for biology 2024 — PMC11185611.
- Long non-coding RNA GRASLND enhances chondrogenesis via suppression of the interferon type II signaling pathway. eLife 2020 — PMC7202894.
- PubMed PMID:1695551 — UniProt-cited evidence.
- PubMed PMID:1373553 — UniProt-cited evidence.
- PubMed PMID:8921913 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for EIF2AK2 (Interferon-induced, double-stranded RNA-activated protein kinase). Expected localisation General cytoplasmic tissue staining (HPA tissue IHC). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A01384-1). Validated IHC antibody: A01384-1 at 2.5-5 μg/mL (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections. Protocol steps: 1. Sample: Formaldehyde-fixed, paraffin-embedded rat lung tissue (datasheet A01384-1); 2. Fixation: Image formalin-fixed; duration unreported (datasheet A01384-1); 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 (datasheet A01384-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% serum (datasheet A01384-1); 8. Primary antibody: Rabbit anti-EIF2AK2, 2.5-5 μg/mL (datasheet A01384-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across paraffin sections (standard IHC practice; not target-specific); Caveat: Nuclear localisation may rise in some cancers (UniProt); Isoform / epitope: Two isoforms; no extracellular domain; epitope coverage unknown (UniProt); Regulation: Interferon-induced expression (UniProt). Evidence: UniProt P19525, HPA tissue IHC · Approved, HPA ICC-IF · cytosol., Datasheet A01384-1, PMC10561166. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/EIF2AK2/immunohistochemistry-ihc-guide.