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CRK / Adapter molecule crk · IHC design guide
Design Immunohistochemistry for CRK
Plan chromogenic CRK IHC in paraffin sections using the reported cytoplasmic and membranous tissue pattern (HPA tissue IHC). Select positive control cells from the reported high-staining tissues (HPA tissue IHC), and check isoform coverage before scoring (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CRK 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 and membranous staining (HPA tissue IHC) | |
| Staining pattern | Membranous and cytoplasmic staining in several tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet PB9101) | |
| Positive control | Duodenum+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Presumed off-target staining can confound scoring (HPA tissue IHC) | |
| Regulation | Adhesion recruits CRK to the plasma membrane (UniProt) | |
| Isoform / epitope | Two isoforms; verify antibody epitope coverage (UniProt) |
Section 1
Recommended CRK IHC & IF Protocols
The catalog antibody protocol (datasheet: PB9101) is accompanied by four published CRK IHC protocols (PMC11896132; PMC4477307; PMC4471787; PMC3695334).
| Sample | Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet PB9101) |
| Fixation | Image fixative and duration unreported (datasheet PB9101); 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 PB9101); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet PB9101) |
| Primary antibody | Rabbit anti-CRK, 2-5μg/ml (datasheet PB9101) |
| Primary incubation | Overnight at 4 °C (datasheet PB9101) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet PB9101) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CRK-positive staining in glandular cells of duodenum (HPA tissue IHC: High). HPA tissue profile: Membranous and cytoplasmic expression in several tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet: PB9101); the published citrate step has no stated heating conditions (PMC11896132).
Section 2
What Is the Expected CRK Staining Pattern?
CRK should show membranous and cytoplasmic staining in several tissues (HPA tissue IHC). Expect strong staining in duodenal glandular cells, kidney tubular cells and hepatocytes (HPA: High). CRK has no transmembrane segment and can move to the plasma membrane during cell adhesion (UniProt P46108 topology and localisation). HPA rates its tissue IHC profile Approved, with medium consistency between staining and RNA data (HPA tissue IHC).
| Membranous and cytoplasmic staining in duodenal glandular cells or kidney tubular cells. | This matches the reported compartments and High staining in those cells (HPA tissue IHC). Judge the named cell population, since adjacent cells need not have the same intensity. |
| Predominantly nuclear staining in a tissue section, with little membranous or cytoplasmic signal. | This departs from the reported tissue IHC profile (HPA tissue IHC). Check specificity before calling it CRK; nucleoplasmic localisation is uncertain in ICC-IF and does not establish a nuclear IHC pattern (HPA subcellular). |
| Strong staining of adipocytes in adipose tissue or hematopoietic cells in bone marrow. | Those specific cell populations are reported as Not detected (HPA tissue IHC). Consider cross-reactivity or endogenous detection activity, and inspect a no-primary control (general IHC practice). |
| Diffuse chromogen over cells and tissue spaces, obscuring cell boundaries. | That distribution cannot be scored reliably against the membranous and cytoplasmic profile (HPA tissue IHC). It may reflect nonspecific detection or background; compare with a no-primary control (general IHC practice). |
| No convincing signal in duodenal glandular cells or hepatocytes. | Both are reported High (HPA tissue IHC). First assess whether the run detected antigen in a known-positive section; a blank section alone cannot establish absent CRK expression (general IHC practice). |
💡Expected CRK appearanceA convincing positive shows strong membranous and cytoplasmic signal in a named High cell population, such as kidney tubular cells (HPA tissue IHC); widespread acellular colour or staining confined to an unexpected cell population warrants a control check (general IHC practice).
| Compartment and membrane association | CRK is cytoplasmic and can move to the plasma membrane during adhesion (UniProt P46108); tissue IHC reports both membranous and cytoplasmic staining (HPA tissue IHC). Membrane signal need not imply a transmembrane protein (UniProt P46108 topology). |
| Cell population and staining level | Hepatocytes and salivary glandular cells are High, while colon glandular cells are Medium (HPA tissue IHC). Compare the identified cells rather than assigning one intensity to an entire organ. |
| Tissue IHC evidence limit | The profile is Approved, with medium staining–RNA consistency; presumed off-target binding was observed and disregarded (HPA tissue IHC). Treat an unexpected pattern as a finding to validate, not as established CRK localisation. |
| Antibody-specific validation | CAB010485 is Approved for IHC, while HPA068087 is Approved for ICC and has no listed IHC rating (HPA antibodies). Validation in one application does not supply an IHC rating for the other antibody. |
| Isoforms and antibody recognition | Crk-II and Crk-I are listed isoforms (UniProt P46108). The supplied record gives no antibody epitope or isoform recognition data, so a difference between preparations cannot be assigned to an isoform from staining alone. |
| IF/ICC Q: Should its pattern match tissue IHC exactly? | No exact match is established: plasma membrane is the main approved ICC-IF location; actin filaments and basal body are additional approved locations, while nucleoplasm is uncertain (HPA subcellular). Interpret those images within ICC-IF, separately from the tissue IHC profile (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive cells are blank. | The section or detection run may have failed; High staining is reported in duodenal glandular cells and hepatocytes (HPA tissue IHC). | Run a known-positive section alongside the sample and confirm that the detection reagents and counterstain worked (general IHC practice). |
| Only nuclei stain strongly. | A nuclear-dominant tissue pattern does not match the reported membranous and cytoplasmic IHC profile (HPA tissue IHC). | Inspect a no-primary control and repeat with an IHC-validated antibody if available (general IHC practice; HPA antibodies). |
| An HPA Not detected cell population stains strongly. | Adipose-tissue adipocytes and bone-marrow hematopoietic cells are reported Not detected (HPA tissue IHC); cross-reactivity or endogenous activity is possible (general IHC practice). | Check a no-primary control and whether background follows the cells or the detection chemistry (general IHC practice). |
| Chromogen is diffuse across the section. | Background can obscure the cell boundaries needed to judge CRK's tissue pattern (HPA tissue IHC; general IHC practice). | Compare with a no-primary control, then review blocking, washes and detection development using the assay controls (general IHC practice). |
| Colon glandular cells look weaker than kidney tubular cells. | Colon glandular cells are Medium and kidney tubular cells are High in the HPA profile (HPA tissue IHC). | Score each named population against its reported level before treating the difference as an assay failure (HPA tissue IHC). |
| An ICC-IF image shows nucleoplasmic signal but the IHC section does not. | Nucleoplasmic ICC-IF localisation is uncertain, while tissue IHC reports membranous and cytoplasmic staining (HPA subcellular; HPA tissue IHC). | Evaluate each application with its own validated antibody and controls; do not use the uncertain ICC-IF location as an IHC acceptance criterion (HPA antibodies; HPA subcellular). |
Experiment evidence
IHC & IF images showing CRK staining
PB9101 · IHCIHC analysis of Crk p38 using anti-Crk p38 antibody (PB9101). Crk p38 was detected in a paraffin-embedded section of human lung cancer tissu
PB9101 · IHCIHC analysis of Crk p38 using anti-Crk p38 antibody (PB9101). Crk p38 was detected in a paraffin-embedded section of mouse brain tissue. Hea
M02533-3 · IFIF analysis of CRKII using anti-CRKII antibody (M02533-3) and anti-Beta Tubulin antibody (M01857-3). CRKII was detected in immunocytochemica
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000167193-CRK/tissue/cerebral+cortex#imid_3638020
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000167193-CRK/tissue/colon#imid_3637930
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000167193-CRK/tissue/liver#imid_3637939
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000167193-CRK/tissue/kidney#imid_3637948
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA068087; green = CRK, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA068087; green = CRK, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CRK IHC & IF
🧪Run duodenum first and look for staining in its glandular cells (HPA: High in duodenal glandular cells). Use adipose tissue adipocytes as the negative comparator (HPA: Not detected in adipocytes); on the duodenum slide, check adjacent non-glandular cells for background, without treating them as a validated CRK-negative population (HPA: no internal-negative annotation supplied).
Positive control tissue: Duodenum (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CRK in A-549, HaCaT, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Plasma membrane (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit IgG isotype control for the rabbit catalog antibody (hero caption: rabbit primary and anti-rabbit secondary; standard IHC practice). Use CRK knockout material as a biological specificity control where available, and block endogenous peroxidase before HRP–DAB detection while checking the duodenum no-primary slide for tissue background (hero caption: HRP–DAB detection; standard IHC practice).
⚠️Feasibility: A CRK-specific fixation window or fixation effect is unreported, and the selected PB9101 paraffin-section caption does not state the fixative (hero caption: fixative not stated). Heat retrieval in EDTA at pH 8.0 was used in that caption, but whether CRK staining depends on retrieval is unreported (hero caption: EDTA retrieval). The supplied evidence does not establish that frozen sections or IF are easier; for duodenum, inspect luminal material and glandular background against the no-primary control (HPA: High in duodenal glandular cells; standard IHC practice).
HPA tissue IHC evidence for CRK
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data. Presumed off target binding observed and disregarded.). 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 |
|---|---|---|---|---|
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | High | Protein (IHC) | HPA → |
| Liver | Hepatocytes | High | Protein (IHC) | HPA → |
| Salivary gland | Glandular cells | High | Protein (IHC) | HPA → |
| Small intestine | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CRK IHC Tips
Troubleshoot CRK staining in paraffin sections by checking retrieval, compartment, cell identity and controls before comparing staining intensity.
How should I adjust retrieval when CRK staining is weak or uneven?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet PB9101). The PB9101 tissue image used this retrieval before incubation with 2 μg/ml primary antibody overnight at 4°C (datasheet PB9101). If staining is weak, vary heating and cooling conditions on matched sections while keeping antibody concentration and detection steps fixed (standard IHC practice). Include a section with no primary antibody to distinguish retrieval-related background from specific staining (standard IHC practice). Compare cytoplasmic and membranous staining in the same cell populations, because both patterns are reported for CRK (HPA tissue IHC; UniProt P46108 subcellular location).
Could fixation explain weak CRK staining in my paraffin sections?
CRK-specific sensitivity to fixation is unknown from the supplied evidence: the PB9101 paraffin-section caption does not state a fixative (datasheet PB9101). Record the specimen fixative and processing history, then compare sections processed under known conditions before attributing a staining difference to CRK abundance (standard IHC practice). Keep retrieval at EDTA pH 8.0 while assessing fixation as a variable (datasheet PB9101). Overfixation or uneven fixation can reduce accessible epitopes in paraffin-section IHC generally, but its effect on CRK here has not been established (standard IHC practice; datasheet PB9101). Use tissue morphology and an independently validated control stain to assess processing quality (standard IHC practice).
What CRK staining pattern should I expect in tissue sections?
Assess cytoplasmic and membranous signal first, because both are reported in tissue IHC and CRK is annotated in the cytoplasm and at the cell membrane (HPA tissue IHC; UniProt P46108 subcellular location). Membrane enrichment can vary with cell adhesion, which is associated with CRK translocation to the plasma membrane (UniProt P46108 subcellular location). CRK has no annotated transmembrane segment, so membranous staining should not by itself be interpreted as evidence of an integral membrane protein (UniProt P46108 topology). HPA also reports plasma membrane localisation in ICC/IF, with nucleoplasmic localisation marked uncertain (HPA subcellular). Review an unexpected nuclear-dominant IHC pattern against morphology, controls and antibody specificity before scoring it as CRK (HPA subcellular; standard IHC practice).
Can this stain distinguish Crk-I from Crk-II or report phosphorylation?
CRK has 2 annotated isoforms, Crk-I and Crk-II, so isoform attribution requires epitope information or independent validation (UniProt P46108 isoforms; standard IHC practice). The supplied PB9101 caption gives a staining workflow but no epitope mapping or isoform selectivity (datasheet PB9101). CRK contains an SH2 domain at residues 13–118 and SH3 domains at 132–192 and 235–296; these annotations alone do not identify what PB9101 detects (UniProt P46108 domains; datasheet PB9101). Phosphoserines are annotated at several sites, including residues 40 and 41, but ordinary CRK IHC cannot assign a signal to a phosphorylation state without phospho-specific validation (UniProt P46108 modified residues; standard IHC practice). Score the result as CRK immunoreactivity unless that additional validation is available (standard IHC practice).
How can IF help resolve ambiguous CRK localisation seen by IHC?
Use a cell-type marker relevant to the compartment being assessed, then examine whether CRK signal overlaps the expected cells rather than relying on tissue-wide fluorescence (HPA tissue IHC; standard IF practice). HPA reports CRK mainly at the plasma membrane in ICC/IF and additionally at actin filaments and basal bodies, while nucleoplasmic localisation is uncertain (HPA subcellular). Choose fluorophores and acquisition settings after checking tissue autofluorescence with an unstained section, and separate emission channels for multiplexing (standard IF practice). For an epitope accessible from the cytoplasmic side of the membrane, optimise permeabilisation without assuming CRK spans the membrane; its topology has no transmembrane segment (UniProt P46108 topology; standard IF practice). Confirm the IF pattern with appropriate single-label and secondary-only controls (standard IF practice).
How do I troubleshoot diffuse brown staining in CRK IHC?
First compare the stained section with a no-primary control to assess signal from the detection system and tissue itself (standard IHC practice). The PB9101 image used 10% goat serum blocking, a peroxidase-conjugated secondary and DAB development (datasheet PB9101). A peroxidase block and careful DAB development are general chromogenic IHC steps; the caption does not establish that CRK requires a particular block duration (standard IHC practice; datasheet PB9101). If diffuse staining persists, check washing, secondary-antibody background and whether chromogen development obscures cellular boundaries (standard IHC practice). Evaluate putative CRK signal within intact cells, because HPA reports membranous and cytoplasmic expression but also notes presumed off-target binding in its tissue assessment (HPA tissue IHC).
How should I score CRK staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and scoring compartment before examining group differences, because CRK can appear in both cytoplasm and membrane (HPA tissue IHC; UniProt P46108 subcellular location). For cell-based scoring, record the percentage of positive cells and staining intensity, or use an H-score spanning 0–300 with prespecified intensity categories (standard IHC practice). If counting discrete positive cells, report density per mm² of viable evaluable tissue and state how regions were selected (standard IHC practice). Normalise comparisons to the same cell population, evaluable area and staining batch, using reference sections where possible (standard IHC practice). Keep membrane and cytoplasmic scores separate when redistribution is the biological question, since adhesion can shift CRK toward the plasma membrane (UniProt P46108 subcellular location).
How can I separate true CRK staining from tissue artefact?
A credible result has cellular boundaries and a cytoplasmic or membranous distribution consistent with reported CRK patterns (HPA tissue IHC; UniProt P46108 subcellular location). Check cell identity before calling a field negative: HPA reports high staining in kidney tubular cells and liver hepatocytes, while several other listed cell populations are not detected (HPA tissue IHC). Discount staining restricted to section edges, folds or necrotic areas, and compare suspect DAB signal with a no-primary and peroxidase-block control (standard IHC practice). A nuclear-dominant pattern needs particular scrutiny because HPA labels nucleoplasmic CRK localisation uncertain in ICC/IF (HPA subcellular). Interpret a clean pattern cautiously: the HPA tissue assessment is approved but notes medium RNA–protein consistency and presumed off-target binding (HPA tissue IHC).
Boster reagents
Best CRK / Adapter molecule crk IHC Antibodies
Anti-CRK antibodies have IHC images from paraffin sections of human lung cancer and mouse and rat brain (PB9101 image captions), plus an IF image from HeLa cells (M02533-3 image caption).
Real IHC data
Anti-Crk p38 Antibody ®
Cat # PB9101
Real IF data
Anti-CRKII Rabbit Monoclonal Antibody
Cat # M02533-3
PB9101 has IHC images from paraffin sections of human lung cancer and mouse and rat brain (PB9101 image captions). M02533-3 has an IF image from HeLa cells (M02533-3 image caption) and is listed for ICC/IF (catalog applications).
Which to pick: Choose PB9101 for tissue IHC: it is listed for IHC in human, mouse and rat (catalog applications and reactivity), and its images show paraffin sections from all three species (PB9101 image captions). Choose M02533-3, a rabbit monoclonal, for IF/ICC (catalog: clone 24C53; catalog applications); its reactivity lists human, mouse and rat, while the supplied IF image shows HeLa cells (catalog reactivity; M02533-3 image caption). PB9101's captions support paraffin sections; the fixative is unreported (PB9101 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P46108 (CRK_HUMAN, Adapter molecule crk).
- Human Protein Atlas. CRK tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CRK subcellular location (ICC-IF): Mainly localized to the plasma membrane. In addition localized to the nucleoplasm, actin filaments and basal body..
- Human Protein Atlas. CRK antibody validation summary (2 antibodies).
- MiR-194-5p inhibited metastasis and EMT of nephroblastoma cells through targeting Crk. The Kaohsiung journal of medical sciences 2020 — PMC11896132.
- Significance of KRAS/PAK1/Crk pathway in non-small cell lung cancer oncogenesis. BMC cancer 2015 — PMC4477307.
- Adaptor protein CRK induces epithelial-mesenchymal transition and metastasis of bladder cancer cells through HGF/c-Met feedback loop. Cancer science 2015 — PMC4471787.
- Increased cellular distribution of vimentin and ret in the cingulum of rat offspring after developmental exposure to decabromodiphenyl ether or 1,2,5,6,9,10-hexabromocyclododecane. Journal of toxicologic pathology 2013 — PMC3695334.
- PubMed PMID:1630456 — UniProt-cited evidence.
- PubMed PMID:8378094 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CRK (Adapter molecule crk). Expected localisation Cytoplasmic and membranous staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet PB9101). Validated IHC antibody: PB9101 at 2-5μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Duodenum (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet PB9101); 2. Fixation: Image fixative and duration unreported (datasheet PB9101); 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 PB9101); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet PB9101); 8. Primary antibody: Rabbit anti-CRK, 2-5μg/ml (datasheet PB9101); 9. Primary incubation: Overnight at 4 °C (datasheet PB9101). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Presumed off-target staining can confound scoring (HPA tissue IHC); Isoform / epitope: Two isoforms; verify antibody epitope coverage (UniProt); Regulation: Adhesion recruits CRK to the plasma membrane (UniProt). Evidence: UniProt P46108, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the plasma membrane. In addition localized to…, Datasheet PB9101, PMC11896132, PMC4477307. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CRK/immunohistochemistry-ihc-guide.