This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
UCN / Urocortin · IHC design guide
Design Immunohistochemistry for UCN
Plan chromogenic UCN staining in paraffin sections using the catalog antibodies’ IHC dilutions of 1:50–1:200 (datasheet: A01807) or 1:100–1:300 (datasheet: A01807-1). Interpret cytoplasmic and plasma staining (HPA tissue IHC) in light of UCN secretion (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
UCN 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 | Cytoplasm and plasma signal (HPA tissue IHC) | |
| Staining pattern | Broad cytoplasmic staining with plasma staining (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adipose tissue+4 more · see all | |
| Negative control | None in HPA (detected in all 45 tissues); use no-primary + isotype controls |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Secretion can separate protein and RNA tissue patterns (HPA tissue IHC) | |
| Regulation | Released in response to stress (UniProt) | |
| Isoform / epitope | No isoforms; signal peptide and propeptide are processed (UniProt) |
Section 1
Recommended UCN IHC & IF Protocols
Compare the catalog antibody’s paraffin-section IHC protocol with four published UCN chromogenic IHC workflows (datasheet; PMC10761541; PMC2826606; PMC2982926; PMC3168230).
| Sample | Paraffin-embedded human colorectal carcinoma tissue; fixative not specified (datasheet A01807) |
| Fixation | Image fixative and duration unreported (datasheet A01807); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-UCN, 1:50-1:200 (datasheet A01807) |
| 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 | UCN-positive staining in adipocytes of adipose tissue (HPA tissue IHC: Medium). HPA tissue profile: Ubiquitous cytoplasmic expression with additional plasma staining. No signal in the no-primary control. |
💡Decision noteFor paraffin sections, start with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page antigen retrieval rule); optimize against controls.
Section 2
What Is the Expected UCN Staining Pattern?
UCN is a secreted protein with no transmembrane segment (UniProt P55089 topology). In paraffin sections, expect predominantly cytoplasmic staining with additional plasma staining (HPA tissue IHC: Approved). Medium staining is reported in adrenal glandular cells, adipocytes and caudate neuronal cells (HPA tissue IHC: Medium). Skin keratinocytes and follicular sheaths are also reported sites (UniProt P55089 tissue specificity). Because UCN is secreted, the site of protein staining need not match the site of RNA expression (HPA tissue IHC: reliability description).
| Cytoplasmic chromogen in adrenal glandular cells, with possible plasma staining (HPA tissue IHC: Medium; profile). | This fits the reported UCN pattern (HPA tissue IHC: Approved). Judge signal by its location and the stained cells together: plasma staining can accompany the cellular pattern, while UCN secretion means protein and RNA maps may differ (UniProt P55089 subcellular location; HPA tissue IHC: reliability description). |
| Strong, predominantly nuclear staining, with little cytoplasmic signal. | Treat this as a compartment mismatch: the reported tissue pattern is cytoplasmic, and ICC-IF places UCN mainly in vesicles with additional plasma membrane localisation (HPA tissue IHC: profile; HPA subcellular: approved). Review controls and staining distribution before scoring it as UCN. |
| Staining is concentrated in an unexpected cell population while expected cells are unstained. | Consider antibody cross-reactivity or endogenous chromogenic activity (general IHC practice). Compare the same slide with its expected cell populations: adrenal glandular cells, adipocytes and caudate neuronal cells have Medium staining in HPA; rectal glandular and lymph-node germinal-center cells are reported Low, not negative (HPA tissue IHC). |
| Broad, diffuse colour obscures cell boundaries or appears similarly across compartments. | This is difficult to score as a cellular UCN pattern (HPA tissue IHC: cytoplasmic profile). Secreted UCN can also produce plasma staining, so extracellular colour alone does not prove artefact (UniProt P55089 subcellular location; HPA tissue IHC: profile). Use controls to assess nonspecific background (general IHC practice). |
| No visible signal in an HPA-reported Medium tissue, such as adrenal glandular cells (HPA tissue IHC). | The result does not establish absent UCN. Check whether the control section and detection worked, then assess antibody concentration and antigen retrieval as general IHC variables (general IHC practice). The supplied sources give no target-specific fixation sensitivity or retrieval condition for UCN. |
💡Expected UCN appearanceCall a result consistent with UCN when cytoplasmic staining is discernible in an HPA-reported Medium cell population, such as adrenal glandular cells, with possible plasma staining (HPA tissue IHC: Approved; Medium; profile); dominant nuclear signal or uniform colour across unrelated cells warrants control-based review (HPA subcellular: approved; general IHC practice).
| Secretion and processing (UniProt P55089) | UCN has a signal peptide at residues 1–25, a propeptide at 26–82 and an amidated residue at 122 (UniProt P55089 processing). Antibody epitope location is unspecified here, so these annotations cannot predict which processed forms the antibody detects or prescribe a retrieval condition. |
| Tissue and cell context (HPA tissue IHC; UniProt P55089 tissue specificity) | HPA reports Medium staining in several tissues and Low staining in rectal glandular and lymph-node germinal-center cells; it lists no negative tissue (HPA tissue IHC). UniProt additionally reports skin structures and colonic lamina propria plasma cells (UniProt P55089 tissue specificity). Do not confuse those cells with HPA's plasma staining. |
| Evidence and antibody choice (HPA tissue IHC; HPA antibodies) | The tissue profile has Approved reliability, with a caution that RNA and protein locations may differ for secreted proteins (HPA tissue IHC). CAB010911 is Approved for IHC; the supplied antibody list does not report an Enhanced IHC result (HPA antibodies). These ratings support interpretation, not a guaranteed result in every section. |
| Antigen retrieval (general IHC practice) | Retrieval conditions can be assessed with a positive control when chromogenic IHC signal is weak (general IHC practice). Neither supplied source identifies a UCN-specific retrieval condition or shows that fixation changes UCN epitope access, sensitivity or tissue positivity; avoid assigning a target-specific cause to a weak result. |
| IF/ICC Q: Where should UCN signal appear? | A: Mainly in vesicles, with additional plasma membrane localisation (HPA subcellular: approved). HPA lists ICC-IF images from SK-MEL-30, THP-1 and U2OS, and HPA069971 as Approved for ICC (HPA subcellular; HPA antibodies). These ICC-IF observations are a localisation reference, not a paraffin-section protocol. |
| Situation | Likely cause | Next action |
|---|---|---|
| Adrenal glandular cells show no stain despite a working counterstain (HPA tissue IHC: Medium). | A counterstain alone does not verify antibody detection; low assay sensitivity or an unsuitable staining condition remains possible (general IHC practice). | Run a reported Medium tissue as a positive control and check the antibody and detection steps (HPA tissue IHC: Medium; general IHC practice). Optimise retrieval only as a general IHC variable; UCN-specific retrieval and fixation effects are unreported here. |
| Diffuse colour persists in a section processed without primary antibody. | The primary antibody cannot account for colour in that control; detection-system background or endogenous activity is possible (general IHC practice). | Check blocking and detection controls; if using peroxidase chromogen, assess endogenous peroxidase blocking (general IHC practice). Reassess cellular UCN staining against the reported cytoplasmic pattern (HPA tissue IHC: profile). |
| Many unrelated cells stain equally, obscuring the expected distribution. | Nonspecific antibody binding or excessive chromogen development may reduce cell-level discrimination (general IHC practice). HPA's broad cytoplasmic profile means breadth alone is insufficient to diagnose artefact (HPA tissue IHC: profile). | Compare positive and no-primary controls, review antibody concentration and detection time, and score by cell type and compartment (general IHC practice; HPA tissue IHC: profile). |
| Nuclear colour dominates while cytoplasmic staining is absent. | This conflicts with HPA's cytoplasmic tissue profile and vesicular ICC-IF localisation (HPA tissue IHC: profile; HPA subcellular: approved). | Check no-primary and positive controls, then review chromogen and counterstain separation before interpreting nuclear colour as target signal (general IHC practice). |
| A Low tissue stains more strongly than an HPA-reported Medium tissue. | Section handling, detection conditions or biological variation may affect comparisons (general IHC practice); HPA's Low and Medium labels describe its reported staining, not fixed thresholds for every run (HPA tissue IHC). | Compare matched sections within the same run and confirm the stained cell types before changing the UCN interpretation (general IHC practice; HPA tissue IHC). |
| Protein staining appears away from the cell population suggested by RNA data. | UCN is secreted, and HPA explicitly expects RNA and protein tissue locations to differ (UniProt P55089 subcellular location; HPA tissue IHC: reliability description). | Assess cytoplasmic cell staining and possible plasma staining against the HPA protein profile; use staining controls to investigate a discordant result rather than treating RNA location alone as the IHC reference (HPA tissue IHC: profile; general IHC practice). |
Experiment evidence
IHC & IF images showing UCN staining
A01807 · IHCImmunohistochemistry (IHC) analyzes of UCN (A103) pAb in paraffin-embedded human colorectal carcinoma tissue at 1:100.
A01807-1 · IHCImmunohistochemistry analysis of paraffin-embedded human lung carcinoma tissue, using Urocortin Antibody. The picture on the right is blocke
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000163794-UCN/tissue/cerebral+cortex#imid_3181272
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000163794-UCN/tissue/colon#imid_3181206
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000163794-UCN/tissue/liver#imid_3181207
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000163794-UCN/tissue/kidney#imid_3181208
HPA · ICC-IF SK-MEL-30Human Protein Atlas immunofluorescence, SK-MEL-30, antibody HPA069971; green = UCN, blue = nucleus, red = microtubules.
HPA · ICC-IF SK-MEL-30Human Protein Atlas immunofluorescence, SK-MEL-30, antibody HPA069971; green = UCN, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for UCN IHC & IF
🧪Run adrenal gland first and assess staining in its glandular cells (HPA: Medium in adrenal gland glandular cells; UniProt P55089: expressed in adrenal glands). HPA detects UCN in all 45 scored tissues, so there is no HPA-supported negative tissue; use no-primary and isotype controls, and assess morphologically distinct non-glandular cells on the same slide for background-level staining where present (HPA: no negative tissue rows; standard IHC practice).
Positive control tissue: Adipose tissue (Adipocytes, HPA Medium)
Negative control tissue: None in HPA: UCN is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show UCN in SK-MEL-30, THP-1, U2OS, with annotated localisation: Vesicles (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only section and a control immunoglobulin matched to the primary antibody’s host species and, for a monoclonal antibody, its isotype; assess target specificity with UCN knockout material if available or peptide competition if the immunizing peptide is available (standard IHC practice). For chromogenic adrenal IHC, quench endogenous peroxidase and check for endogenous biotin background if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: The selected A01807 caption documents paraffin-section IHC in human colorectal carcinoma at 1:100, but its fixative is unreported; the supplied evidence reports no target-specific fixation window or effect (selected A01807 tissue-IHC caption). Antigen-retrieval dependence and whether frozen sections or IF are easier are unreported in the supplied evidence; HPA does show ICC-IF images for UCN in SK-MEL-30, THP-1 and U2OS (HPA subcellular record). Check adrenal sections for endogenous peroxidase background when interpreting chromogenic staining (standard IHC practice).
HPA tissue IHC evidence for UCN
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Secreted protein, tissue location of RNA and protein is expected to differ.). 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 |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Breast | Adipocytes | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: UCN is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced UCN IHC Tips
Troubleshoot chromogenic UCN staining in paraffin sections by checking retrieval, controls, cellular location, and scoring before interpreting tissue differences.
What retrieval should I start with for weak UCN staining in paraffin sections?
Start with citrate buffer at pH 6.0, heated to 95–98 °C for 20 minutes (page retrieval setting). Let sections cool in buffer before washing, and keep retrieval conditions identical across comparison slides to limit technical variation (standard IHC practice). If staining remains weak, compare a modestly longer retrieval time on serial sections while watching for tissue damage; treat that change as an experiment, not a UCN-specific requirement (standard IHC practice). Include a tissue with documented UCN staining, such as adrenal glandular cells, and a no-primary control so weak signal can be distinguished from failed detection or background (HPA: Medium in adrenal glandular cells; standard IHC practice).
Could fixation explain weak or uneven UCN staining?
UCN-specific sensitivity to fixation is unknown from the supplied evidence, so do not assign a weak result to a particular fixative without a controlled comparison. The selected antibody image shows paraffin-embedded colorectal carcinoma at 1:100, but its caption does not report the fixative (caption: A01807). Record fixation method and duration for each specimen, then compare similarly processed sections under the same citrate pH 6.0, 95–98 °C, 20-minute retrieval conditions (page retrieval setting; standard IHC practice). Review tissue preservation and include a no-primary control before changing antibody concentration, because uneven processing and detection background can mimic differences in staining (standard IHC practice).
Where should convincing UCN signal appear within stained cells and tissue?
Look for cytoplasmic staining, potentially concentrated in vesicular structures, while allowing some plasma membrane signal (HPA: ubiquitous cytoplasmic expression; HPA subcellular: vesicles, with additional plasma membrane localization). UCN is a secreted protein with no transmembrane segment, so a exclusively crisp membrane outline would need stronger control evidence (UniProt P55089: secreted; no transmembrane segment). Additional plasma staining may occur and should be recorded separately from cellular staining (HPA: additional plasma staining). Examine sections at high magnification to distinguish intracellular deposits from extracellular pigment, and compare the same compartment in a no-primary control before scoring it as UCN (standard IHC practice).
How can UCN processing affect interpretation of an antibody epitope?
UCN is recorded as a 124-amino-acid precursor with a signal peptide at residues 1–25 and a propeptide at 26–82 (UniProt P55089: sequence and processing). The supplied record lists no isoforms and notes an amidated valine at residue 122 (UniProt P55089: isoforms and modified residues). Check the antibody’s documented immunogen or epitope before assuming that staining detects precursor and processed UCN equally; the supplied caption does not specify that epitope (caption: A01807). Compare staining with an independently characterized antibody or an appropriate specificity control where available, and report the antibody-defined signal without assigning it to a molecular form that the epitope evidence cannot resolve (standard IHC practice).
How should I adapt the UCN question to a multiplex IF experiment?
Plan IF as a separately optimized assay: the selected antibody evidence here is a paraffin-section IHC image at 1:100, with no fixative reported (caption: A01807). Pair UCN with a validated marker of the expected cell population, such as a plasma-cell marker when investigating colon mucosa, and confirm that both channels identify the intended cells (UniProt P55089: UCN detected in lamina propria plasma cells; standard IF practice). Choose fluorophores after inspecting tissue autofluorescence, favoring spectrally separated, brighter channels for weak signal (standard IF practice). Because UCN is secreted and mainly vesicular, optimize permeabilisation for access to intracellular epitopes, then check whether that treatment alters apparent membrane staining (UniProt P55089: secreted; HPA subcellular: vesicles and plasma membrane; standard IF practice).
How do I reduce diffuse UCN chromogen without losing cellular signal?
Run a no-primary slide beside the UCN slide to identify secondary-reagent or detection background before changing the antibody (standard IHC practice). Use a peroxidase block before horseradish-peroxidase detection, block nonspecific binding, and wash consistently between incubations (standard chromogenic IHC practice). If diffuse DAB persists, titrate the primary around the documented 1:100 image condition while holding development time and section processing constant; that caption is an example, not a validated optimum for every specimen (caption: A01807; standard IHC practice). Because HPA describes cytoplasmic staining with additional plasma staining, assess extracellular signal separately rather than subtracting all diffuse color as artefact (HPA: tissue IHC profile).
What should I measure when comparing UCN staining across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then apply the same threshold and imaging conditions to every section (standard IHC practice). For cellular UCN, report percent-positive cells and an H-score based on staining intensity; normalize cell counts to the number of evaluable cells in the defined region (standard IHC practice). If measuring extracellular or plasma-associated deposits, report positive area or optical density per mm² of evaluable tissue separately from cellular scores (HPA: additional plasma staining; standard IHC practice). Exclude folds, necrosis, and section edges by a prespecified rule, and compare biological replicates rather than treating multiple fields from one section as independent specimens (standard IHC practice).
Which apparent UCN-positive patterns deserve extra skepticism?
Give greatest weight to reproducible cellular staining in a plausible location, such as adrenal glandular cells or colon lamina propria plasma cells (HPA: Medium in adrenal glandular cells; UniProt P55089: colon plasma-cell detection). UCN is secreted, and HPA reports cytoplasmic staining with additional plasma staining, so extracellular color alone does not establish which nearby cell produced it (UniProt P55089: secreted; HPA: tissue IHC profile). Treat an exclusively nuclear pattern, edge-only signal, or staining restricted to necrotic regions as suspect until controls support it (HPA subcellular: vesicles and plasma membrane; standard IHC practice). Check no-primary and peroxidase-block controls for endogenous enzyme or detection artefacts before interpreting differences between specimens (standard chromogenic IHC practice).
Boster reagents
Best UCN / Urocortin IHC Antibodies
Two anti-UCN antibodies have real IHC figures from paraffin-embedded human carcinoma tissue; both list Human, Mouse and Rat reactivity, while one also lists IF/ICC applications (catalog; IHC image captions).
Real IHC data
Anti-UCN (A103) Antibody
Cat # A01807
Real IHC data
Anti-Urocortin UCN Antibody
Cat # A01807-1
A01807 lists IHC and shows staining in paraffin-embedded human colorectal carcinoma tissue at 1:100 (catalog; A01807 IHC image caption). A01807-1 lists IHC, IF and ICC and shows staining in paraffin-embedded human lung carcinoma tissue with a peptide-blocked comparison (catalog; A01807-1 IHC image caption).
Which to pick: For tissue IHC, choose either SKU according to the relevant illustrated sample: A01807 for paraffin-embedded colorectal carcinoma tissue or A01807-1 for paraffin-embedded lung carcinoma tissue (respective IHC image captions). For IF/ICC, choose A01807-1 because those applications are listed only for it; no IF figure is supplied (catalog). Both list Human, Mouse and Rat reactivity, but their figures show human tissue only; neither IHC caption reports the fixative (catalog; respective IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P55089 (UCN1_HUMAN, Urocortin).
- Human Protein Atlas. UCN tissue IHC expression (reliability: Approved).
- Human Protein Atlas. UCN subcellular location (ICC-IF): Mainly localized to vesicles. In addition localized to the plasma membrane..
- Human Protein Atlas. UCN antibody validation summary (2 antibodies).
- The expression of corticotropin-releasing hormone family peptides in premalignant and malignant vulvar lesions. Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico 2024 — PMC10761541.
- Postnatal developmental profile of urocortin 1 and cocaine- and amphetamine-regulated transcript in the perioculomotor region of C57BL/6J mice. Brain research 2010 — PMC2826606.
- Expression of urocortin peptides in canine myocardium and plasma. Veterinary journal (London, England : 1997) 2011 — PMC2982926.
- Urocortin-like immunoreactivity in the primary lymphoid organs of the duck (Anas platyrhynchos). European journal of histochemistry : EJH 2009 — PMC3168230.
- PubMed PMID:8612563 — UniProt-cited evidence.
- PubMed PMID:9628819 — UniProt-cited evidence.
- PubMed PMID:15815621 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for UCN (Urocortin). Expected localisation Cytoplasm and plasma signal (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A01807 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adipose tissue (Adipocytes); negative None in HPA (detected in all 45 tissues); use no-primary + isotype controls. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colorectal carcinoma tissue; fixative not specified (datasheet A01807); 2. Fixation: Image fixative and duration unreported (datasheet A01807); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-UCN, 1:50-1:200 (datasheet A01807); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Secretion can separate protein and RNA tissue patterns (HPA tissue IHC); Isoform / epitope: No isoforms; signal peptide and propeptide are processed (UniProt); Regulation: Released in response to stress (UniProt). Evidence: UniProt P55089, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to vesicles. In addition localized to the plasma…, Datasheet A01807, PMC10761541, PMC2826606. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/UCN/immunohistochemistry-ihc-guide.