This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
CLCF1 / Cardiotrophin-like cytokine factor 1 · IHC design guide
Design Immunohistochemistry for CLCF1
Plan chromogenic CLCF1 IHC around the reported cytoplasmic tissue pattern and plasma positivity (HPA tissue IHC). Bone marrow hematopoietic cells and lung macrophages show medium staining, but HPA rates the tissue profile uncertain because secreted protein may differ from local RNA expression (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CLCF1 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 staining in several tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in several tissues; plasma positive (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Bone marrow+4 more · see all | |
| Negative control | Caudate+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep formalin fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Secretion can separate tissue RNA from protein staining (HPA tissue IHC) | |
| Regulation | CRLF1/CNTFR-dependent secretion (UniProt) | |
| Isoform / epitope | 2 isoforms; signal peptide 1–27; epitope impact unknown (UniProt) |
Section 1
Recommended CLCF1 IHC & IF Protocols
The catalog antibody IHC-P protocol is accompanied by published CLCF1 IHC methods for mouse IRI tissue (PMC9073803) and human kidney tissue (PMC12305993).
| Sample | Paraffin-embedded human brain tissue; fixative not specified (datasheet A08886) |
| Fixation | Image fixative and duration unreported (datasheet A08886); 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-CLCF1, 1:50 recommended; image 1:200 (datasheet A08886) |
| 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 | CLCF1-positive staining in hematopoietic cells of bone marrow (HPA tissue IHC: Medium). HPA tissue profile: Cytoplasmic expression in several tissues and positivity in plasma. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval); neither published IHC method specifies retrieval-buffer pH (PMC9073803; PMC12305993).
Section 2
What Is the Expected CLCF1 Staining Pattern?
CLCF1 is a secreted cytokine with no transmembrane segment (UniProt Q9UBD9 topology). In paraffin sections, expect mainly cytoplasmic staining in selected cells, including bone marrow hematopoietic cells and lung macrophages; HPA also reports positivity in plasma (HPA tissue IHC). Treat this as a provisional pattern: HPA rates its tissue staining Uncertain and notes that a secreted protein’s RNA and protein locations may differ (HPA tissue IHC).
| Cytoplasmic signal in bone marrow hematopoietic cells or lung macrophages. | This fits HPA’s Medium staining in both cell populations (HPA tissue IHC). Judge the labelled cells and their compartment separately from overall slide colour; the reported pattern has Uncertain reliability and awaits external verification (HPA tissue IHC). |
| Signal is confined to nuclei, with little cytoplasmic staining in an IHC section. | That departs from the reported tissue IHC profile of cytoplasmic expression (HPA tissue IHC). Check morphology and detection controls before calling it CLCF1. Nuclear bodies are an approved ICC-IF location, so the IF observation alone cannot validate nuclear IHC staining (HPA subcellular; HPA tissue IHC). |
| Strong staining appears in caudate glia or prostate glandular cells. | HPA reports CLCF1 as Not detected in those respective cell populations (HPA tissue IHC). Investigate antibody cross-reactivity, endogenous detection activity, or tissue identification before accepting an unexpected positive; an HPA negative is a reference observation, not proof that every specimen must be negative (HPA tissue IHC). |
| Colour spreads across unrelated cells, extracellular spaces, and blank areas. | A uniform haze is less persuasive than cell-associated cytoplasmic staining (HPA tissue IHC). Secreted CLCF1 and reported plasma positivity complicate extracellular interpretation (UniProt Q9UBD9 subcellular; HPA tissue IHC); compare background controls and morphology before assigning the signal to producing cells (general IHC practice). |
| No signal appears in bone marrow hematopoietic cells on a stained section. | HPA reports Medium staining there, so inspect the section, antibody and detection controls, and staining conditions (HPA tissue IHC; general IHC practice). This is a useful reference tissue, but HPA’s Uncertain reliability does not make a negative result conclusive (HPA tissue IHC). |
💡Expected CLCF1 appearanceA plausible positive is predominantly cytoplasmic staining in the appropriate cells at a Medium reference level, such as bone marrow hematopoietic cells or lung macrophages; indiscriminate haze or strong signal in HPA Not detected populations needs investigation (HPA tissue IHC; general IHC practice).
| Secretion and tissue location (UniProt Q9UBD9; HPA tissue IHC) | CLCF1 is secreted, and HPA notes plasma positivity and possible RNA–protein location differences (UniProt Q9UBD9 subcellular; HPA tissue IHC). Cytoplasmic staining can support a cellular pattern, but extracellular signal cannot by itself identify the producing cell (general IHC interpretation). |
| Mature protein and signal peptide (UniProt Q9UBD9 processing) | The precursor has a signal peptide at residues 1–27; the annotated mature chain spans 28–225 (UniProt Q9UBD9 processing). Epitope location would affect which form an antibody recognizes, but the supplied evidence does not identify this antibody’s epitope. |
| Isoforms and glycosylation (UniProt Q9UBD9) | UniProt lists 2 isoforms and one glycosylation site at residue 29 (UniProt Q9UBD9 isoforms; UniProt Q9UBD9 glycosylation). Their effects on this antibody’s paraffin-section staining are unreported; avoid assigning an unexpected pattern to either feature without further evidence. |
| Tissue reference and confidence (HPA tissue IHC) | HPA reports Medium staining in several distinct cell populations and Not detected staining in others, but rates the tissue profile Uncertain pending external verification (HPA tissue IHC). Use these observations as comparison points rather than definitive acceptance criteria. |
| IF/ICC pattern? (HPA subcellular; HPA antibodies) | For IF/ICC, HPA approves nuclear bodies and vesicles as locations and lists antibody HPA042444 as ICC Approved (HPA subcellular; HPA antibodies). Its IHC status is Uncertain; the IF/ICC locations do not establish a paraffin IHC nuclear pattern (HPA antibodies; HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| Reference cells show no detectable chromogen. | The tissue may lack identifiable reference cells, or the IHC staining workflow may have failed (general IHC practice). HPA’s Medium bone marrow signal is an uncertain reference, not a guaranteed result (HPA tissue IHC). | Confirm hematopoietic cells in the section, then check antibody application, detection reagents, and a working assay control (general IHC practice). Interpret any persistent negative alongside HPA’s Uncertain tissue reliability (HPA tissue IHC). |
| Most cells and blank regions stain similarly. | Nonspecific reagent binding or detection background can obscure a cellular pattern (general IHC practice). HPA describes cytoplasmic staining in several tissues and plasma positivity, not uniform staining of the entire section (HPA tissue IHC). | Inspect a no-primary control and review blocking, washes, and chromogen development (general IHC practice). Score cell-associated signal only where morphology and controls permit; avoid treating generalized haze as CLCF1 (general IHC interpretation). |
| An HPA Not detected population stains strongly. | Possible explanations include cross-reactivity, endogenous detection activity, or misidentified cells (general IHC practice). The comparison is against HPA observations for the named population, not a universal absence claim (HPA tissue IHC). | Verify cell identity and compare no-primary and detection controls; assess whether staining tracks the expected cytoplasmic profile in HPA Medium populations (general IHC practice; HPA tissue IHC). |
| Only nuclei stain in paraffin IHC. | A nuclear-only IHC result conflicts with HPA’s cytoplasmic tissue profile (HPA tissue IHC). HPA’s approved nuclear-body localisation comes from ICC-IF, a different application (HPA subcellular; HPA antibodies). | Check counterstain interpretation, tissue morphology, and detection controls before assigning nuclear CLCF1 (general IHC practice). Record the IHC–ICC difference as unresolved if controls are clean (HPA tissue IHC; HPA subcellular). |
| Extracellular or plasma signal dominates the section. | CLCF1 is secreted, and HPA reports plasma positivity; signal away from cells therefore needs careful attribution (UniProt Q9UBD9 subcellular; HPA tissue IHC). Diffuse background remains another possibility (general IHC practice). | Compare signal with vessel and cell morphology and a no-primary control (general IHC practice). Report extracellular staining separately from cytoplasmic staining; do not infer its cell of origin from location alone (UniProt Q9UBD9 subcellular). |
| IF/ICC shows puncta while paraffin IHC appears cytoplasmic. | HPA approves nuclear bodies and vesicles in ICC-IF, while its tissue IHC summary describes cytoplasmic staining (HPA subcellular; HPA tissue IHC). Those application-specific observations need not yield identical images. | Interpret each result using its own application and controls (general IHC/IF practice). Use the dedicated IF/ICC guide for IF design; do not transfer an ICC-approved location into an IHC positive call without supporting IHC evidence (HPA antibodies; HPA tissue IHC). |
Experiment evidence
IHC & IF images showing CLCF1 staining
A08886 · IHCImmunohistochemical analysis of paraffin-embedded human-brain, antibody was diluted at 1:200
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000175505-CLCF1/tissue/cerebral+cortex#imid_11690030
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000175505-CLCF1/tissue/colon#imid_11688220
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000175505-CLCF1/tissue/liver#imid_11688237
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000175505-CLCF1/tissue/kidney#imid_11688238
HPA · ICC-IF SiHaHuman Protein Atlas immunofluorescence, SiHa, antibody HPA042444; green = CLCF1, blue = nucleus, red = microtubules.
HPA · ICC-IF SiHaHuman Protein Atlas immunofluorescence, SiHa, antibody HPA042444; green = CLCF1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CLCF1 IHC & IF
🧪Run bone marrow first and look for CLCF1 staining in hematopoietic cells (HPA: Medium in bone marrow hematopoietic cells); use caudate glial cells as the negative tissue comparator (HPA: Not detected in caudate glial cells). On the marrow slide, treat cells outside the identified hematopoietic population as candidate internal negatives only if their signal matches the no-primary background (HPA: Medium in hematopoietic cells; standard IHC control practice).
Positive control tissue: Bone marrow (Hematopoietic cells, HPA Medium)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CLCF1 in SiHa, U-251MG, U2OS, with annotated localisation: Nuclear bodies (approved), Vesicles (approved) (HPA subcellular).
Technical controls: Include no-primary or secondary-only and host-species- and clonality-matched isotype controls, plus a matched CLCF1 knockout sample if available or a validated immunizing-peptide block (standard IHC control practice). Quench endogenous peroxidase in bone marrow before chromogenic detection, and block endogenous biotin if using avidin–biotin detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported in the supplied evidence; the selected A08886 paraffin-section caption gives 1:200 dilution but does not report the fixative (A08886 tissue-IHC caption). Antigen-retrieval dependence is unreported, so compare retrieved and unretrieved sections under the antibody's IHC-P conditions (standard IHC optimization practice). The supplied evidence does not establish frozen sections or IF as easier; in bone marrow, endogenous peroxidase may produce chromogenic background (standard IHC practice).
HPA tissue IHC evidence for CLCF1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — Secreted protein, tissue location of RNA and protein is expected to differ. Pending external verification.). 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 |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Breast | Myoepithelial cells | Medium | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Medium | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Medium | Protein (IHC) | HPA → |
| Kidney | Cells in tubules | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Salivary gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CLCF1 IHC Tips
Troubleshoot CLCF1 staining in paraffin sections by checking retrieval, compartment, cell identity and controls before scoring chromogenic signal.
How should I retrieve CLCF1 in paraffin sections?
Start with citrate buffer at pH 6.0 for heat induced antigen retrieval at 95–98 °C for 20 minutes (page retrieval specification). Apply the same cooling and wash procedure to every section so differences in chromogenic intensity reflect staining rather than handling (standard IHC practice). The selected A08886 image documents paraffin embedded human brain stained at 1:200, but its caption does not specify retrieval conditions (A08886 tissue IHC caption). If staining remains weak, compare a second retrieval condition on adjacent sections while holding antibody dilution and detection constant (standard IHC practice). Check morphology and the negative control before accepting any gain in signal (standard IHC practice).
Can fixation explain weak CLCF1 staining?
The supplied CLCF1 evidence does not establish target specific fixation sensitivity, so treat fixation as an experimental variable rather than a known cause of weak staining (supplied evidence; standard IHC practice). The A08886 caption identifies paraffin embedded human brain and 1:200 antibody dilution, but does not state the fixative (A08886 tissue IHC caption). Record fixative, fixation duration and processing history for each specimen, then compare sections processed together with identical retrieval and detection (standard IHC practice). Preserve comparable section thickness and use a no primary control when assessing diffuse chromogen (standard IHC practice). Do not infer a fixation response from tissue staining patterns (HPA: tissue reliability uncertain).
Which staining compartments should I expect for CLCF1?
Evaluate cytoplasmic and extracellular staining separately because CLCF1 is secreted and lacks a transmembrane segment (UniProt Q9UBD9). HPA describes cytoplasmic expression across several tissues and positivity in plasma, while rating tissue IHC reliability uncertain because secreted protein can appear away from its RNA source (HPA: tissue profile and reliability). Its cell based imaging also reports nuclear bodies and vesicles; that observation alone does not establish a nuclear tissue IHC pattern (HPA: subcellular summary). Score the stained cell type and compartment together, using morphology and a matched negative control to assess questionable deposits (standard IHC practice). Avoid assigning extracellular signal to the nearest cell without independent evidence (UniProt Q9UBD9 secretion).
How can epitope position affect CLCF1 interpretation?
CLCF1 has 2 annotated isoforms, a signal peptide at residues 1–27, and a mature chain spanning residues 28–225 (UniProt Q9UBD9). It also has an annotated glycosylation site at residue 29, close to the mature chain's amino terminus (UniProt Q9UBD9). Confirm the catalog antibody's stated immunogen or epitope before claiming that tissue staining measures both isoforms or the secreted mature protein (standard IHC practice). If epitope information is unavailable, report the result as antibody reactivity and compare its distribution with an independently validated reagent or orthogonal measurement (standard IHC practice). Keep retrieval and detection identical during that comparison (standard IHC practice).
How should I investigate CLCF1 by multiplex immunofluorescence?
Treat IF as a separate assay: the A08886 caption documents paraffin section IHC at 1:200, not an IF dilution or fixation method (A08886 tissue IHC caption). Pair CLCF1 with a validated marker of the cell population under investigation; lung macrophages or bone marrow hematopoietic cells are reported medium positive examples (HPA: tissue IHC). Select fluorophores in channels with low specimen autofluorescence, and inspect unstained tissue and single stain controls before interpreting overlap (standard IF practice). Choose permeabilisation after checking the antibody epitope: intracellular access may be needed, whereas extracellular staining should be assessed without assuming membrane permeabilisation is beneficial (standard IF practice; UniProt Q9UBD9 secretion).
What causes diffuse or misleading chromogenic CLCF1 signal?
First distinguish extracellular CLCF1 from background: secretion is annotated, and HPA reports plasma positivity with uncertain tissue IHC reliability (UniProt Q9UBD9; HPA: tissue profile and reliability). Run no primary and detection only controls, block endogenous peroxidase for peroxidase based detection, and compare deposits with tissue edges and damaged regions (standard IHC practice). If chromogen is widespread, reassess antibody concentration, blocking and wash conditions on adjacent sections while keeping retrieval constant (standard IHC practice). The A08886 image used 1:200, which is a documented starting point for that paraffin specimen rather than a universal optimum (A08886 tissue IHC caption; standard IHC practice).
How should I score CLCF1 across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring because secreted CLCF1 may be detected away from its source (UniProt Q9UBD9; HPA: tissue reliability uncertain). For cell associated chromogenic staining, record the percentage of positive cells and intensity, then calculate an H score using a consistent 0–3 intensity scale (standard IHC practice). For discrete positive cells, report density per mm² of viable tissue or per relevant cell population rather than total section area alone (standard IHC practice). Apply the same threshold, sampling plan and section processing across groups, and score extracellular deposits separately (standard IHC practice). Document any excluded folds, edges or necrotic areas (standard IHC practice).
When is a CLCF1 positive result convincing?
A convincing result shows reproducible signal in an anatomically identified cell population or extracellular compartment, with little corresponding signal in the no primary control (standard IHC practice; UniProt Q9UBD9 secretion). HPA reports medium staining in lung macrophages and bone marrow hematopoietic cells, but calls its tissue IHC reliability uncertain (HPA: tissue IHC and reliability). Treat isolated strong nuclear staining cautiously: nuclear bodies are reported in cell based imaging, yet that does not validate nuclear chromogenic staining in tissue (HPA: subcellular summary). Check edge effects, necrosis and endogenous peroxidase with matched controls before assigning positive cells (standard IHC practice). Interpret a negative brain region cautiously because reported neuronal staining is low or undetected by region (HPA: tissue IHC).
Boster reagents
Best CLCF1 / Cardiotrophin-like cytokine factor 1 IHC Antibodies
A08886 is listed for IHC and IF in human, mouse, and rat; its supplied image shows IHC in paraffin-embedded human brain (catalog applications and reactivity; IHC image caption).
Real IHC data
Anti-NNT-1 CLCF1 Antibody
Cat # A08886
A08886 will render with an IHC image of paraffin-embedded human brain at 1:200 (IHC image caption). IF is listed, but no IF image is supplied; human, mouse, and rat reactivity is listed (catalog applications and reactivity; IF image alts).
Which to pick: For tissue IHC, choose A08886 when working with paraffin sections; its image shows human brain, and the fixative is unreported (IHC image caption). For IF, A08886 is listed at 1:50, but ICC validation is unreported (catalog applications and IF dilution). A08886 lists human, mouse, and rat reactivity, though the supplied IHC image documents human tissue only (catalog reactivity; IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9UBD9 (CLCF1_HUMAN, Cardiotrophin-like cytokine factor 1).
- Human Protein Atlas. CLCF1 tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. CLCF1 subcellular location (ICC-IF): Localized to the nuclear bodies and vesicles..
- Human Protein Atlas. CLCF1 antibody validation summary (1 antibodies).
- CLCF1 is up-regulated in renal ischemia reperfusion injury and may associate with FOXO3. Annals of translational medicine 2022 — PMC9073803.
- CLCF1 Is a Novel Potential Immune-Related Target With Predictive Value for Prognosis and Immunotherapy Response in Glioma. Frontiers in immunology 2022 — PMC8898905.
- CLCF1 contributes to high-glucose-induced EndMT by modulating JAK2/STAT3 signaling. European journal of medical research 2025 — PMC12305993.
- Saikosaponin D and paeoniflorin improve the HCC immune microenvironment via CLCF1/PD-L1 mediated crosstalk between CAFs and tumor cells. Scientific reports 2026 — PMC13234363.
- PubMed PMID:10500198 — UniProt-cited evidence.
- PubMed PMID:10448081 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CLCF1 (Cardiotrophin-like cytokine factor 1). Expected localisation Cytoplasmic staining in several tissues (HPA tissue IHC). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A08886 at 1:50 recommended; image 1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Bone marrow (Hematopoietic cells); negative Caudate. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded human brain tissue; fixative not specified (datasheet A08886); 2. Fixation: Image fixative and duration unreported (datasheet A08886); 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-CLCF1, 1:50 recommended; image 1:200 (datasheet A08886); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep formalin fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Secretion can separate tissue RNA from protein staining (HPA tissue IHC); Isoform / epitope: 2 isoforms; signal peptide 1–27; epitope impact unknown (UniProt); Regulation: CRLF1/CNTFR-dependent secretion (UniProt). Evidence: UniProt Q9UBD9, HPA tissue IHC · Uncertain, HPA ICC-IF · nuclear bodies and vesicles., Datasheet A08886, PMC9073803, PMC12305993. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CLCF1/immunohistochemistry-ihc-guide.