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CSTF2 / Cleavage stimulation factor subunit 2 · IHC design guide
Design Immunohistochemistry for CSTF2
Plan CSTF2 paraffin IHC around its general nuclear staining pattern (HPA tissue IHC). Use high-staining glandular or respiratory epithelial cells as positive references, while accounting for medium concordance between staining and RNA expression (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CSTF2 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 | Nuclear staining (HPA tissue IHC) | |
| Staining pattern | General nuclear staining across tissues (HPA tissue IHC) | |
| Antigen retrieval | Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen) | |
| Positive control | Adrenal gland+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 samples (standard IHC practice; not target-specific) | |
| Caveat | Staining and RNA expression have medium concordance (HPA tissue IHC) | |
| Regulation | Low tissue specificity of RNA expression (HPA tissue RNA) | |
| Isoform / epitope | 2 isoforms; epitope effects unspecified (UniProt) |
Section 1
Recommended CSTF2 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is provided separately; the published CSTF2 IHC options below cover oral, gastric, and liver cancer specimens (PMC11842344; PMC12751398; PMC9304882).
| Sample | Paraffin-embedded human colon carcinoma tissue; fixative not specified (datasheet A07037) |
| Fixation | Image fixative and duration unreported (datasheet A07037); 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 Tris-EDTA buffer, pH 9.0, 20 min at 95–98 °C (standard rule: nuclear 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-CSTF2, 1:50-1:200 (datasheet A07037) |
| 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 | CSTF2-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General nuclear expression. No signal in the no-primary control. |
💡Decision noteFor the catalog antibody, start with pH 9.0 Tris-EDTA retrieval at 95–98 °C for 20 min (page retrieval setting); CSTF2 is nuclear (UniProt P33240).
Section 2
What Is the Expected CSTF2 Staining Pattern?
CSTF2 is a nuclear protein with no transmembrane segment (UniProt P33240: localization and topology). In paraffin-section IHC, expect nuclear staining across multiple cell types, with high staining reported in selected glandular and respiratory epithelial cells (HPA: general nuclear expression; High in listed cells). HPA rates tissue IHC reliability as Supported, with medium consistency between antibody staining and RNA expression (HPA: tissue IHC reliability).
| Nuclear chromogen in glandular cells of appendix or rectum. | This matches the expected compartment and cell type; HPA reports High staining in these cells (HPA: appendix and rectum). Judge the nuclear pattern alongside tissue structure and the matched control, since HPA's Supported rating indicates medium consistency with RNA expression (HPA: tissue IHC reliability). |
| Predominantly cytoplasmic or membrane-like staining with little nuclear signal. | This conflicts with UniProt's nuclear localization and HPA's general nuclear IHC profile (UniProt P33240: localization; HPA: tissue IHC profile). Treat the compartment mismatch as a possible artefact; check whether background, detection, or section quality obscures nuclei before assigning CSTF2 positivity (general IHC practice). |
| Strong staining confined to an unexpected cell population. | Compare it with HPA's cell-specific observations before interpreting it as CSTF2. For example, HPA reports High staining in bronchial respiratory epithelial cells (HPA: bronchus). An isolated discordant pattern raises possible cross-reactivity or endogenous detection activity; morphology and controls are needed to distinguish them (general IHC practice). |
| Diffuse chromogen across tissue, including spaces outside cells. | A field-wide deposit lacks the nuclear distribution expected for CSTF2 (UniProt P33240: localization; HPA: general nuclear expression). Inspect the no-primary control and reagent background, then assess blocking, washes, and chromogen development (general IHC practice). Diffuse colour alone should not be scored as nuclear target staining. |
| No nuclear signal in a well-preserved expected-positive section. | Absence is unexpected in cells reported High by HPA, such as adrenal glandular cells (HPA: adrenal gland). Check section integrity and the IHC workflow with a positive control (general IHC practice). HPA provides no negative tissue in this payload, so a low-staining tissue is not a validated negative control (HPA: tissue IHC). |
💡Expected CSTF2 appearanceCall a result positive when identifiable nuclei show the expected chromogenic signal in HPA-listed cells, including High-staining glandular or respiratory epithelial populations; isolated cytoplasmic colour or diffuse tissue deposit is suspicious (HPA: tissue IHC profile; UniProt P33240: nuclear localization).
| Cell and tissue context | HPA reports High staining in adrenal, appendix, gallbladder, and rectal glandular cells, and bronchial and nasopharyngeal respiratory epithelial cells (HPA: tissue IHC). These are useful expected-positive comparisons; HPA also reports low tissue RNA specificity (HPA: RNA specificity). |
| Lower-staining comparisons | Cholangiocytes, cardiomyocytes, peripheral nerve, and bone-marrow hematopoietic cells are listed as Low, not negative (HPA: tissue IHC). Use them as relative staining comparisons, without treating absence of colour as a required biological result. |
| Antibody evidence | Both listed antibodies, HPA000427 and CAB004680, have Supported IHC status (HPA: antibody validation). This supports comparison with the reported pattern but does not establish Enhanced IHC validation for either antibody (HPA: antibody validation). |
| Target structure | CSTF2 has no signal peptide, propeptide, or transmembrane segment; its annotated chain spans residues 1–577 (UniProt P33240: processing and topology). A membrane-outline pattern therefore lacks support from the target annotation. The payload gives no antibody epitope location (UniProt P33240; HPA: supplied antibody data). |
| Isoforms and modifications | Two isoforms and several modified residues are annotated, including phosphorylation and arginine methylation (UniProt P33240: isoforms and modified residues). Their effects on the listed antibodies' IHC signal are unreported here; do not infer variant-specific staining or retrieval needs from these annotations. |
| IF/ICC: what pattern is expected? | HPA reports mainly nucleoplasmic signal with additional nuclear-body signal in ICC-IF; HPA000427 has Enhanced ICC validation (HPA: subcellular localization; antibody validation). This is an IF interpretation reference, not an IHC-P protocol or evidence that every paraffin-section nucleus must show resolvable nuclear bodies. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected-positive nuclei are blank. | The IHC run, section, or antibody conditions may have failed; HPA reports High staining in appendix glandular cells (HPA: appendix). | Verify a known-positive section, detection reagents, and the catalog antibody's IHC-P instructions; review retrieval conditions as general IHC practice. No CSTF2-specific fixation sensitivity is supplied. |
| Most colour sits in cytoplasm or along cell borders. | The location disagrees with nuclear CSTF2 (UniProt P33240: localization; HPA: general nuclear expression); background or cross-reactivity is possible. | Inspect morphology and a no-primary control, then check blocking, washes, and detection settings (general IHC practice). Score only convincing nuclear signal. |
| High colour appears in an unlisted cell type. | HPA's reported cell-level pattern does not validate that population; cross-reactivity or endogenous detection activity may contribute (HPA: tissue IHC; general IHC practice). | Compare adjacent expected-positive cells and a no-primary control; confirm the unexpected pattern with an independently validated approach before calling it CSTF2. |
| Brown haze covers cells and surrounding tissue. | Nonlocalised deposit cannot be reconciled with the reported nuclear pattern (HPA: tissue IHC; UniProt P33240: localization). | Review no-primary staining, blocking, washes, and chromogen development (general IHC practice); reassess only after nuclei can be distinguished from background. |
| A low-staining comparison looks blank. | HPA labels cholangiocytes and cardiomyocytes Low, while listing no negative tissues in this payload (HPA: tissue IHC). | Keep a High-staining tissue on the same run to verify detection. Describe the blank comparison as an observation, not proof of a CSTF2-negative tissue. |
| Nuclear-body puncta are not resolved in IHC. | HPA's additional nuclear-body localization comes from ICC-IF images, whereas tissue IHC is summarized as general nuclear expression (HPA: subcellular localization; tissue IHC). | Evaluate the paraffin section against the supported nuclear IHC pattern. Use the separate IF/ICC guide when nuclear-body resolution is the experimental objective. |
Experiment evidence
IHC & IF images showing CSTF2 staining
A07037 · IHCImmunohistochemistry (IHC) analyzes of CstF-64 (R46) pAb in paraffin-embedded human colon carcinoma tissue at 1:50.showing nucleus staining.
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000101811-CSTF2/tissue/cerebral+cortex#imid_1349084
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000101811-CSTF2/tissue/colon#imid_1349010
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000101811-CSTF2/tissue/liver#imid_1349011
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000101811-CSTF2/tissue/kidney#imid_1349004
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000427; green = CSTF2, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA000427; green = CSTF2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CSTF2 IHC & IF
🧪Run appendix first and expect nuclear staining in glandular cells (HPA: High in appendix glandular cells; UniProt P33240: nucleus). HPA detects CSTF2 in all 45 scored tissues, so there is no HPA-supported negative tissue; use no-primary and isotype controls, and do not designate any cell type within the positive slide as an internal negative (HPA: no negative tissue rows).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: None in HPA: CSTF2 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 CSTF2 in A-431, U-251MG, U2OS, U2OS, siRNA 1 (10x), U2OS, siRNA 1 (40x), U2OS, siRNA 2 (10x), U2OS, siRNA 2 (40x), U2OS, scrambled (10x), U2OS, scrambled (40x), with annotated localisation: Nucleoplasm (enhanced) (HPA subcellular).
Technical controls: Include a secondary-only, no-primary section and a control matched to the primary antibody’s host species and clonality, such as nonimmune IgG for a polyclonal antibody (standard IHC practice). Use CSTF2 knockout material or a peptide block, where available, to test target specificity, and quench endogenous peroxidase before chromogenic detection because appendix leukocytes can contribute peroxidase signal (standard IHC practice).
⚠️Feasibility: No supplied target or application evidence reports a CSTF2-specific fixation window or retrieval dependency; optimize antigen retrieval empirically (supplied evidence). The selected A07037 paraffin-section caption does not state a fixative, so its fixation conditions are unreported (A07037 caption: fixative not stated). The evidence does not establish whether frozen sections or IF are easier; for IF, interpret nucleoplasmic and nuclear-body signal against appropriate background controls, particularly where appendix tissue autofluorescence is present (HPA subcellular: nucleoplasm and nuclear bodies; standard IF practice).
HPA tissue IHC evidence for CSTF2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — 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 → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| None in HPA: CSTF2 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead. | ||||
Section 3
Advanced CSTF2 IHC Tips
Troubleshoot CSTF2 chromogenic IHC by checking nuclear staining, section processing, and controls before interpreting differences between samples (UniProt P33240; standard IHC practice).
Which retrieval conditions should I start with for weak CSTF2 nuclear staining?
Start with Tris-EDTA pH 9.0 heat-induced antigen retrieval at 95–98 °C for 20 min (page retrieval setting). Compare sections processed in the same batch, allowing them to cool consistently before antibody incubation so retrieval differences do not masquerade as expression differences (standard IHC practice). Expect predominantly nuclear staining, because CSTF2 is annotated as nuclear and the tissue profile reports general nuclear expression (UniProt P33240; HPA tissue IHC). If staining remains weak, compare a longer retrieval condition on adjacent sections while monitoring tissue integrity, and retain the condition that preserves morphology and nuclear contrast (standard IHC practice).
Can I attribute weak CSTF2 staining to fixation?
The A07037 image shows nuclear staining in paraffin-embedded human colon carcinoma at 1:50, but its caption does not identify the fixative (caption A07037). Target-specific fixation sensitivity is unknown; neither the HPA nuclear pattern nor UniProt topology or modifications establishes how CSTF2 responds to fixation (HPA tissue IHC; UniProt P33240). For a new series, record fixative, fixation duration, processing conditions, and section age, then compare matched sections using identical retrieval and chromogenic detection (standard IHC practice). If signal varies, inspect morphology and a concurrently processed control before changing antibody dilution, since processing differences can change epitope accessibility (standard IHC practice).
What CSTF2 staining pattern should I accept in a paraffin section?
Score convincing CSTF2 signal primarily within nuclei: UniProt places CSTF2 in the nucleus, and HPA reports general nuclear tissue staining (UniProt P33240; HPA tissue IHC). In cell imaging, HPA further places it mainly in the nucleoplasm with additional nuclear-body signal, while UniProt notes colocalisation with DDX1 in cleavage bodies (HPA subcellular; UniProt P33240). Use a nuclear counterstain and inspect individual cells at high magnification to distinguish nuclear chromogen from overlying pigment or diffuse cytoplasmic deposit (standard IHC practice). Treat isolated cytoplasmic staining as questionable until it survives antibody omission, detection controls, and comparison with a well-preserved positive section (standard IHC practice).
How should isoforms and epitope uncertainty affect CSTF2 IHC interpretation?
CSTF2 has 2 annotated isoforms and an RNA-recognition motif at residues 16–94, but the supplied A07037 caption does not map the antibody epitope (UniProt P33240; caption A07037). Therefore, a nuclear stain cannot by itself establish which isoform contributes to the signal or whether retrieval exposes both equally (UniProt P33240; standard IHC practice). UniProt also lists modified residues, including phosphoserine at 14 and methylarginines at 308, 468, and 475; their effect on this antibody is unreported (UniProt P33240). If isoform-specific interpretation matters, obtain epitope documentation and validate against an independent, isoform-discriminating assay before assigning staining to either isoform (standard IHC practice).
How can I check a nuclear CSTF2 pattern by multiplex IF?
For a separate IF assay, pair CSTF2 with a validated glandular-cell marker when examining rectum, where HPA reports high glandular-cell staining (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and place the CSTF2 readout in a far-red channel if tissue autofluorescence compromises shorter wavelengths (standard IF practice). CSTF2 is nuclear and lacks a transmembrane segment, so use a permeabilisation condition that permits antibody access to nuclear epitopes, then verify that nuclear morphology remains intact (UniProt P33240; standard IF practice). Compare antibody-omission controls and single-channel images before calling nucleoplasmic or nuclear-body colocalisation, patterns reported by HPA cell imaging (HPA subcellular; standard IF practice).
How do I reduce diffuse or misleading CSTF2 chromogenic background?
First inspect the no-primary control: the A07037 caption describes PBS replacing primary antibody, followed by biotinylated secondary antibody and avidin-peroxidase detection (caption A07037). If that control develops color, check secondary-antibody binding, endogenous biotin where relevant, endogenous peroxidase, and incomplete washes before changing the CSTF2 primary incubation (standard IHC practice). Use an appropriate protein block and peroxidase block, keep chromogen development comparable between sections, and assess whether deposit is confined to nuclei (standard IHC practice; UniProt P33240). Compare suspect areas with intact tissue away from section edges, because pooled reagent or damaged morphology can create misleading local signal (standard IHC practice).
What is a defensible way to quantify CSTF2 IHC across samples? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scoring compartment as the nucleus, consistent with UniProt localisation and HPA general nuclear tissue staining (UniProt P33240; HPA tissue IHC). For each prespecified tissue region, report the percentage of positive nuclei and an intensity-weighted H-score, documenting the intensity categories and threshold used (standard IHC practice). Normalise counts to the number of evaluable nuclei or, for spatial counts, report positive nuclei per mm² of viable tissue rather than per whole image (standard IHC practice). Apply identical retrieval, imaging, and threshold settings across comparison groups, and exclude necrotic or folded areas before comparing scores (standard IHC practice).
When is apparent CSTF2 positivity more likely to be artefact?
Favor reproducible nuclear staining over isolated cytoplasmic or extracellular color, since CSTF2 is nuclear and HPA reports general nuclear expression (UniProt P33240; HPA tissue IHC). A07037 provides a paraffin-section example of nuclear staining in human colon carcinoma, with a PBS-for-primary negative control described in its caption (caption A07037). Question signal confined to tissue edges, necrotic areas, or cells with poor nuclear preservation, and investigate color that also appears in the no-primary control (standard IHC practice). Endogenous peroxidase activity and nonspecific detection can mimic positivity; assess blocking and control sections before treating such deposits as biological CSTF2 expression (standard IHC practice).
Boster reagents
Best CSTF2 / Cleavage stimulation factor subunit 2 IHC Antibodies
The catalog antibody has a real IHC image from a paraffin-embedded human colon carcinoma section (IHC image caption). IF is listed, with Human, Mouse and Rat reactivity (catalog: A07037).
Real IHC data
Anti-CstF-64 (R46) CSTF2 Antibody
Cat # A07037
A07037 has an IHC image showing nuclear staining in a paraffin-embedded human colon carcinoma section at 1:50, with a PBS-for-primary control (IHC image caption). A07037 lists IF and IHC applications and Human, Mouse and Rat reactivity; no IF image is supplied (catalog: A07037).
Which to pick: For tissue IHC, choose A07037: its own image documents staining in a paraffin-embedded section, but does not report the fixative (IHC image caption). For IF/ICC, A07037 lists IF and a 1:50–1:200 dilution range, although no IF image is supplied (catalog: A07037). For cross-species work, A07037 lists Human, Mouse and Rat reactivity and is rabbit polyclonal; the supplied IHC image documents human tissue only (catalog: A07037; IHC image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P33240 (CSTF2_HUMAN, Cleavage stimulation factor subunit 2).
- Human Protein Atlas. CSTF2 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. CSTF2 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the nuclear bodies..
- Human Protein Atlas. CSTF2 antibody validation summary (2 antibodies).
- Expression of CSTF2 in oral squamous cell carcinoma and its relationship with immune infiltration and poor prognosis. Frontiers in oral health 2025 — PMC11842344.
- The role of CSTF2 in gastric cancer: implications for therapy. European journal of medical research 2025 — PMC12751398.
- CSTF2 Promotes Hepatocarcinogenesis and Hepatocellular Carcinoma Progression via Aerobic Glycolysis. Frontiers in oncology 2022 — PMC9304882.
- CSTF2 Acts as a Prognostic Marker Correlated with Immune Infiltration in Hepatocellular Carcinoma. Cancer management and research 2022 — PMC9481280.
- PubMed PMID:1741396 — UniProt-cited evidence.
- PubMed PMID:15772651 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CSTF2 (Cleavage stimulation factor subunit 2). Expected localisation Nuclear staining (HPA tissue IHC). Antigen retrieval: Tris-EDTA pH 9.0 HIER, 95–98 °C, 20 min (rule: nuclear antigen). Validated IHC antibody: A07037 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); 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 colon carcinoma tissue; fixative not specified (datasheet A07037); 2. Fixation: Image fixative and duration unreported (datasheet A07037); 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 Tris-EDTA buffer, pH 9.0 , 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-CSTF2, 1:50-1:200 (datasheet A07037); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across samples (standard IHC practice; not target-specific); Caveat: Staining and RNA expression have medium concordance (HPA tissue IHC); Isoform / epitope: 2 isoforms; epitope effects unspecified (UniProt); Regulation: Low tissue specificity of RNA expression (HPA tissue RNA). Evidence: UniProt P33240, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A07037, PMC11842344, PMC12751398. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CSTF2/immunohistochemistry-ihc-guide.