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IFI44L / Interferon-induced protein 44-like · IHC design guide
Design Immunohistochemistry for IFI44L
Plan paraffin-section IFI44L IHC with the IHC-validated antibody at 2–5 μg/ml (datasheet A10975-1). Assess cytoplasmic staining alongside HPA-described high and undetected cell populations, bearing in mind that tissue-IHC reliability is uncertain (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
IFI44L 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 most tissues (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in glandular and myoepithelial cells (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A10975-1) | |
| Positive control | Breast+4 more · see all | |
| Negative control | Endometrium+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A10975-1) | |
| Caveat | Low consistency between IHC staining and RNA expression (HPA tissue IHC) | |
| Regulation | Type I interferon inducible (UniProt) | |
| Isoform / epitope | 2 isoforms; antibody epitope coverage is unknown (UniProt) |
Section 1
Recommended IFI44L IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet: A10975-1) and a published rat-heart IHC protocol (PMC8657967) provide starting conditions for IFI44L staining.
| Sample | Paraffin-embedded human non-small cell lung cancer tissue; fixative not specified (datasheet A10975-1) |
| Fixation | Image fixative and duration unreported (datasheet A10975-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A10975-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A10975-1) |
| Primary antibody | Rabbit anti-IFI44L, 2-5 μg/ml (datasheet A10975-1) |
| Primary incubation | Overnight at 4 °C (datasheet A10975-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A10975-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | IFI44L-positive staining in myoepithelial cells of breast (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval (datasheet: A10975-1); consider microwave 0.1 M citrate, pH 6.0 for rat heart (PMC8657967).
Section 2
What Is the Expected IFI44L Staining Pattern?
For paraffin-section IHC, expect mainly cytoplasmic IFI44L staining in the cell types where HPA reports protein expression, including breast myoepithelial cells and duodenal glandular cells (HPA: tissue IHC). UniProt lists IFI44L as cytoplasmic with no transmembrane segment (UniProt Q53G44). Interpret this pattern cautiously: HPA rates its tissue IHC profile Uncertain because antibody staining and RNA expression show low consistency (HPA: tissue IHC reliability).
| Cell-associated cytoplasmic chromogen is strong in breast myoepithelial cells or duodenal glandular cells. | This matches two HPA High observations and its broader cytoplasmic tissue profile (HPA: breast and duodenum IHC). Compare the labelled cells with tissue morphology; the HPA profile remains Uncertain and cannot establish that every positive cell represents specific IFI44L detection (HPA: tissue IHC reliability). |
| Staining is predominantly nuclear, membranous, or extracellular in a paraffin section. | Membranous or extracellular staining conflicts with UniProt's cytoplasmic location and lack of a transmembrane segment (UniProt Q53G44). Nuclear IHC warrants antibody and control review, but is not automatically artefact: HPA reports approved nucleoplasmic ICC-IF staining, while its tissue IHC profile is cytoplasmic and Uncertain (HPA: subcellular; HPA: tissue IHC). |
| Strong chromogen appears in cells that HPA lists as Not detected, such as lung alveolar cells. | Check whether the signal follows cell boundaries and survives detection controls; cross-reactivity or endogenous detection activity can mimic cellular staining (general IHC practice). HPA's Not detected finding applies to the specified cells in its sampled tissue, not to every cell in the organ or every biological state (HPA: lung IHC). |
| Colour spreads across stroma, lumens, section edges, or many unrelated cell types. | A diffuse field is less persuasive than cell-associated cytoplasmic staining in HPA High populations (HPA: tissue IHC). Excess background can arise from nonspecific antibody binding, endogenous enzyme activity, or detection chemistry; interpret it using appropriate reagent controls and the tissue's morphology (general IHC practice). |
| No signal appears in a reference section containing an HPA High cell population. | First confirm that the expected cells are present: HPA reports High staining in prostate glandular cells and smooth muscle cells, among others (HPA: tissue IHC). An absent result may reflect assay performance or specimen biology; HPA's Uncertain reliability prevents treating one reference section as a definitive IFI44L-positive control (HPA: tissue IHC reliability). |
💡Expected IFI44L appearanceA plausible positive IHC result is High, cell-associated cytoplasmic chromogen in an HPA High population such as breast myoepithelial cells (HPA: breast IHC); diffuse background or similarly strong signal in an HPA Not detected population raises concern for false positivity (HPA: tissue IHC; general IHC practice).
| Tissue-profile confidence | HPA calls its cytoplasmic tissue IHC profile Uncertain because staining and RNA expression have low consistency; treat cell-specific levels as reference observations that need independent confirmation (HPA: tissue IHC reliability). |
| Cell identity within a tissue | HPA reports High duodenal glandular-cell staining but Not detected lung alveolar-cell staining; assess the named cell population rather than assigning one expected intensity to an entire organ (HPA: duodenum and lung IHC). |
| Interferon-response biology | IFI44L is a type I interferon-stimulated gene with roles in infection responses (UniProt Q53G44). Biological context may inform sample selection, but these annotations do not predict staining intensity in a particular paraffin section. |
| Isoforms and antibody recognition | UniProt lists 2 isoforms (UniProt Q53G44). Their existence alone does not identify which form an antibody detects; use documented epitope information if available, without assuming isoform-specific staining. |
| IHC versus IF/ICC compartment | Does IF/ICC have the same expected compartment? HPA reports approved nucleoplasmic ICC-IF images in HeLa, PC-3, and U2OS, whereas its tissue IHC profile is cytoplasmic and Uncertain (HPA: subcellular; HPA: tissue IHC). Assess each application separately. |
| Situation | Likely cause | Next action |
|---|---|---|
| A proposed positive section is blank. | The section may lack the specific HPA High cell population, or the IHC workflow may have failed (HPA: tissue IHC; general IHC practice). | Verify cell identity and section quality; run an independently characterized positive control alongside the specimen, then review retrieval, primary-antibody conditions, and detection using the assay's validated settings (general IHC practice). |
| Chromogen is strong in an HPA Not detected population. | Nonspecific binding or endogenous detection activity is possible; HPA lists endometrial stromal cells as Not detected in its tissue IHC samples (HPA: endometrium IHC; general IHC practice). | Compare with an appropriate primary-antibody omission or detection control, check morphology, and reassess specificity with independent evidence before calling these cells IFI44L-positive (general IHC practice). |
| Most of the section shows diffuse background. | Nonspecific reagent binding, inadequate blocking, or endogenous enzyme activity can obscure cell-associated staining (general IHC practice). | Review blocking, washes, primary-antibody concentration, and detection controls within the validated IHC workflow; score only interpretable cell-associated signal (general IHC practice). |
| Nuclear staining dominates tissue IHC. | The result differs from HPA's cytoplasmic tissue profile, although approved nucleoplasmic ICC-IF staining makes a blanket artefact call unjustified (HPA: tissue IHC; HPA: subcellular). | Document the compartment and antibody used, inspect negative controls, and seek independent antibody or orthogonal confirmation before assigning nuclear IHC signal to IFI44L (general IHC practice). |
| Different reference tissues give inconsistent intensities. | HPA reports distinct cell-specific levels and rates its tissue IHC profile Uncertain (HPA: tissue IHC reliability). | Compare matching cell populations and processing conditions, record staining intensity by cell type, and avoid using HPA levels as universal acceptance thresholds (HPA: tissue IHC; general IHC practice). |
| IF/ICC and IHC suggest different compartments. | HPA reports approved nucleoplasmic ICC-IF localization but an Uncertain cytoplasmic tissue IHC profile (HPA: subcellular; HPA: tissue IHC). | Report each application and its validation status separately; use the dedicated IF/ICC guide for that application and seek independent confirmation before reconciling the locations (HPA: subcellular; HPA: tissue IHC). |
Experiment evidence
IHC & IF images showing IFI44L staining
A10975-1 · IHCIHC analysis of IFI44L using anti-IFI44L antibody (A10975-1). IFI44L was detected in a paraffin-embedded section of human non-small cell lun
A10975-1 · IHCIHC analysis of IFI44L using anti-IFI44L antibody (A10975-1). IFI44L was detected in a paraffin-embedded section of human stomach cancer tis
A10975-1 · IFIF analysis of IFI44L using anti-IFI44L antibody (A10975-1). IFI44L was detected in an immunocytochemical section of SiHa cells. Enzyme anti
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000137959-IFI44L/tissue/cerebral+cortex#imid_16500779
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000137959-IFI44L/tissue/colon#imid_16500121
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000137959-IFI44L/tissue/liver#imid_16500122
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000137959-IFI44L/tissue/kidney#imid_16500131
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA053247; green = IFI44L, blue = nucleus, red = microtubules.
HPA · ICC-IF HeLaHuman Protein Atlas immunofluorescence, HeLa, antibody HPA053247; green = IFI44L, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for IFI44L IHC & IF
🧪Run breast first and score myoepithelial cells for staining (HPA: High in breast myoepithelial cells). Use endometrium as the negative tissue, scoring stromal cells (HPA: Not detected in endometrial stromal cells); on the breast slide, assess non-myoepithelial cells for background without assuming they are target-negative.
Positive control tissue: Breast (Myoepithelial cells, HPA High)
Negative control tissue: Endometrium (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show IFI44L in HeLa, PC-3, U2OS, with annotated localisation: Nucleoplasm (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit IgG isotype control; the caption identifies a rabbit primary but does not state its clonality (selected-SKU IHC caption: rabbit anti-IFI44L). Confirm specificity with IFI44L-knockout material where available, and block endogenous peroxidase for the HRP–DAB workflow (selected-SKU IHC caption: peroxidase secondary and DAB).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the fixative is unreported in the selected-SKU paraffin-section caption. The caption uses heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required (selected-SKU IHC caption). The supplied evidence does not establish whether frozen sections or IF are easier or identify a breast-specific artefact; IF localisation also needs independent validation because the reported locations differ (UniProt Q53G44: cytoplasm; HPA subcellular: approved nucleoplasm).
HPA tissue IHC evidence for IFI44L
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — Low consistency between antibody staining and RNA expression data. 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 |
|---|---|---|---|---|
| Breast | Myoepithelial cells | High | Protein (IHC) | HPA → |
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Prostate | Glandular cells | High | Protein (IHC) | HPA → |
| Rectum | Glandular cells | High | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Endometrium | Cells in endometrial stroma | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Lung | Alveolar cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced IFI44L IHC Tips
Troubleshoot IFI44L staining in paraffin sections by checking retrieval, compartment, cell identity, controls, and the limits of the available tissue evidence.
Which retrieval condition should I start with for weak IFI44L staining?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet A10975-1). The catalog antibody detected IFI44L in a paraffin-embedded human non-small cell lung cancer section under that condition, followed by 2 μg/ml primary antibody overnight at 4°C (datasheet A10975-1). If staining remains weak, compare a modest range of heating durations on adjacent sections while keeping dilution and detection constant; use a different buffer only as a documented fallback after that comparison (standard IHC practice). Include a section processed without primary antibody and assess morphology alongside signal, since excessive retrieval can make apparent staining harder to interpret (standard IHC practice).
Could fixation explain weak or uneven IFI44L staining in my sections?
Target-specific sensitivity of IFI44L staining to fixation is unknown from the supplied evidence; the catalog image identifies a paraffin-embedded section but does not state its fixative (datasheet A10975-1). Record the fixative, fixation duration, tissue thickness, and processing history before comparing sections, because these variables can change antigen accessibility and morphology in IHC (standard IHC practice). Compare similarly processed sections in the same staining run and keep retrieval, antibody concentration, and DAB development consistent (standard IHC practice). If signal differs, test processing variables systematically and confirm that any gain preserves the expected cellular pattern rather than treating the difference as a demonstrated IFI44L-specific fixation effect (UniProt Q53G44; standard IHC practice).
How should I interpret cytoplasmic and nuclear IFI44L staining?
Assess cytoplasmic staining first: IFI44L is annotated as cytoplasmic, with no transmembrane segment or signal peptide (UniProt Q53G44). Tissue IHC describes cytoplasmic expression in most tissues, although its staining reliability is uncertain (HPA tissue IHC: profile and reliability). Nuclear signal needs separate scrutiny because cell imaging reports an approved nucleoplasmic location, creating a method-dependent localisation discrepancy to resolve rather than ignore (HPA subcellular: Nucleoplasm). Compare compartment boundaries against the counterstain, inspect adjacent cells and section edges, and require a consistent pattern across replicates before assigning nuclear staining to IFI44L (standard IHC practice). Report cytoplasmic and nuclear scores separately if both persist (standard IHC practice).
Can epitope choice account for different IFI44L staining patterns?
IFI44L has 2 annotated isoforms and a TLDc domain spanning residues 1–159 (UniProt Q53G44). The supplied catalog caption does not identify the antibody epitope, so staining cannot establish which isoform contributes to a cell's signal (datasheet A10975-1; UniProt Q53G44). Obtain the immunogen or epitope location before comparing antibodies, then check whether that sequence is shared by the isoforms under study (standard IHC practice). IFI44L has no annotated glycosylation sites or modified residues in this record, but those annotations alone do not establish epitope accessibility after tissue processing (UniProt Q53G44). Keep epitope and retrieval differences in view when staining patterns disagree (standard IHC practice).
How can I check an IFI44L IHC pattern with multiplex immunofluorescence?
Use multiplex IF as a separate check of the IHC interpretation, pairing IFI44L with a validated marker for the cell population being assessed; glandular cells are among the reported tissue IHC positive populations (HPA tissue IHC: glandular cells). Select spectrally separated fluorophores after inspecting unstained tissue autofluorescence, particularly before treating a weak channel as positive (standard IF practice). Because IFI44L lacks a transmembrane segment and is annotated in the cytoplasm, plan permeabilisation for access to an intracellular epitope, then check whether the resulting compartment pattern is credible (UniProt Q53G44; standard IF practice). Evaluate any nuclear signal separately because cell imaging reports nucleoplasmic localisation (HPA subcellular: Nucleoplasm). Match cell identities between IF and chromogenic sections before comparing their scores (standard IHC/IF practice).
What should I check when DAB staining appears diffuse or widespread?
The catalog tissue example used 10% goat serum block, 2 μg/ml primary antibody overnight at 4°C, a peroxidase-linked secondary, and DAB detection (datasheet A10975-1). Check a no-primary control and include a general endogenous peroxidase block before interpreting diffuse brown signal (standard chromogenic IHC practice). If background persists, titrate the primary antibody and DAB development while holding retrieval and imaging conditions constant, and inspect whether signal tracks tissue edges or damaged regions (standard IHC practice). Judge the result against cellular localisation rather than overall darkness: tissue IHC describes predominantly cytoplasmic IFI44L, but its antibody staining has uncertain consistency with RNA data (HPA tissue IHC: profile and reliability).
How should I quantify IFI44L staining across tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region before scoring, then record the percentage of positive cells and staining intensity to calculate an H-score where that scale suits the study (standard IHC practice). Report cytoplasmic staining separately from nuclear staining because tissue IHC describes a cytoplasmic pattern while cell imaging reports nucleoplasmic localisation (HPA tissue IHC: profile; HPA subcellular: Nucleoplasm). If counting positive cells, express density per mm² of viable, assessable tissue and normalise comparisons to the same cell population, sampled area, and staining batch (standard IHC practice). Apply fixed thresholds and blinded scoring where feasible, and retain no-primary control results alongside scores (standard IHC practice).
When is an apparent IFI44L-positive cell convincing rather than artefactual?
A convincing call combines cell-resolved staining, reproducibility across comparable sections, and appropriate controls with a plausible compartment pattern (standard IHC practice). Cytoplasmic staining fits the tissue IHC profile and UniProt annotation, while nucleoplasmic staining has separate cell-imaging support and merits explicit reporting (HPA tissue IHC: profile; UniProt Q53G44; HPA subcellular: Nucleoplasm). Check the cell type rather than labeling an entire organ positive: HPA reports high staining in several glandular-cell populations but no detection in lung alveolar cells (HPA tissue IHC: positive and negative cell populations). Discount isolated staining at cut edges, necrotic areas, or sites that remain brown without primary antibody, including possible endogenous enzyme signal (standard IHC practice). HPA rates the tissue staining reliability uncertain, so corroborate consequential conclusions independently (HPA tissue IHC: reliability).
Boster reagents
Best IFI44L / Interferon-induced protein 44-like IHC Antibodies
A10975-1 has IHC images from human paraffin-embedded lung and stomach cancer sections and an IF image from SiHa cells (catalog image captions). Reactivity is listed for human, monkey, mouse and rat (catalog).
Real IHC data
Anti-IFI44L Antibody ®
Cat # A10975-1
A10975-1 will render with its own IHC figure from a human paraffin-embedded non-small cell lung cancer section (IHC image caption). Its additional IHC caption shows human stomach cancer tissue, and its IF caption shows SiHa cells (catalog image captions).
Which to pick: Choose A10975-1 for paraffin-section IHC: its figure used heat retrieval in EDTA at pH 8.0 and 2 μg/ml primary antibody; the fixative is unreported (IHC image caption). Choose the same SKU for IF/ICC because its IF caption shows SiHa cells at 5 μg/ml (IF image caption). For other species, its catalog lists monkey, mouse and rat reactivity, while the supplied IHC and IF images show human material only (catalog reactivity; image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q53G44 (IF44L_HUMAN, Interferon-induced protein 44-like).
- Human Protein Atlas. IFI44L tissue IHC expression (reliability: Uncertain).
- Human Protein Atlas. IFI44L subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. IFI44L antibody validation summary (2 antibodies).
- IFI44L is a novel tumor suppressor in human hepatocellular carcinoma affecting cancer stemness, metastasis, and drug resistance via regulating met/Src signaling pathway. BMC cancer 2018 — PMC5977745.
- Identified Three Interferon Induced Proteins as Novel Biomarkers of Human Ischemic Cardiomyopathy. International journal of molecular sciences 2021 — PMC8657967.
- Identification of NET formation and the renoprotective effect of degraded NETs in lupus nephritis. American journal of physiology. Renal physiology 2024 — PMC11483074.
- Comprehensive Analysis of Immune Implication and Prognostic Value of IFI44L in Non-Small Cell Lung Cancer. Frontiers in oncology 2021 — PMC8761744.
- PubMed PMID:17974005 — UniProt-cited evidence.
- PubMed PMID:16710414 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for IFI44L (Interferon-induced protein 44-like). Expected localisation Cytoplasmic staining in most tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A10975-1). Validated IHC antibody: A10975-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Breast (Myoepithelial cells); negative Endometrium. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human non-small cell lung cancer tissue; fixative not specified (datasheet A10975-1); 2. Fixation: Image fixative and duration unreported (datasheet A10975-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: EDTA pH 8.0 (datasheet A10975-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A10975-1); 8. Primary antibody: Rabbit anti-IFI44L, 2-5 μg/ml (datasheet A10975-1); 9. Primary incubation: Overnight at 4 °C (datasheet A10975-1). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A10975-1); Caveat: Low consistency between IHC staining and RNA expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; antibody epitope coverage is unknown (UniProt); Regulation: Type I interferon inducible (UniProt). Evidence: UniProt Q53G44, HPA tissue IHC · Uncertain, HPA ICC-IF · nucleoplasm., Datasheet A10975-1, PMC8657967. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/IFI44L/immunohistochemistry-ihc-guide.