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FLNB / Filamin-B · IHC design guide
Design Immunohistochemistry for FLNB
Plan FLNB paraffin IHC using the catalog antibody validated for IHC (datasheet A02562-1). Compare cytoplasmic and membranous staining across cell types, with endothelial cells expected to stain strongly (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
FLNB Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Cytoplasmic and membranous tissue staining (HPA tissue IHC) | |
| Staining pattern | Most cells show cytoplasmic/membranous staining; endothelial cells are abundant (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02562-1) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Caudate+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Cardiomyocytes may lack detectable staining (HPA tissue IHC) | |
| Regulation | Staining varies by cell type (HPA tissue IHC) | |
| Isoform / epitope | Nine isoforms; check whether the epitope spans splice variants (UniProt) |
Section 1
Recommended FLNB IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A02562-1) is paired with a published FLNB IHC-P protocol for human PDAC resections (PMC7611572).
| Sample | Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A02562-1) |
| Fixation | Image fixative and duration unreported (datasheet A02562-1); 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 A02562-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02562-1) |
| Primary antibody | Rabbit anti-FLNB, 1-2 μg/ml (datasheet A02562-1) |
| Primary incubation | Overnight at 4 °C (datasheet A02562-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A02562-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | FLNB-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and membranous expression in most cells. Highly abundant in endothelial cells. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet A02562-1); the published PDAC excerpt does not specify a retrieval method (PMC7611572).
Section 2
What Is the Expected FLNB Staining Pattern?
FLNB staining should be mainly cytoplasmic and membranous, with particularly abundant signal in endothelial cells and high signal in several glandular or epithelial cell populations (HPA tissue IHC: Enhanced reliability). A cell cortex or actin-associated pattern is consistent with its annotated location and role; FLNB has no transmembrane segment, so membrane-associated staining does not imply that it spans the membrane (UniProt O75369: location, function and topology).
| Cytoplasmic and membranous signal is clear in endothelium and in bronchial respiratory epithelium or colon glandular cells (HPA tissue IHC). | This matches the reported pattern: endothelial cells are highly abundant, and the listed epithelial cells stain at a high level (HPA tissue IHC). Judge signal within identified cells and alongside a negative reagent control (general IHC practice). |
| Predominantly nuclear signal appears without a convincing cytoplasmic or membranous pattern. | Nuclear-only staining conflicts with the reported IHC pattern and annotated cell cortex and cytoskeletal locations (HPA tissue IHC; UniProt O75369: location). Treat it as a possible artefact; check controls and staining conditions before assigning it to FLNB (general IHC practice). |
| Strong signal appears in cardiomyocytes or lymph-node germinal-center cells, while nearby expected cells stain weakly (HPA tissue IHC). | HPA reports FLNB as not detected in those specific cell populations (HPA tissue IHC). Consider cross-reactivity or endogenous chromogen activity, and compare cellular shape and reagent controls (general IHC practice). An unexpected result alone does not prove either cause. |
| Color forms a diffuse haze across tissue and empty slide areas, obscuring cell boundaries. | A field-wide deposit does not resemble HPA's cellular cytoplasmic and membranous pattern (HPA tissue IHC). Check blocking, washing and detection controls for background (general IHC practice); do not score the haze as FLNB-positive cells. |
| No signal is visible in endothelium or in high-staining glandular cells on an otherwise interpretable section (HPA tissue IHC). | The missing expected reference pattern makes a negative call uncertain (HPA tissue IHC). First check tissue preservation, retrieval, antibody and detection steps with appropriate controls (general IHC practice). HPA levels describe observed populations, not a guarantee that every section will stain. |
💡Expected FLNB appearanceCall a positive result when identifiable endothelium shows abundant cytoplasmic and membranous staining, with high staining in an HPA-listed glandular or epithelial population (HPA tissue IHC); isolated nuclear signal or diffuse noncellular color is suspect (HPA tissue IHC; general IHC practice).
| Cellular location | FLNB is annotated at the cell cortex, cytoskeleton and stress fibers (UniProt O75369: location). HPA describes cytoplasmic and membranous tissue staining (HPA tissue IHC); interpret the membrane edge as an associated pattern, consistent with the absence of a transmembrane segment (UniProt O75369: topology). |
| Cell-population contrast | HPA reports high staining in bronchial respiratory epithelium and several glandular populations, but no detected signal in cardiomyocytes or lymph-node germinal-center cells (HPA tissue IHC). Use the named cell populations when comparing areas of a section; whole-organ labels can hide these differences. |
| Isoforms and tissue context | UniProt lists nine FLNB isoforms and reports heart expression for several, whereas HPA reports cardiomyocytes as not detected by tissue IHC (UniProt O75369: isoforms and tissue specificity; HPA tissue IHC). Expression annotation therefore does not establish a positive cardiomyocyte IHC result; the supplied sources do not map staining to individual isoforms. |
| Evidence strength | HPA rates the tissue profile Enhanced: paired antibodies show similar staining, while agreement with RNA expression is medium (HPA tissue IHC: reliability). HPA004747 and HPA004886 are IHC Enhanced; CAB019322 is IHC Supported (HPA antibodies). These ratings support the reported profile without validating every unexpected stained cell. |
| IF/ICC Q&A: What pattern should appear? | Mainly plasma membrane, with additional Golgi, cytosol and actin-filament localization (HPA subcellular ICC-IF). That image-based pattern complements the tissue IHC interpretation; the supplied ICC-IF record does not establish a paraffin-section IF pattern (HPA subcellular ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| Endothelium and an HPA high-staining epithelial population both appear blank (HPA tissue IHC). | A failed staining run or weak detection is possible (general IHC practice); the image alone cannot identify the failed step. | Check a known-positive section and reagent controls, then review retrieval, primary-antibody and detection steps (general IHC practice). Do not infer FLNB-specific fixation sensitivity from these sources. |
| Only a few cells stain weakly in an expected positive area (HPA tissue IHC). | Section quality, cellular composition or insufficient signal may explain the weak result (general IHC practice); HPA tissue levels are population descriptions. | Identify endothelial or glandular cells morphologically, compare a positive control and repeat with a controlled change to the general IHC workflow if needed (general IHC practice). |
| Nuclei dominate the staining while cell edges remain indistinct. | The compartment disagrees with the HPA tissue profile and UniProt localization (HPA tissue IHC; UniProt O75369: location); nonspecific detection is possible. | Inspect reagent controls and compare with the expected cytoplasmic and membranous signal in identifiable positive cells (HPA tissue IHC; general IHC practice). |
| A uniform brown haze masks tissue structures. | Residual detection reagent or inadequate washing can cause chromogenic background (general IHC practice). | Review blocking, washes and detection timing; compare the negative reagent control before scoring cells (general IHC practice). |
| Brown deposits track blood-rich areas or pigment instead of cell outlines. | Endogenous enzyme activity or pigment can mimic chromogenic signal (general IHC practice). | Compare a no-primary control and tissue morphology; review the relevant endogenous-activity block for the detection chemistry (general IHC practice). |
| Cardiomyocytes stain strongly despite HPA reporting them as not detected (HPA tissue IHC). | Cross-reactivity or background is possible, although an unexpected result alone does not establish its cause (general IHC practice). | Confirm the cell identity and controls, then compare an independently validated FLNB antibody if available (general IHC practice; HPA antibodies). Do not use UniProt heart expression alone to validate this IHC pattern (UniProt O75369: tissue specificity; HPA tissue IHC). |
Experiment evidence
IHC & IF images showing FLNB staining
A02562-1 · IHCIHC analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (A02562-1). Filamin B/FLNB was detected in a paraffin-embedded section of
A02562-1 · IHCIHC analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (A02562-1). Filamin B/FLNB was detected in a paraffin-embedded section of
A02562-1 · IFIF analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (A02562-1). Filamin B/FLNB was detected in an immunocytochemical section of
M02562-1 · IHCIHC analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (M02562-1). Filamin B/FLNB was detected in a paraffin-embedded section of
M02562-1 · IHCIHC analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (M02562-1). Filamin B/FLNB was detected in a paraffin-embedded section of
M02562-1 · IFIF analysis of Filamin B/FLNB using anti-Filamin B/FLNB antibody (M02562-1). Filamin B/FLNB was detected in an immunocytochemical section of
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000136068-FLNB/tissue/cerebral+cortex#imid_1781568
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000136068-FLNB/tissue/colon#imid_1781494
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000136068-FLNB/tissue/liver#imid_1781495
HPA · PancreasHuman Protein Atlas IHC, Pancreas. Source: https://www.proteinatlas.org/ENSG00000136068-FLNB/tissue/pancreas#imid_1781462
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB019322; green = FLNB, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB019322; green = FLNB, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for FLNB IHC & IF
🧪Run colon first: its glandular cells should stain for FLNB (HPA: High in colon glandular cells). Use lymph node germinal center cells as the negative tissue comparison (HPA: Not detected in lymph node germinal center cells); assess nonglandular cells on the colon slide for background, but do not assume they are FLNB-negative (HPA: High is specified for glandular cells).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Caudate (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show FLNB in A-431, U-251MG, U2OS, with annotated localisation: Plasma membrane (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched nonimmune rabbit IgG isotype control (selected-SKU caption: rabbit primary antibody); a confirmed FLNB knockout specimen provides a biological negative. For colon, check endogenous peroxidase and biotin when using the caption’s biotin-based DAB detection (selected-SKU caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU tissue-IHC caption does not report a fixative (selected-SKU caption: fixative not stated). The paraffin-section example uses heat retrieval in EDTA at pH 8.0, but the evidence does not establish whether FLNB staining requires that retrieval condition (selected-SKU caption: EDTA retrieval). HPA provides ICC-IF images, but the supplied evidence does not establish that frozen sections or IF are easier than paraffin IHC (HPA: ICC-IF images in A-431, U-251MG and U2OS); endogenous biotin in colon epithelium is a potential artefact with the caption’s SABC detection method (selected-SKU caption: SABC).
HPA tissue IHC evidence for FLNB
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data. Paired antibodies with high similarity supports the protein expression profile.). 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 → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Colon | Glandular cells | High | 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 → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Hippocampus | Glial cells | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced FLNB IHC Tips
Troubleshoot FLNB staining in paraffin sections by checking retrieval, cellular pattern and tissue controls before assigning a biological interpretation.
What retrieval conditions should I try first when FLNB staining is weak?
Start with heat-mediated retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A02562-1). Keep the retrieval method consistent across comparison slides, then repeat the catalog antibody’s reported incubation at 2 μg/ml overnight at 4°C to isolate the retrieval variable (datasheet A02562-1; standard IHC practice). If staining remains weak, compare a second retrieval condition on serial sections while holding antibody concentration and detection constant; that alternative is an optimisation trial, not a reported condition for this antibody (standard IHC practice). Assess endothelial staining alongside the cells of interest, since FLNB is highly abundant in endothelial cells (HPA tissue IHC).
Can I attribute weak FLNB staining to overfixation?
Target-specific fixation sensitivity is unknown: the catalog image shows a paraffin-embedded human lung cancer section but does not state its fixative (datasheet A02562-1). Record the fixative and processing history for each specimen, and compare similarly processed sections before attributing a difference to FLNB abundance (standard IHC practice). If staining varies with processing, run matched sections through the same EDTA pH 8.0 retrieval and 2 μg/ml primary incubation used in the catalog example (datasheet A02562-1). Use an internal positive cell population and a no-primary control to distinguish loss of detectable signal from widespread staining or detection failure (standard IHC practice).
Should FLNB stain the membrane, cytoplasm, or both in tissue sections?
Expect cytoplasmic and membranous staining in many cells, with particularly abundant endothelial staining (HPA tissue IHC). FLNB is a cytoplasmic, cortical and cytoskeletal protein without a transmembrane segment, so a peripheral pattern can reflect its association with membrane-linked actin rather than passage through the membrane (UniProt O75369 localisation, topology and function). Compare staining with cell borders and cytoplasm on the counterstained section, then examine whether the pattern repeats in intact cells away from section edges (standard IHC practice). Treat isolated nuclear staining cautiously because the supplied localisation records place FLNB mainly at the plasma membrane and in cytoplasmic structures (HPA subcellular; UniProt O75369 localisation).
Could an isoform difference explain discordant FLNB staining?
FLNB has 9 recorded isoforms, and their reported tissue distributions differ, so an antibody’s epitope matters when comparing specimens (UniProt O75369 isoforms and tissue specificity). The supplied catalog caption establishes tissue staining conditions but does not identify the antibody epitope or which isoforms it detects (datasheet A02562-1). Check the antibody’s documented immunogen or epitope against the isoforms relevant to your specimen before interpreting a weak or absent stain as loss of all FLNB (standard IHC practice; UniProt O75369 isoforms). If comparing antibodies, score matched cell populations and retrieval conditions; agreement between antibodies can strengthen an expression pattern but cannot by itself establish isoform specificity (standard IHC practice).
How should I follow up a chromogenic FLNB pattern with IF?
Use IF on its own validated workflow and multiplex FLNB with a marker for the expected cell population, such as an endothelial marker when assessing vessel-associated signal (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and inspect an unstained specimen for autofluorescence before assigning a dim FLNB signal (standard IF practice). FLNB lacks a transmembrane segment and has cytoplasmic and cortical locations; permeabilisation may be needed to expose an intracellular epitope, but the catalog caption does not identify that epitope (UniProt O75369 topology and localisation; datasheet A02562-1). Compare single-stain controls and cellular patterns before treating channel overlap as colocalisation (standard IF practice).
How can I reduce diffuse or vascular background without losing FLNB signal?
First compare the stained section with a no-primary control, since diffuse chromogen in both points to the detection workflow rather than specific primary-antibody binding (standard IHC practice). The catalog example used 10% goat serum block, 2 μg/ml primary overnight at 4°C, a biotinylated secondary for 30 minutes at 37°C, and SABC–DAB detection (datasheet A02562-1). Check peroxidase blocking and the contribution of biotin-based detection when background persists; these are general chromogenic IHC controls, not FLNB-specific findings (standard IHC practice). Preserve staining in intact endothelial cells while optimising washes or antibody concentration, because endothelial FLNB can be abundant (HPA tissue IHC; standard IHC practice).
How should I score FLNB staining across specimens with different cell composition? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because FLNB staining is cytoplasmic and membranous in many cells and is particularly abundant in endothelial cells (HPA tissue IHC). For a selected population, report the percentage of positive cells and an intensity-based H-score using the same scoring rules across sections (standard IHC practice). For vascular questions, report positive-cell or vessel-associated density per mm² of evaluable tissue, and document how vessels were identified (standard IHC practice). Normalise comparisons to the same tissue area or eligible cell count, exclude necrotic and folded regions, and keep acquisition and threshold settings consistent (standard IHC practice).
What distinguishes genuine FLNB staining from a tissue artefact?
A credible pattern follows intact cell borders or cytoplasm and can include strong endothelial staining, consistent with the reported tissue profile and FLNB’s cortical and cytoskeletal locations (HPA tissue IHC; UniProt O75369 localisation). Inspect the counterstain and adjacent morphology before calling a cell positive; staining confined to section edges, folds or necrosis is suspect (standard IHC practice). Check a no-primary control for endogenous enzyme or detection-system colour, particularly before interpreting weak diffuse DAB signal (standard IHC practice). Weigh unexpected nuclear-only staining against the supplied localisation evidence, and require a reproducible pattern in intact cells before making a biological claim (HPA subcellular; UniProt O75369 localisation; standard IHC practice).
Boster reagents
Best FLNB / Filamin-B IHC Antibodies
Two anti-FLNB antibodies have human paraffin-section IHC images and cell-based IF images (catalog image captions); one also lists mouse and rat reactivity (A02562-1 catalog reactivity).
Real IHC data
Anti-Filamin B/FLNB Antibody ®
Cat # A02562-1
Real IHC data
Anti-Filamin B/FLNB Antibody ® (monoclonal, 11E2D2)
Cat # M02562-1
A02562-1 lists IHC and IF/ICC applications with human, mouse, and rat reactivity; its images show human paraffin sections and U2-OS cells (A02562-1 applications, reactivity, and image captions). M02562-1 lists IHC and IF/ICC applications with human reactivity; its images show human paraffin sections and Hela cells (M02562-1 applications, reactivity, and image captions).
Which to pick: For human tissue IHC, either SKU has its own paraffin-section figure using EDTA retrieval; the fixative is unreported in both captions (A02562-1 and M02562-1 IHC captions). For IF/ICC, choose A02562-1 for its U2-OS image or M02562-1 if a monoclonal antibody and its Hela image suit the experiment (A02562-1 IF caption; M02562-1 clone and IF caption). For mouse or rat work, choose A02562-1 based on its listed reactivity and IHC application; its IHC image documents a human paraffin section (A02562-1 reactivity, applications, and IHC caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O75369 (FLNB_HUMAN, Filamin-B).
- Human Protein Atlas. FLNB tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. FLNB subcellular location (ICC-IF): Mainly localized to the plasma membrane. In addition localized to the Golgi apparatus, cytosol and actin filaments..
- Human Protein Atlas. FLNB antibody validation summary (3 antibodies).
- Single-cell-resolved differentiation of human induced pluripotent stem cells into pancreatic duct-like organoids on a microwell chip. Nature biomedical engineering 2021 — PMC7611572.
- The molecular basis of force selectivity by PIEZO2. Nature 2026 — PMC13149025.
- Biallelic pathogenic variants in FLNB are associated with paediatric steroid-resistant nephrotic syndrome via podocyte cytoskeletal dysfunction. Journal of medical genetics 2026 — PMC13539846.
- TGFβ and BMP Dependent Cell Fate Changes Due to Loss of Filamin B Produces Disc Degeneration and Progressive Vertebral Fusions. PLoS genetics 2016 — PMC4809497.
- PubMed PMID:9651345 — UniProt-cited evidence.
- PubMed PMID:9694715 — UniProt-cited evidence.
- PubMed PMID:11807098 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for FLNB (Filamin-B). Expected localisation Cytoplasmic and membranous tissue staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02562-1). Validated IHC antibody: A02562-1 at 1-2 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Caudate. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung cancer tissue; fixative not specified (datasheet A02562-1); 2. Fixation: Image fixative and duration unreported (datasheet A02562-1); 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 A02562-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02562-1); 8. Primary antibody: Rabbit anti-FLNB, 1-2 μg/ml (datasheet A02562-1); 9. Primary incubation: Overnight at 4 °C (datasheet A02562-1). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Cardiomyocytes may lack detectable staining (HPA tissue IHC); Isoform / epitope: Nine isoforms; check whether the epitope spans splice variants (UniProt); Regulation: Staining varies by cell type (HPA tissue IHC). Evidence: UniProt O75369, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the plasma membrane. In addition localized to…, Datasheet A02562-1, PMC7611572. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/FLNB/immunohistochemistry-ihc-guide.