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ATP1B1 / Sodium/potassium-transporting ATPase subunit beta-1 · IHC design guide
Design Immunohistochemistry for ATP1B1
Plan chromogenic ATP1B1 IHC in paraffin sections using the membranous tissue pattern (HPA tissue IHC). This guide highlights high staining in collecting ducts and glandular cells (HPA tissue IHC) and the catalog antibody’s 2–5 μg/ml IHC range (datasheet A03469-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ATP1B1 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 | Membranous staining in many tissues (HPA tissue IHC) | |
| Staining pattern | Membranous pattern; high in glandular cells and collecting ducts (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A03469-1) | |
| Positive control | Duodenum+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining and RNA show medium consistency (HPA tissue IHC) | |
| Regulation | Kidney-enhanced RNA expression (HPA tissue RNA) | |
| Isoform / epitope | 2 isoforms; epitope coverage is unknown; topology spans both sides (UniProt) |
Section 1
Recommended ATP1B1 IHC & IF Protocols
The catalog antibody protocol is accompanied by published ATP1B1 staining methods for rat CA1 sections, lung cancer tissue microarrays, and corneas (PMC5970211; PMC7174014; PMC5379100).
| Sample | Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A03469-1) |
| Fixation | Image fixative and duration unreported (datasheet A03469-1); 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 A03469-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A03469-1) |
| Primary antibody | Rabbit anti-ATP1B1, 2-5 μg/ml (datasheet A03469-1) |
| Primary incubation | Overnight at 4 °C (datasheet A03469-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A03469-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | ATP1B1-positive staining in glandular cells of duodenum (HPA tissue IHC: High). HPA tissue profile: Membranous expression in many tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for paraffin sections (datasheet A03469-1). A corneal confirmatory assay omitted retrieval (PMC5379100).
Section 2
What Is the Expected ATP1B1 Staining Pattern?
ATP1B1 is a membrane protein with one transmembrane segment and a large extracellular region (UniProt P05026 topology). In paraffin-section IHC, expect membranous staining in many tissues, including high staining in kidney collecting ducts and glandular cells of the duodenum, gallbladder, salivary gland, and small intestine (HPA: tissue IHC). HPA rates the tissue IHC pattern Enhanced, with medium consistency between staining and RNA data (HPA: tissue IHC reliability).
| Membrane outlines in kidney collecting ducts and glandular cells, with strong signal in the listed high-staining tissues (HPA: tissue IHC). | This fits ATP1B1's cell-membrane location and HPA's membranous IHC profile (UniProt P05026 subcellular location; HPA: tissue IHC). Judge the membrane pattern and the stained cell population together; tissue-wide color alone is less informative. |
| Predominantly nuclear or uniform cytoplasmic color, without a convincing membrane pattern. | That distribution conflicts with the principal membrane localization (UniProt P05026 subcellular location; HPA: tissue IHC). Treat it as possible nonspecific staining or a detection artifact, and review the controls before assigning it to ATP1B1. |
| Strong staining in a cell population HPA reports as not detected, such as adipocytes in adipose tissue (HPA: not detected in adipocytes). | The mismatch raises concern for cross-reactivity or endogenous detection activity. It does not prove either cause: HPA reports tissue and cell-specific observations, and its IHC profile has medium consistency with RNA data (HPA: tissue IHC reliability). |
| Broad, low-contrast color across cells and surrounding section areas, obscuring membrane outlines. | This is background rather than a readily interpretable membrane pattern (HPA: membranous tissue IHC profile). Compare a no-primary control and assess blocking, washes, antibody concentration, and chromogen development as general IHC checks. |
| No detectable signal in kidney collecting ducts or another listed high-staining population (HPA: tissue IHC). | The result falls short of the supplied positive reference pattern. Check a same-run positive section and reagent performance before calling the sample negative; HPA's Enhanced rating supports the reported pattern but does not guarantee every section will stain (HPA: tissue IHC reliability). |
💡Expected ATP1B1 appearanceA convincing positive shows distinct membranous signal in the listed high-staining collecting-duct or glandular cells, while dominant nuclear color or indiscriminate tissue-wide color is suspect (HPA: tissue IHC; UniProt P05026 subcellular location).
| Topology and epitope access (UniProt P05026 topology) | ATP1B1 spans the membrane at residues 35–62, with residues 1–34 cytoplasmic and 63–303 extracellular (UniProt P05026 topology). An antibody's mapped epitope would guide interpretation of access after tissue processing; no epitope map or ATP1B1-specific retrieval effect is supplied here. |
| Tissue and cell population (HPA: tissue IHC) | HPA reports high staining in kidney collecting ducts and selected glandular cells, medium staining in appendix glandular cells and cerebral-cortex neuropil, and no detection in several specified populations (HPA: tissue IHC). Compare the same cell type when judging intensity. |
| Validation and agreement (HPA: tissue IHC reliability; HPA: HPA012911 IHC Enhanced) | The supplied antibody has an Enhanced IHC status, while the tissue profile has medium consistency between antibody staining and RNA expression (HPA: HPA012911 IHC Enhanced; HPA: tissue IHC reliability). Use the reported distribution as a reference, with section-level controls. |
| Protein variants and glycosylation (UniProt P05026) | UniProt lists two isoforms and three glycosylation sites at residues 158, 193, and 265 (UniProt P05026 isoforms and glycosylation). Their effect on this antibody's paraffin-section staining cannot be determined without an antibody epitope or assay-specific evidence. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in a kidney collecting-duct positive reference (HPA: High in collecting ducts). | A weak IHC run or missed target remains possible; a blank reference section alone cannot distinguish them. | Review section quality, primary-antibody dilution, antigen retrieval, detection reagents, and chromogen development as general IHC checks. Run a listed high-staining section alongside the test tissue (HPA: tissue IHC). |
| Color is mainly nuclear or diffuse within cells. | The location disagrees with the reported membrane pattern (UniProt P05026 subcellular location; HPA: tissue IHC). | Check a no-primary control, inspect membrane outlines at appropriate magnification, and reassess blocking and detection conditions. Do not score nuclear color alone as ATP1B1-positive. |
| Unexpected strong color appears in an HPA not-detected cell population (HPA: tissue IHC). | Cross-reactivity or endogenous detection activity is possible; the slide does not identify which. | Compare the no-primary control and a listed positive cell population on the same run. Score the named cell population rather than treating an entire organ as uniformly positive or negative (HPA: tissue IHC). |
| Diffuse background hides cell borders. | Excess primary antibody, incomplete blocking or washing, or detection background are general IHC possibilities. | Use the no-primary control to locate detection background; then adjust blocking, washes, primary dilution, or development time one variable at a time. Reassess whether a membrane pattern emerges (HPA: tissue IHC profile). |
| Staining intensity varies across tissues or cell populations. | HPA reports high, medium, low, and not-detected levels in different named populations (HPA: tissue IHC). | Record compartment and intensity for each cell population. Compare with the corresponding HPA entry; avoid using kidney collecting-duct intensity as a required threshold for all tissues (HPA: High in collecting ducts). |
| Does the IHC pattern establish what IF/ICC should show? | HPA summarizes the IF/ICC location as membrane but supplies no ICC/IF image-bearing cell lines or detailed main location (HPA: subcellular ICC-IF). | Use membrane localization as the limited expectation (UniProt P05026 subcellular location; HPA: subcellular ICC-IF). Interpret IF/ICC with its own controls and guide; the supplied IHC evidence does not establish an IF/ICC protocol. |
Experiment evidence
IHC & IF images showing ATP1B1 staining
A03469-1 · IHCIHC analysis of ATP1B1 using anti-ATP1B1 antibody (A03469-1). ATP1B1 was detected in a paraffin-embedded section of human breast cancer tiss
A03469-1 · IHCIHC analysis of ATP1B1 using anti-ATP1B1 antibody (A03469-1). ATP1B1 was detected in a paraffin-embedded section of human colonic adenocarci
A03469-1 · IFIF analysis of ATP1B1 using anti-ATP1B1 antibody (A03469-1). ATP1B1 was detected in an immunocytochemical section of SiHa cells. Enzyme anti
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000143153-ATP1B1/tissue/cerebral+cortex#imid_20780888
HPA · Parathyroid glandHuman Protein Atlas IHC, Parathyroid gland. Source: https://www.proteinatlas.org/ENSG00000143153-ATP1B1/tissue/parathyroid+gland#imid_20780779
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000143153-ATP1B1/tissue/colon#imid_20780806
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000143153-ATP1B1/tissue/liver#imid_20780823
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ATP1B1 IHC & IF
🧪Run kidney first: collecting duct cells should stain strongly (HPA: High in kidney collecting ducts). Use adipose tissue as the negative comparator (HPA: Not detected in adipocytes); on the kidney slide, cells outside stained collecting ducts can show background staining, but should not be assumed to be ATP1B1-negative.
Positive control tissue: Duodenum (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for ATP1B1; derive a cell-line control from the positive tissue's cell type (Glandular cells) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit IgG isotype control (caption: rabbit primary; standard IHC practice). Use ATP1B1 knockout tissue or a validated peptide block as a biological specificity control, and check for endogenous peroxidase and biotin signal in kidney when using the caption’s biotin-based DAB detection (standard IHC practice; caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A03469-1 paraffin-section caption does not state a fixative (caption: fixative unreported). The caption uses heat-mediated retrieval in EDTA at pH 8.0, but does not establish that retrieval is required for ATP1B1 (caption: EDTA retrieval). The supplied evidence does not establish whether frozen sections or IF are easier; for kidney IHC, assess endogenous biotin signal because the caption uses SABC detection (caption: SABC; standard IHC practice).
HPA tissue IHC evidence for ATP1B1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — 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 |
|---|---|---|---|---|
| Duodenum | Glandular cells | High | Protein (IHC) | HPA → |
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Kidney | Collecting ducts | High | Protein (IHC) | HPA → |
| Salivary gland | Glandular cells | High | Protein (IHC) | HPA → |
| Small intestine | Glandular cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
| Cervix | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Esophagus | Squamous epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ATP1B1 IHC Tips
Troubleshoot ATP1B1 staining by checking retrieval, membrane localisation and cell type before comparing signal intensity across paraffin sections.
Which retrieval condition should I try first for weak ATP1B1 staining in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet A03469-1). The catalog antibody stained a paraffin section after that retrieval, followed by 2 μg/ml primary antibody overnight at 4°C (datasheet A03469-1). If staining remains weak, compare a second retrieval buffer on adjacent sections while holding antibody concentration, detection and development constant (general IHC practice). Check whether any gained signal follows cell borders, since ATP1B1 is membrane associated (UniProt P05026 topology; HPA: membranous expression). Compare against a known positive compartment, such as kidney collecting ducts, before accepting stronger diffuse staining as an improvement (HPA: High in kidney collecting ducts).
How should I troubleshoot fixation-related loss of ATP1B1 staining?
The selected paraffin-section caption does not report its fixative, so ATP1B1 sensitivity to fixation cannot be determined from it (datasheet A03469-1: fixative not stated). Record the fixative and processing history for each section, then compare similarly processed positive and test tissue with the same EDTA pH 8.0 retrieval (datasheet A03469-1; general IHC practice). Include a no-primary control to distinguish processing-related background from antibody signal (general IHC practice). Assess whether staining outlines cells in the expected compartment rather than relying on total brown intensity (HPA: membranous expression; general IHC practice). Avoid attributing differences between differently fixed specimens specifically to ATP1B1 without a controlled comparison (general IHC practice).
What staining pattern is credible for ATP1B1, and how should I assess cytoplasmic signal?
Expect a predominantly membranous pattern across many tissues, while judging the relevant cell population separately (HPA: membranous expression in many tissues). ATP1B1 has one transmembrane segment at residues 35–62, with residues 1–34 cytoplasmic and 63–303 extracellular (UniProt P05026 topology). Review intact cell borders at the same magnification used for controls; diffuse cytoplasmic DAB alone is less persuasive than a reproducible membrane outline (UniProt P05026 localisation; general IHC practice). In epithelia, inspect lateral borders because ATP1B1 contributes to lateral pump polarisation and cell adhesion (UniProt P05026 function). Keep scoring compartment specific when both border and cytoplasmic colour appear (general IHC practice).
Can this staining distinguish ATP1B1 isoforms or reveal which side of the membrane is recognised?
ATP1B1 has 2 annotated isoforms, but the supplied caption does not identify an antibody epitope or establish isoform selectivity (UniProt P05026 isoforms; datasheet A03469-1 caption). Do not assign isoform 1 or 2 from DAB staining alone (UniProt P05026 isoforms; general IHC practice). The protein spans the membrane at residues 35–62, leaving residues 1–34 cytoplasmic and 63–303 extracellular (UniProt P05026 topology). Three annotated glycosylation sites occur at residues 158, 193 and 265; their effects on this antibody's tissue staining are untested here (UniProt P05026 glycosylation; datasheet A03469-1 caption). Obtain epitope or isoform-specific validation before interpreting compartmental differences as isoform changes (general IHC practice).
How can I plan an IF follow-up to a convincing ATP1B1 IHC pattern?
For multiplex IF, pair ATP1B1 with a validated marker of the cell population being examined, and compare the resulting cell borders with the chromogenic IHC pattern (HPA: membranous expression; general IF practice). Choose fluorophores after viewing an unstained section in each channel, placing the ATP1B1 readout in a channel with low tissue autofluorescence (general IF practice). If the antibody recognises the extracellular region at residues 63–303, test minimal permeabilisation; access to the cytoplasmic region at residues 1–34 generally requires permeabilisation (UniProt P05026 topology; general IF practice). The supplied evidence does not locate this antibody's epitope, so compare permeabilised and unpermeabilised conditions experimentally (datasheet A03469-1 caption; general IF practice). Keep single-label and secondary-only controls for bleed-through and background assessment (general IF practice).
How do I separate ATP1B1 signal from chromogenic IHC background?
The selected example used 10% goat serum block, biotinylated goat anti-rabbit secondary antibody, an avidin–biotin detection complex and DAB (datasheet A03469-1). Include a no-primary section and a peroxidase block when using enzyme-based chromogenic detection (general IHC practice). Because the example used a biotin-based system, assess endogenous biotin as a possible contributor if staining persists without primary antibody (datasheet A03469-1; general IHC practice). Compare background with membrane outlines in a positive population such as kidney collecting ducts, reported as High (HPA: kidney collecting ducts). Reduce nonspecific colour through controlled antibody titration and development time, preserving the same conditions across comparison sections (general IHC practice).
What should I measure when comparing ATP1B1 IHC across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and membrane compartment before scoring, since the reported tissue pattern is membranous and expression varies by cell type (HPA: membranous expression and tissue profile). For comparable sections, record the percentage of membrane-positive target cells and membrane intensity, then calculate an H-score if the intensity scale is predefined (general IHC practice). Alternatively, measure positive membrane area or cell density per mm² within a fixed tissue region (general IHC practice). Normalise to evaluable target cells or tissue area, excluding necrosis and damaged edges by the same rules in every section (general IHC practice). Keep retrieval, 2 μg/ml primary concentration and DAB development consistent when comparing the catalog workflow (datasheet A03469-1; general IHC practice).
When is an apparent ATP1B1-positive cell likely to be a staining artefact?
Give greatest weight to reproducible membrane staining in an appropriate cell population, consistent with the reported membranous tissue profile (HPA: membranous expression in many tissues). High staining in kidney collecting ducts or glandular cells of duodenum can serve as an anatomical comparison, whereas adipocytes are reported as Not detected (HPA: kidney collecting ducts High; duodenum glandular cells High; adipocytes Not detected). Isolated nuclear colour or uniform staining across unrelated compartments warrants review against no-primary controls and section morphology (UniProt P05026 localisation; general IHC practice). Inspect cut edges and necrotic regions separately, where staining artefacts can obscure the cell-border pattern (general IHC practice). Residual colour after omitting primary antibody can implicate endogenous enzyme activity or the detection system rather than ATP1B1 (general IHC practice).
Boster reagents
Best ATP1B1 / Sodium/potassium-transporting ATPase subunit beta-1 IHC Antibodies
The IHC-validated antibody has paraffin-section images from human breast, colon, gall bladder and liver cancers, plus IF images from human cells and mouse and rat kidney (A03469-1 image captions).
Real IHC data
Anti-ATP1B1 Antibody ®
Cat # A03469-1
The rendered card is A03469-1, with its own IHC image from a human breast cancer paraffin section (A03469-1 IHC caption). Its other captions show human colon, gall bladder and liver cancer IHC, human cell ICC/IF, and mouse and rat kidney IF (A03469-1 image captions).
Which to pick: Choose A03469-1 for tissue IHC when a documented paraffin-section example is useful: its IHC caption specifies EDTA retrieval at pH 8.0 and 2 μg/ml primary antibody; the fixative is unreported (A03469-1 IHC caption). Choose A03469-1 for IF/ICC because its application list includes both and its captions show human cell ICC/IF and mouse and rat kidney IF (A03469-1 catalog; A03469-1 IF captions). For cross-species IHC, both SKUs list human, mouse and rat reactivity, but A03469-1 has an illustrated human paraffin IHC example; M03469 is a rabbit monoclonal, clone 23A21, listed for IHC without an IHC image caption (catalog).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P05026 (AT1B1_HUMAN, Sodium/potassium-transporting ATPase subunit beta-1).
- Human Protein Atlas. ATP1B1 tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. ATP1B1 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. ATP1B1 antibody validation summary (1 antibodies).
- Neuroglobin mediates neuroprotection of hypoxic postconditioning against transient global cerebral ischemia in rats through preserving the activity of Na(+)/K(+) ATPases. Cell death & disease 2018 — PMC5970211.
- Na/K-ATPase Gene Expression in the Human Cochlea: A Study Using mRNA in situ Hybridization and Super-Resolution Structured Illumination Microscopy. Frontiers in molecular neuroscience 2022 — PMC9009265.
- Immunohistochemical validation study of 15-gene biomarker panel predictive of benefit from adjuvant chemotherapy in resected non-small-cell lung cancer: analysis of JBR.10. ESMO open 2020 — PMC7174014.
- Genome-wide association study identifies three novel loci in Fuchs endothelial corneal dystrophy. Nature communications 2017 — PMC5379100.
- PubMed PMID:3008098 — UniProt-cited evidence.
- PubMed PMID:2559024 — UniProt-cited evidence.
- PubMed PMID:7536695 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ATP1B1 (Sodium/potassium-transporting ATPase subunit beta-1). Expected localisation Membranous staining in many tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A03469-1). Validated IHC antibody: A03469-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Duodenum (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast cancer tissue; fixative not specified (datasheet A03469-1); 2. Fixation: Image fixative and duration unreported (datasheet A03469-1); 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 A03469-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A03469-1); 8. Primary antibody: Rabbit anti-ATP1B1, 2-5 μg/ml (datasheet A03469-1); 9. Primary incubation: Overnight at 4 °C (datasheet A03469-1). Watch for: Fixation: Keep fixation consistent across paraffin sections. (standard IHC practice; not target-specific); Caveat: Antibody staining and RNA show medium consistency (HPA tissue IHC); Isoform / epitope: 2 isoforms ; epitope coverage is unknown; topology spans both sides (UniProt); Regulation: Kidney-enhanced RNA expression (HPA tissue RNA). Evidence: UniProt P05026, HPA tissue IHC · Enhanced, HPA ICC-IF · Membrane, Datasheet A03469-1, PMC5970211, PMC7174014. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ATP1B1/immunohistochemistry-ihc-guide.