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MAD2L2 / Mitotic spindle assembly checkpoint protein MAD2B · IHC design guide
Design Immunohistochemistry for MAD2L2
Plan MAD2L2 IHC in paraffin sections using the catalog antibody at 2–5 μg/ml (datasheet A02357-4). Expect cytoplasmic staining in most tissue types, and compare signal within the same cell type when scoring (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
MAD2L2 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 tissue types (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining across most tissue types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02357-4) | |
| Positive control | Bone marrow+4 more · see all | |
| Negative control | Breast+2 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 | Lung macrophages stain strongly; score cell types separately (HPA tissue IHC) | |
| Regulation | No stimulus-linked regulation specified (UniProt) | |
| Isoform / epitope | One 1–211 chain; no listed isoforms or processing (UniProt) |
Section 1
Recommended MAD2L2 IHC & IF Protocols
Compare the catalog antibody’s IHC-P protocol (datasheet A02357-4) with two published MAD2L2 IHC workflows (PMC10956193; PMC6090455).
| Sample | Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A02357-4) |
| Fixation | Image fixative and duration unreported (datasheet A02357-4); 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 A02357-4); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02357-4) |
| Primary antibody | Rabbit anti-MAD2L2, 2-5 μg/ml (datasheet A02357-4) |
| Primary incubation | Overnight at 4 °C (datasheet A02357-4) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A02357-4) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | MAD2L2-positive staining in hematopoietic cells of bone marrow (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissue types. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A02357-4). Citrate pH 6.0 was used with a different antibody (PMC6090455).
Section 2
What Is the Expected MAD2L2 Staining Pattern?
MAD2L2 is found in the cytoplasm and nucleus, with no transmembrane segment (UniProt Q9UI95). In paraffin section IHC, expect mainly cytoplasmic staining across many tissues, especially bone marrow hematopoietic cells, lung macrophages and testis Leydig cells (HPA tissue IHC). HPA rates the tissue IHC evidence Approved, with medium consistency between staining and RNA expression; use its cell level observations as guides rather than universal cutoffs (HPA tissue IHC).
| Cytoplasmic signal in bone marrow hematopoietic cells, lung macrophages or testis Leydig cells. | This matches the High staining reported in each cell population and the mainly cytoplasmic tissue profile (HPA tissue IHC). Assess the relevant cells within each section; staining elsewhere in the tissue does not replace that comparison. |
| Predominantly membranous or extracellular staining, without the expected cytoplasmic pattern. | Treat this as a possible artefact: MAD2L2 has no transmembrane segment or signal peptide, and HPA describes cytoplasmic tissue IHC (UniProt Q9UI95; HPA tissue IHC). Nuclear signal alone needs separate assessment because nuclear localisation is also reported (UniProt Q9UI95). |
| Strong signal in a cell population reported as undetected, such as skeletal muscle myocytes. | Consider cross-reactivity or endogenous detection activity and check controls before scoring it as MAD2L2. The undetected call applies to the named cells in the sampled tissue; it does not establish that every cell there must be negative (HPA tissue IHC). |
| Diffuse chromogen across cells and surrounding tissue, obscuring cell boundaries. | The distribution cannot support a reliable cell level call. Check the no-primary control and detection background, then reassess whether distinct cytoplasmic signal remains in an HPA High population (general IHC practice; HPA tissue IHC). |
| No signal in bone marrow hematopoietic cells, lung macrophages or testis Leydig cells. | A blank result in a reported High population raises concern about the staining run or antibody performance (HPA tissue IHC). Verify tissue identity, section quality and a working positive control before interpreting other samples as negative (general IHC practice). |
💡Expected MAD2L2 appearanceCall a result positive when identifiable bone marrow hematopoietic cells, lung macrophages or testis Leydig cells show clear, predominantly cytoplasmic High staining; diffuse background or isolated membranous deposits are suspect (HPA tissue IHC; UniProt Q9UI95).
| Tissue and cell selection | Expression is ubiquitous by UniProt, but HPA reports different IHC levels by cell population: High in three listed positive populations and undetected in breast adipocytes, hippocampal glia and skeletal muscle myocytes (UniProt Q9UI95; HPA tissue IHC). Choose controls by named cell type, not organ name alone. |
| IF/ICC Q: Should nuclear staining be expected? | Yes. HPA reports supported nucleoplasmic localisation in ICC-IF, while its tissue IHC summary is cytoplasmic; UniProt also lists nucleus and cytoplasm (HPA subcellular; HPA tissue IHC; UniProt Q9UI95). Interpret each assay against its own observed pattern. |
| Antibody validation | CAB008110 is Approved for IHC, while HPA003882 is Supported for ICC-IF; those labels apply to different antibodies and applications (HPA antibodies). HPA's tissue IHC reliability is Approved with medium staining versus RNA consistency, so an unexpected slide still needs controls (HPA tissue IHC). |
| Topology and processing | The annotated protein is a single 1–211 chain with no signal peptide, propeptide, transmembrane segment or glycosylation sites (UniProt Q9UI95). These annotations support scrutiny of membrane or extracellular staining; they do not establish epitope access or fixation sensitivity. |
| Situation | Likely cause | Next action |
|---|---|---|
| The reported High cells are blank. | A failed staining run, unsuitable section or ineffective antigen retrieval is possible; no MAD2L2-specific retrieval requirement is supplied (HPA tissue IHC; general IHC practice). | Check tissue identity and a run positive control, then review the antibody's validated IHC conditions and the run's retrieval and detection steps (general IHC practice). |
| Macrophages or hematopoietic cells stain heavily in an HRP chromogen run. | These are HPA High populations, but endogenous peroxidase can also contribute chromogen in some cells (HPA tissue IHC; general IHC practice). | Compare a no-primary control and confirm the peroxidase blocking step before attributing all signal to MAD2L2 (general IHC practice). |
| A skeletal muscle myocyte appears strongly positive. | HPA reports myocytes as undetected; cross-reactivity, background or cell misidentification could explain the mismatch (HPA tissue IHC). | Confirm the cell type and staining compartment, inspect controls, and repeat with an independently validated IHC antibody if the finding matters (general IHC practice). |
| The section shows mainly membrane or extracellular deposits. | That pattern conflicts with the HPA cytoplasmic IHC profile and UniProt's lack of a transmembrane segment or signal peptide (HPA tissue IHC; UniProt Q9UI95). | Inspect the no-primary control and chromogen precipitate, then score only cell associated staining that remains credible after background review (general IHC practice). |
| Staining is faint in liver hepatocytes or bronchial respiratory epithelial cells. | HPA reports Low staining in both populations, so weak signal there alone does not establish a failed assay (HPA tissue IHC). | Compare with a named High population in the same run before changing the assay or calling the sample negative (HPA tissue IHC; general IHC practice). |
| IHC looks cytoplasmic, but an ICC-IF image looks nuclear. | The observations come from different assays: HPA tissue IHC is chiefly cytoplasmic, and HPA ICC-IF supports nucleoplasmic localisation (HPA tissue IHC; HPA subcellular). | Check each result against its assay specific evidence and antibody validation; do not use the ICC-IF localisation alone to reject a controlled cytoplasmic IHC result (HPA antibodies; HPA tissue IHC; HPA subcellular). |
Experiment evidence
IHC & IF images showing MAD2L2 staining
A02357-4 · IHCIHC analysis of MAD2L2 using anti-MAD2L2 antibody (A02357-4). MAD2L2 was detected in a paraffin-embedded section of human liver cancer tissu
A02357-4 · IFIF analysis of MAD2L2 using anti-MAD2L2 antibody (A02357-4). MAD2L2 was detected in an immunocytochemical section of U2OS cells. Enzyme anti
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000116670-MAD2L2/tissue/cerebral+cortex#imid_2271204
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000116670-MAD2L2/tissue/colon#imid_2271138
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000116670-MAD2L2/tissue/liver#imid_2271139
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000116670-MAD2L2/tissue/kidney#imid_2271132
HPA · ICC-IF REHHuman Protein Atlas immunofluorescence, REH, antibody HPA003882; green = MAD2L2, blue = nucleus, red = microtubules.
HPA · ICC-IF REHHuman Protein Atlas immunofluorescence, REH, antibody HPA003882; green = MAD2L2, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for MAD2L2 IHC & IF
🧪Run lung first: macrophages should stain (HPA: High in lung macrophages). Use skeletal muscle myocytes as the negative tissue (HPA: Not detected in skeletal muscle myocytes); on the lung slide, candidate internal negative cells should show counterstain without specific signal, but their negative status must be verified rather than assumed from cell type (standard IHC practice).
Positive control tissue: Bone marrow (Hematopoietic cells, HPA High)
Negative control tissue: Breast (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show MAD2L2 in REH, SK-MEL-30, U2OS, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control; a rabbit IgG control matched for isotype and clonality where applicable (caption: rabbit primary antibody); and a MAD2L2 knockout control or peptide-block control if the immunizing peptide is available (standard IHC practice). Check endogenous peroxidase activity in lung macrophages before interpreting DAB staining (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A02357-4 paraffin-section caption does not state the fixative (caption: fixative unreported). The demonstrated IHC procedure uses heat retrieval in EDTA at pH 8.0, but the caption does not establish that retrieval is required (caption: EDTA retrieval, pH 8.0). Frozen sections have no demonstrated advantage here; ICC-IF images support nucleoplasmic localization but do not establish easier tissue IF, while lung macrophage peroxidase can complicate chromogenic interpretation (HPA: nucleoplasm supported; standard IHC practice).
HPA tissue IHC evidence for MAD2L2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 |
|---|---|---|---|---|
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Lung | Macrophages | High | Protein (IHC) | HPA → |
| Testis | Leydig cells | High | Protein (IHC) | HPA → |
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
Section 3
Advanced MAD2L2 IHC Tips
Troubleshoot MAD2L2 staining in paraffin sections by checking retrieval, compartment, cell identity and controls before comparing chromogenic signal across samples.
How should I adjust retrieval when MAD2L2 staining is weak?
Start with heat-mediated retrieval in EDTA pH 8.0 for paraffin sections stained with A02357-4 (datasheet A02357-4). The catalog image used 2 μg/ml primary antibody overnight at 4°C; hold that condition steady while recording retrieval heat exposure and cooling (caption A02357-4; standard IHC practice). If staining remains weak, compare an alternative buffer or heat exposure on adjacent sections, checking tissue morphology and background before changing antibody concentration (standard IHC practice). Include a no-primary control and a tissue compartment with expected signal when judging whether retrieval changed specific staining (HPA: high in lung macrophages; standard IHC practice).
Could fixation explain inconsistent MAD2L2 staining between paraffin samples?
Target-specific fixation sensitivity is unknown: the A02357-4 tissue caption identifies a paraffin section but does not state its fixative (caption A02357-4). Record each sample’s fixative, duration and processing history, and compare sections processed together before attributing intensity differences to biology (standard IHC practice). Excessive or uneven fixation can alter antigen accessibility and morphology in IHC generally, but no MAD2L2-specific effect is established here (standard IHC practice; caption A02357-4). Repeat the validated EDTA pH 8.0 retrieval under consistent conditions and inspect positive and no-primary controls alongside the test sections (datasheet A02357-4; standard IHC practice).
Should MAD2L2 appear in nuclei or cytoplasm on IHC?
Both compartments warrant evaluation: tissue IHC reports predominantly cytoplasmic staining, while subcellular imaging supports nucleoplasmic localisation (HPA tissue IHC; HPA subcellular). UniProt also places MAD2L2 at the spindle and chromosomes and reports recruitment to double-strand break sites during G1 and S phase (UniProt Q9UI95). Score nuclear and cytoplasmic chromogen separately within identified cell types, using the counterstain to define nuclei and excluding overlapping cells where assignment is uncertain (standard IHC practice). A change in compartment alone does not establish altered DNA repair activity; compare controls and tissue context before interpreting it (UniProt Q9UI95; standard IHC practice).
Could an isoform or hidden epitope explain discordant staining?
The supplied record annotates 0 isoforms and a 211-residue chain, so it does not identify an isoform-specific staining pattern (UniProt Q9UI95). Its HORMA domain spans residues 13–203, but the antibody’s immunogen coordinates are unavailable here; do not assume the recognized epitope lies in that domain (UniProt Q9UI95; caption A02357-4). MAD2L2 has no annotated transmembrane segment or modified residues in this record, which does not establish that every epitope survives processing (UniProt Q9UI95; standard IHC practice). If staining differs across samples, check retrieval and controls, then compare an independently validated epitope when available (standard IHC practice).
How can IF help assess a disputed IHC localisation pattern?
Use IF as an independent localisation check: nucleoplasmic staining is supported by subcellular imaging, whereas tissue IHC commonly shows cytoplasmic signal (HPA subcellular; HPA tissue IHC). For multiplexing, pair MAD2L2 with a marker identifying the cell population under study, such as a macrophage marker when assessing lung macrophages (HPA: high in lung macrophages; standard IF practice). Choose fluorophores outside prominent tissue autofluorescence and include single-channel controls to assess bleed-through (standard IF practice). Because MAD2L2 is intracellular and has no transmembrane segment, optimise permeabilisation for nuclear and cytoplasmic access; its antibody epitope location remains unspecified (UniProt Q9UI95; standard IF practice).
How do I distinguish MAD2L2 signal from brown background?
Run a no-primary section and inspect it for endogenous pigment, nonspecific secondary binding and enzyme-associated colour before scoring MAD2L2 (standard IHC practice). The A02357-4 paraffin image used 10% goat serum blocking, a peroxidase-conjugated secondary and DAB development; those conditions provide a starting comparison (caption A02357-4). Add an endogenous peroxidase block as a general chromogenic IHC step, then keep DAB development consistent between test and control sections (standard IHC practice). If diffuse staining persists, assess washing, primary concentration and secondary binding one variable at a time while retaining expected cell localisation as a specificity check (standard IHC practice; HPA tissue IHC).
What should I measure when MAD2L2 staining varies across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because tissue IHC reports broad cytoplasmic expression while subcellular imaging supports nucleoplasmic localisation (HPA tissue IHC; HPA subcellular). Record the percentage of positive viable cells and staining intensity separately; an H-score can combine percentages at intensity grades into a 0–300 summary (standard IHC practice). For focal populations, report positive-cell density per mm² of viable tissue and document the area selected (standard IHC practice). Normalise comparisons to the same cell type, compartment, tissue area and staining batch, and exclude necrosis and section edges by a prespecified rule (standard IHC practice).
When is apparent MAD2L2 positivity likely to be misleading?
Check cell identity first: high staining in lung macrophages is reported, whereas hepatocytes are listed as low, so adjacent cell populations can change an apparent tissue-level result (HPA tissue IHC). Cytoplasmic signal can be plausible in tissue IHC, and nucleoplasmic signal has independent support; either pattern still needs appropriate controls (HPA tissue IHC; HPA subcellular; standard IHC practice). Treat staining confined to cut edges, necrosis or a no-primary section as suspect, and check endogenous peroxidase contribution to DAB colour (standard IHC practice). HPA rates tissue staining as approved with medium RNA agreement, so interpret intensity alongside morphology and an independent comparator rather than as a direct activity readout (HPA tissue IHC; standard IHC practice).
Boster reagents
Best MAD2L2 / Mitotic spindle assembly checkpoint protein MAD2B IHC Antibodies
A02357-4 has IHC data from human liver cancer paraffin sections and IF data from U2OS cells (A02357-4 image captions). M02357 lists human, mouse and rat reactivity (M02357 catalog).
Real IHC data
Anti-MAD2L2 Antibody ®
Cat # A02357-4
A02357-4 will render with its IHC figure from human liver cancer paraffin tissue; its separate IF caption shows U2OS cells (A02357-4 image captions). M02357 will render with listed IHC and IF/ICC applications and human, mouse and rat reactivity, but no IHC or IF figure is supplied (M02357 catalog).
Which to pick: For tissue IHC, pick A02357-4: its own caption shows staining in a human liver cancer paraffin section at 2 μg/ml after EDTA retrieval at pH 8.0; the fixative is unreported (A02357-4 IHC image caption). For IF/ICC, pick A02357-4 because its IF caption shows U2OS cells stained at 5 μg/ml (A02357-4 IF image caption). For mouse or rat work, consider M02357, a rabbit monoclonal listed for IHC and IF/ICC in human, mouse and rat, with no image evidence supplied here (M02357 catalog).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q9UI95 (MD2L2_HUMAN, Mitotic spindle assembly checkpoint protein MAD2B).
- Human Protein Atlas. MAD2L2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. MAD2L2 subcellular location (ICC-IF): Localized to the nucleoplasm..
- Human Protein Atlas. MAD2L2 antibody validation summary (2 antibodies).
- Identification of a Novel CD8(+) T cell exhaustion-related gene signature for predicting survival in hepatocellular carcinoma. BMC cancer 2023 — PMC10694949.
- A critical function of Mad2l2 in primordial germ cell development of mice. PLoS genetics 2013 — PMC3757036.
- AURKB promotes bladder cancer progression by deregulating the p53 DNA damage response pathway via MAD2L2. Journal of translational medicine 2024 — PMC10956193.
- Effects of downregulated expression of microRNA-187 in gastric cancer. Experimental and therapeutic medicine 2018 — PMC6090455.
- PubMed PMID:10527948 — UniProt-cited evidence.
- PubMed PMID:10366450 — UniProt-cited evidence.
- PubMed PMID:10660610 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for MAD2L2 (Mitotic spindle assembly checkpoint protein MAD2B). Expected localisation Cytoplasmic staining in most tissue types (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02357-4). Validated IHC antibody: A02357-4 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Bone marrow (Hematopoietic cells); negative Breast. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human liver cancer tissue; fixative not specified (datasheet A02357-4); 2. Fixation: Image fixative and duration unreported (datasheet A02357-4); 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 A02357-4); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02357-4); 8. Primary antibody: Rabbit anti-MAD2L2, 2-5 μg/ml (datasheet A02357-4); 9. Primary incubation: Overnight at 4 °C (datasheet A02357-4). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Lung macrophages stain strongly; score cell types separately (HPA tissue IHC); Isoform / epitope: One 1–211 chain; no listed isoforms or processing (UniProt); Regulation: No stimulus-linked regulation specified (UniProt). Evidence: UniProt Q9UI95, HPA tissue IHC · Approved, HPA ICC-IF · nucleoplasm., Datasheet A02357-4, PMC10956193, PMC6090455. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/MAD2L2/immunohistochemistry-ihc-guide.