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MTR / Methionine synthase · IHC design guide
Design Immunohistochemistry for MTR
Plan chromogenic IHC on paraffin sections with the IHC-validated antibody at 0.5–1 μg/mL (datasheet: A01140-2). Compare high-staining breast glandular cells with adipocytes reported as undetected, and interpret cytoplasmic and membranous staining in light of the reported antibody–RNA inconsistency (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
MTR 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 | IHC: cytoplasmic and membranous; molecular location: cytoplasm (HPA tissue IHC; UniProt) | |
| Staining pattern | Cytoplasmic and membranous staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6 HIER, heat-mediated (datasheet A01140-2) | |
| Positive control | Breast+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 conditions consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Antibody staining has low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | No specific expression regulator is reported (UniProt) | |
| Isoform / epitope | 2 isoforms; confirm antibody epitope coverage (UniProt) |
Section 1
Recommended MTR IHC & IF Protocols
The catalog antibody's IHC-P protocol is accompanied by two published MTR IHC protocols (PMC9208444; PMC5726624).
| Sample | Paraffin-embedded human placenta tissues; fixative not specified (datasheet A01140-2) |
| Fixation | Image fixative and duration unreported (datasheet A01140-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: Citrate pH 6, 20 min (datasheet A01140-2) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A01140-2) |
| Primary antibody | Rabbit anti-MTR, 0.5-1μg/ml (datasheet A01140-2) |
| Primary incubation | Overnight at 4 °C (datasheet A01140-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A01140-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | MTR-positive staining in glandular cells of breast (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic and membranous expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated citrate retrieval at pH 6 (datasheet A01140-2); both published protocols also boil sections in citrate buffer (PMC9208444; PMC5726624).
Section 2
What Is the Expected MTR Staining Pattern?
MTR is a cytoplasmic protein without a transmembrane segment (UniProt Q99707 localization and topology). In tissue IHC, expect cytoplasmic and sometimes membranous staining, including strong signal in breast and thyroid glandular cells, skeletal myocytes, and ciliated cells (HPA tissue IHC). HPA rates the tissue pattern Approved but reports low agreement with RNA expression and pending external verification (HPA tissue IHC reliability).
| Clear cytoplasmic staining in breast or thyroid glandular cells, or skeletal myocytes (HPA tissue IHC). | This fits the observed high staining in those cells (HPA tissue IHC) and MTR’s cytoplasmic localization (UniProt Q99707). Judge intensity against cells on the same section and a separately processed control (standard IHC practice). |
| Apical staining confined to cilia in bronchus, fallopian tube, or nasopharynx (HPA tissue IHC). | HPA reports high signal in ciliated cell axonemes (HPA tissue IHC). This pattern can coexist with cytoplasmic or membranous tissue staining (HPA tissue IHC); do not require every positive cell to look uniformly cytoplasmic. |
| Predominantly nuclear staining, with little expected cytoplasmic signal (UniProt Q99707 localization; HPA tissue IHC). | A nuclear dominant pattern does not match the supplied localization evidence (UniProt Q99707; HPA tissue IHC). Treat it as unconfirmed, then compare the positive control and detection controls before attributing it to MTR (standard IHC practice). |
| Strong staining in adipocytes or prostate glandular cells, especially when the expected positive control is weak (HPA tissue IHC). | HPA reports MTR as not detected in those cell types (HPA tissue IHC). Investigate antibody cross-reactivity or chromogenic detection background with controls; an unexpected positive alone cannot establish either cause (standard IHC practice). |
| Diffuse color across cells and tissue spaces, or no signal in a known positive control (standard IHC practice). | Diffuse color obscures compartment and cell type, while absent control staining leaves a negative specimen uninterpretable (standard IHC practice). Check the detection-only control, tissue preservation, retrieval, and antibody conditions before scoring (standard IHC practice). |
💡Expected MTR appearanceA convincing positive shows distinct cytoplasmic, sometimes membranous, staining in HPA high cell types such as breast glandular cells or skeletal myocytes; widespread nuclear color or diffuse background is suspect (HPA tissue IHC; UniProt Q99707 localization; standard IHC practice).
| Tissue and cell choice | Use breast or thyroid glandular cells and skeletal myocytes as high staining references (HPA tissue IHC). Adipocytes are reported as not detected, but HPA’s Approved profile has low RNA agreement and awaits external verification (HPA tissue IHC reliability). |
| Compartment and cellular structure | UniProt places MTR in the cytoplasm and lists no transmembrane segment (UniProt Q99707). HPA nevertheless describes cytoplasmic and membranous tissue staining and high ciliary axoneme signal in selected epithelia (HPA tissue IHC); score these observations with that source difference visible. |
| Antibody evidence | The supplied antibody HPA054915 has Approved IHC and ICC status (HPA antibodies). The supplied record does not give an Enhanced IHC designation or an epitope, so neither independent pattern reproduction nor epitope specific retrieval behavior is established here (HPA antibodies). |
| IF/ICC Q: What localization should an IF result be compared with? | A: HPA reports approved microtubule and primary cilium localization, with additional cytosol, basal body, and cytokinetic bridge staining (HPA subcellular ICC-IF). These ICC observations give context for IF interpretation; they do not specify an IHC-P retrieval or dilution setting (HPA subcellular ICC-IF). |
| Expression discordance | UniProt lists heart among the highest expression tissues, while HPA reports cardiomyocytes as not detected by tissue IHC (UniProt Q99707 tissue specificity; HPA tissue IHC). Avoid using heart muscle as the sole positive control, and record the source disagreement when interpreting it. |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in breast glandular cells or skeletal myocytes (HPA tissue IHC). | A failed control can reflect tissue processing, retrieval, antibody application, or detection problems (standard IHC practice); the supplied sources do not identify an MTR specific fixation sensitivity. | Confirm that the positive control contains the specified cells, then review retrieval and antibody conditions and verify detection reagents with a suitable control (standard IHC practice). |
| Cardiomyocytes are negative despite an expectation from expression data (UniProt Q99707; HPA tissue IHC). | UniProt reports high heart expression, but HPA reports no detected cardiomyocyte staining and flags low staining versus RNA agreement (UniProt Q99707 tissue specificity; HPA tissue IHC reliability). | Interpret the heart result against a separate HPA high cell type, such as skeletal myocytes, and retain the expression discrepancy in the report (HPA tissue IHC; standard IHC practice). |
| Color is diffuse or appears in tissue spaces (standard IHC practice). | Background from detection chemistry or nonspecific binding may obscure the cellular pattern (standard IHC practice); diffuse color alone does not establish MTR localization. | Inspect a detection-only control, review blocking and washes, and reassess whether the remaining signal resolves to the expected cells and compartments (standard IHC practice; HPA tissue IHC). |
| Unexpected nuclear dominant staining appears (UniProt Q99707 localization; HPA tissue IHC). | The supplied sources place MTR in cytoplasm or describe cytoplasmic and membranous tissue staining, so a nuclear dominant result is unsupported (UniProt Q99707; HPA tissue IHC). | Compare it with a known positive tissue and detection controls; withhold a positive MTR call if the expected cellular pattern cannot be resolved (HPA tissue IHC; standard IHC practice). |
| Strong color appears in adipocytes or prostate glandular cells (HPA tissue IHC). | Those cells are reported as not detected (HPA tissue IHC); cross-reactivity or endogenous chromogenic activity are possibilities that need controls (standard IHC practice). | Run an antibody omission control and review the detection system’s endogenous activity block; compare with a positive cell type before assigning specificity (standard IHC practice; HPA tissue IHC). |
| Ciliated epithelium shows a narrow apical signal without broad cytoplasmic staining (HPA tissue IHC). | HPA reports high staining in cilia axonemes of bronchus, fallopian tube, and nasopharynx, while its overall tissue profile also includes cytoplasmic and membranous staining (HPA tissue IHC). | Identify the stained cell type and apical structure before scoring; compare adjacent nonciliated cells and controls rather than rejecting an axonemal pattern solely for its narrow distribution (HPA tissue IHC; standard IHC practice). |
Experiment evidence
IHC & IF images showing MTR staining
A01140-2 · IHCIHC analysis of MTR using anti-MTR antibody (A01140-2). MTR was detected in paraffin-embedded section of human placenta tissues. Heat mediat
A01140-2 · IHCIHC analysis of MTR using anti-MTR antibody (A01140-2). MTR was detected in paraffin-embedded section of mouse brain tissues. Heat mediated
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000116984-MTR/tissue/cerebral+cortex#imid_20083533
HPA · Parathyroid glandHuman Protein Atlas IHC, Parathyroid gland. Source: https://www.proteinatlas.org/ENSG00000116984-MTR/tissue/parathyroid+gland#imid_20083424
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000116984-MTR/tissue/colon#imid_20083451
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000116984-MTR/tissue/liver#imid_20083468
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA054915; green = MTR, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA054915; green = MTR, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for MTR IHC & IF
🧪Run breast first: glandular cells should stain (HPA: High in breast glandular cells). Use prostate glandular cells as the negative tissue (HPA: Not detected in prostate glandular cells); on the breast slide, assess morphologically identified nonglandular cells as internal negatives only where they show no signal, since their MTR staining level is not specified by the supplied HPA row.
Positive control tissue: Breast (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show MTR in A-431, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), NIH 3T3, with annotated localisation: Microtubules (approved), Primary cilium (approved) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), a concentration-matched rabbit IgG control matched to the primary antibody’s format, and MTR-knockout material or a validated peptide-block control (caption: rabbit anti-MTR antibody; standard IHC controls). For DAB detection on breast sections, block endogenous peroxidase and check for endogenous biotin because the caption uses biotin-based detection (caption: biotinylated secondary, SABC and DAB).
⚠️Feasibility: The selected-SKU paraffin-section caption reports heat retrieval in citrate buffer at pH 6 for 20 minutes, but does not establish that retrieval is required (caption: A01140-2). Its fixative is unreported, and no target-specific fixation window or fixation effect is supplied (caption: fixative not stated). The supplied evidence does not establish whether frozen sections or IF are easier; for breast tissue, use the peroxidase and biotin controls to assess detection artefacts (caption: biotin-based DAB detection; standard IHC practice).
HPA tissue IHC evidence for MTR
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — 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 | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Fallopian tube | Ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Nasopharynx | Ciliated cells (cilia axoneme) | High | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Liver | Cholangiocytes | Not detected | Protein (IHC) | HPA → |
| Lymph node | Germinal center cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced MTR IHC Tips
Troubleshoot MTR staining in paraffin sections by checking retrieval, cellular location, background and scoring against the selected antibody image and independent expression evidence.
What retrieval should I try first if MTR staining is weak in paraffin sections?
Use heat-mediated citrate pH 6 antigen retrieval for 20 minutes in paraffin sections (datasheet A01140-2; selected tissue-IHC caption A01140-2). The selected human placenta image used that retrieval before overnight primary-antibody incubation at 4°C, so it is the closest documented starting point for this antibody (selected tissue-IHC caption A01140-2). If staining remains weak, compare a modestly longer heating interval or EDTA pH 9 on adjacent sections, watching for tissue damage or increased background (standard IHC practice). Keep section thickness, detection chemistry and development time matched while evaluating the retrieval change (standard IHC practice).
Could fixation be hiding MTR in my tissue sections?
Target-specific fixation sensitivity is unknown: the selected paraffin-section image reports retrieval and staining, but does not state a fixative or fixation duration (selected tissue-IHC caption A01140-2). Record the fixative, fixation interval and processing history for each specimen, and compare sections processed together before attributing intensity differences to MTR abundance (standard IHC practice). For routinely formalin-fixed material, keep fixation consistent and begin with citrate pH 6 retrieval for 20 minutes, the documented retrieval for this antibody (selected tissue-IHC caption A01140-2). If archival blocks stain unevenly, test matched control sections across runs and report fixation as a limitation rather than assigning MTR a specific fixation effect (standard IHC practice).
Should MTR appear cytoplasmic, membranous or ciliary in chromogenic IHC?
Expect a cytoplasmic component because MTR is annotated in the cytoplasm and has no transmembrane segment (UniProt Q99707 subcellular location and topology). Tissue IHC also reports cytoplasmic and membranous staining in most tissues, while the separate subcellular IF assessment places MTR mainly at microtubules and the primary cilium (HPA tissue IHC; HPA subcellular). A membrane-like DAB rim alone therefore needs closer inspection rather than automatic assignment to a membrane-spanning MTR pool (UniProt Q99707 topology; standard IHC practice). Compare the cellular pattern with adjacent controls and the expected cell population at matched magnification, especially where cilia may appear as thin apical structures (HPA tissue IHC; standard IHC practice).
Can this antibody distinguish MTR isoforms or a modified epitope in IHC?
MTR has 2 annotated isoforms, but the supplied antibody image does not identify its binding epitope or establish isoform selectivity (UniProt Q99707 isoforms; selected tissue-IHC caption A01140-2). The protein contains homocysteine-binding, pterin-binding, cobalamin-binding and AdoMet activation regions, so an epitope assignment would matter when interpreting altered staining (UniProt Q99707 domains). A phosphothreonine is annotated at residue 1264, but the evidence does not show whether this antibody detects that modification differently (UniProt Q99707 modified residues; selected tissue-IHC caption A01140-2). Obtain epitope mapping or isoform-specific validation before calling a staining difference isoform loss or a phosphorylation change; score the observed MTR signal without that attribution (standard IHC practice).
How should I check MTR localisation by multiplex IF alongside this IHC result?
For a cell-type check, pair MTR with an epithelial marker such as cytokeratin in glandular tissue; glandular cells show high tissue-IHC signal in breast and thyroid (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and consider a far-red MTR channel when tissue autofluorescence is strong in shorter-wavelength channels (standard IF practice). MTR has no transmembrane segment and is annotated in the cytoplasm, so use controlled permeabilisation when probing an intracellular epitope, while checking that it preserves ciliary structures (UniProt Q99707 topology and subcellular location; standard IF practice). The subcellular IF record reports microtubule and primary-cilium localisation, but the selected IHC caption supplies no IF dilution or fixation condition to transfer (HPA subcellular; selected tissue-IHC caption A01140-2).
How can I separate MTR signal from DAB and detection background?
The selected image used a 10% goat-serum block, a biotinylated secondary antibody, streptavidin-biotin detection and DAB, giving several steps to examine when background rises (selected tissue-IHC caption A01140-2). Include a no-primary section and inspect whether colour follows tissue edges, vessels or damaged regions rather than individual cells (standard IHC practice). Block endogenous peroxidase for DAB work and assess endogenous biotin where a biotin-based detection system is used; these are general workflow controls, not evidence of an MTR-specific problem (standard IHC practice). Compare matched sections after changing only one factor, such as block, primary-antibody concentration or DAB development time, so reduced background can be distinguished from lost signal (standard IHC practice).
What is a defensible way to score MTR staining across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and region before scoring, then record the percentage of positive cells and staining intensity within that population (standard IHC practice). An H-score can combine percentages at intensity grades 0–3 into a 0–300 range, provided the same thresholds and imaging conditions are used for every section (standard IHC practice). Normalise positive-cell counts to all eligible cells of the same type, or report positive-cell density per mm² of viable tissue when cell segmentation is unreliable (standard IHC practice). Score cytoplasmic signal separately from apical or membrane-like signal, and document the compartment rule because tissue IHC reports both patterns for MTR (UniProt Q99707 subcellular location; HPA tissue IHC).
When is an apparent MTR-positive cell likely to be artefactual?
Give greatest weight to reproducible cellular staining in an expected compartment: MTR is cytoplasmic by annotation, while tissue IHC also describes membrane-like staining (UniProt Q99707 subcellular location; HPA tissue IHC). Question isolated edge staining, necrotic areas or DAB colour persisting in a no-primary control, since those patterns can arise from section or detection artefacts (standard IHC practice). Cell identity matters: skeletal-muscle myocytes are listed as high, whereas heart cardiomyocytes are listed as not detected despite reported high heart expression at the tissue level (HPA tissue IHC; UniProt Q99707 tissue specificity). Because the HPA tissue assessment notes low consistency with RNA and pending external verification, resolve surprising positives with matched controls and independent target evidence before drawing biological conclusions (HPA tissue IHC).
Boster reagents
Best MTR / Methionine synthase IHC Antibodies
The IHC-validated antibody has images of human placenta and mouse and rat brain paraffin sections (IHC captions: A01140-2). No IF/ICC data are supplied (catalog: A01140-2).
Real IHC data
Anti-MTR Antibody ®
Cat # A01140-2
A01140-2 is listed for IHC in human, mouse and rat (catalog: A01140-2 applications and reactivity). Its IHC captions show human placenta and mouse and rat brain paraffin sections (IHC captions: A01140-2).
Which to pick: Choose A01140-2 for paraffin-section IHC; its captions document citrate retrieval at pH 6 for 20 minutes and 1 μg/ml primary antibody overnight at 4°C, but do not report the fixative (IHC captions: A01140-2). No listed SKU has IF/ICC validation or an IF dilution (catalog: A01140-2 applications and dilutions). For cross-species IHC, A01140-2 lists human, mouse and rat reactivity and has IHC images for all three; its host is rabbit and its clonality is unreported (catalog: A01140-2 reactivity, host and clone; IHC captions: A01140-2).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q99707 (METH_HUMAN, Methionine synthase).
- Human Protein Atlas. MTR tissue IHC expression (reliability: Approved).
- Human Protein Atlas. MTR subcellular location (ICC-IF): Mainly localized to the microtubules and primary cilium. In addition localized to the cytosol, cytokinetic bridge and basal body..
- Human Protein Atlas. MTR antibody validation summary (1 antibodies).
- Increased homocysteine regulated by androgen activates autophagy by suppressing the mammalian target of rapamycin pathway in the granulosa cells of polycystic ovary syndrome mice. Bioengineered 2022 — PMC9208444.
- Attenuated expression of MTR in both prenatally androgenized mice and women with the hyperandrogenic phenotype of PCOS. PloS one 2017 — PMC5726624.
- Modulation of mTOR Within Retinal Pigment Epithelium Affects Cell Viability and Mitochondrial Pathology. International journal of molecular sciences 2025 — PMC12524646.
- Magnetization transfer imaging of ovarian cancer: initial experiences of correlation with tissue cellularity and changes following neoadjuvant chemotherapy. BJR open 2022 — PMC9459873.
- PubMed PMID:8968737 — UniProt-cited evidence.
- PubMed PMID:8968735 — UniProt-cited evidence.
- PubMed PMID:9013615 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for MTR (Methionine synthase). Expected localisation IHC: cytoplasmic and membranous; molecular location: cytoplasm (HPA tissue IHC; UniProt). Antigen retrieval: Citrate pH 6 HIER, heat-mediated (datasheet A01140-2). Validated IHC antibody: A01140-2 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Breast (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human placenta tissues; fixative not specified (datasheet A01140-2); 2. Fixation: Image fixative and duration unreported (datasheet A01140-2); 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: Citrate pH 6 , 20 min (datasheet A01140-2); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A01140-2); 8. Primary antibody: Rabbit anti-MTR, 0.5-1μg/ml (datasheet A01140-2); 9. Primary incubation: Overnight at 4 °C (datasheet A01140-2). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: 2 isoforms; confirm antibody epitope coverage (UniProt); Regulation: No specific expression regulator is reported (UniProt). Evidence: UniProt Q99707, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the microtubules and primary cilium. In addition…, Datasheet A01140-2, PMC9208444, PMC5726624. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/MTR/immunohistochemistry-ihc-guide.