This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
CHRM2 / Muscarinic acetylcholine receptor M2 · IHC design guide
Design Immunohistochemistry for CHRM2
Plan CHRM2 IHC-P using the catalog antibody at 1:25 (datasheet: M02733). Compare the observed cytoplasmic tissue staining (HPA tissue IHC) with the expected membrane location (UniProt), and interpret staining cautiously given its very low consistency with RNA expression (HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CHRM2 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 tissue staining (HPA tissue IHC); membrane expected (UniProt) | |
| Staining pattern | Variable cytoplasmic staining in glandular and neuronal cells (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Adipose tissue |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across sections (standard IHC practice; not target-specific) | |
| Caveat | Staining has very low consistency with RNA expression (HPA tissue IHC) | |
| Regulation | Agonist-driven internalization (UniProt) | |
| Isoform / epitope | No isoforms; extracellular versus cytoplasmic epitopes matter (UniProt) |
Section 1
Recommended CHRM2 IHC & IF Protocols
The catalog antibody protocol is accompanied by one published CHRM2 IHC protocol using mouse lung paraffin sections (PMC6791200).
| Sample | Paraffin-embedded H. brain tissue; fixative not specified (datasheet M02733) |
| Fixation | Image fixative and duration unreported (datasheet M02733); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Mouse monoclonal (clone 1424CT461.78.60) anti-CHRM2, 1:25 (datasheet M02733) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CHRM2-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Ubiquitous cytoplasmic expression in all tissues at variable levels. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 retrieval at 95–98 °C for 20 min (page antigen retrieval guidance); the published excerpt does not specify retrieval (PMC6791200).
Section 2
What Is the Expected CHRM2 Staining Pattern?
CHRM2 is a seven-pass receptor at the cell and postsynaptic membranes (UniProt P08172 topology and subcellular location). In paraffin-section IHC, assess staining in the cell types HPA reports as high, including caudate neurons and cerebellar Purkinje cells (HPA: High). Interpret apparent positivity cautiously: HPA describes widespread cytoplasmic staining, but rates its IHC antibodies Approved with very low agreement between staining and RNA expression, pending external verification (HPA tissue IHC).
| Membrane-associated staining in caudate neurons or cerebellar Purkinje cells, with identifiable cells and limited surrounding stain. | This fits the UniProt cell-membrane location and HPA High staining in these cells (UniProt P08172; HPA tissue IHC). Record the membrane pattern and cellular distribution separately from overall intensity. HPA's low staining–RNA agreement means this appearance remains a provisional CHRM2 interpretation (HPA reliability description). |
| Strong, uniform cytoplasmic staining with little discernible membrane association. | HPA reports variable cytoplasmic staining across tissues, so cytoplasmic signal alone cannot be dismissed (HPA tissue IHC). It also cannot establish receptor localization: UniProt places CHRM2 at cell and postsynaptic membranes (UniProt P08172). Compare cellular boundaries and negative controls, then seek independent confirmation before calling diffuse cytoplasm specific. |
| Staining centered on adipocytes or equally intense across unrelated cells. | HPA reports CHRM2 as Not detected in adipocytes, making prominent adipocyte staining discordant with that reference (HPA tissue IHC). Widespread cell-independent color may reflect nonspecific binding or endogenous detection activity (general IHC practice). Check a no-primary control and the tissue's morphology before assigning cell-specific positivity. |
| Diffuse color over cells and extracellular areas, without a readable cellular pattern. | Treat this as background rather than a CHRM2 localization call (general IHC practice). The reported HPA profile is widespread cytoplasmic staining, but a diffuse field does not identify stained cells (HPA tissue IHC). Inspect the no-primary control and detection reagents; reassess staining only where cell boundaries and counterstain remain interpretable. |
| No convincing signal in a section containing caudate neurons or cerebellar Purkinje cells. | These are useful reference cells because HPA reports High staining in them (HPA tissue IHC), although its antibody evidence remains provisional (HPA reliability description). Review tissue preservation, assay controls, retrieval conditions and antibody use as general IHC troubleshooting. An absent signal alone cannot distinguish failed detection from a discrepancy in the HPA pattern. |
💡Expected CHRM2 appearanceCall a cautious positive when distinct, membrane-associated staining marks HPA High cells such as caudate neurons or Purkinje cells; diffuse field color or strong adipocyte staining is suspect (UniProt P08172 localization; HPA tissue IHC, reliability description).
| Membrane topology and compartment | CHRM2 has seven transmembrane segments, with extracellular and cytoplasmic regions (UniProt P08172 topology). Assess cell borders and processes where morphology permits. The supplied record gives no antibody epitope, so topology cannot predict which retrieval condition or staining side a given IHC antibody requires. |
| Receptor internalization | Agonist-associated phosphorylation promotes CHRM2 internalization; phosphorylated receptor interacts with arrestins during internalization (UniProt P08172). Intracellular puncta can therefore be biologically plausible, but this record supplies no specimen treatment history. Do not identify puncta as internalized CHRM2 from appearance alone. |
| Tissue and cell selection | HPA reports High staining in bronchial respiratory epithelium and adrenal, appendix, colon and duodenal glandular cells, as well as specified neurons (HPA tissue IHC). It reports Low staining in ovarian stroma and skeletal-muscle myocytes, and Not detected in adipocytes (HPA tissue IHC). Use these as comparison patterns, with the HPA reliability caveat. |
| Strength of tissue evidence | Two listed rabbit antibodies, HPA029795 and CAB022338, have Approved IHC status; neither is listed as Enhanced (HPA antibodies). HPA also flags very low staining–RNA consistency and pending external verification (HPA tissue IHC). An Approved label should not be treated as independent confirmation of every stained cell or compartment. |
| IF/ICC: what pattern is supported? | HPA summarizes the subcellular location as Membrane but supplies no main location or cell-line ICC-IF images (HPA subcellular). UniProt also places CHRM2 at cell and postsynaptic membranes (UniProt P08172). These records support a membrane-location expectation, but provide no image-backed IF/ICC staining pattern or IF protocol. |
| Processing and target-specific fixation evidence | UniProt lists one CHRM2 chain, residues 1–466, with no signal peptide or propeptide and no annotated isoforms (UniProt P08172). The supplied UniProt and HPA records do not report a target-specific fixation effect. They cannot justify attributing a weak or absent IHC result specifically to CHRM2 fixation sensitivity. |
| Situation | Likely cause | Next action |
|---|---|---|
| Expected HPA High cells are unstained. | The assay may have failed, or this specimen may differ from the provisional HPA pattern (HPA tissue IHC, reliability description). | Confirm morphology and run appropriate assay controls; review retrieval and antibody conditions as general IHC practice. Interpret a repeat result against an independently supported reference. |
| Color is widespread but cell borders are unreadable. | Excess background or endogenous detection activity can obscure localization (general IHC practice); HPA also describes widespread cytoplasmic staining (HPA tissue IHC). | Inspect a no-primary control, blocking and chromogen development. Score CHRM2 only where cellular distribution can be distinguished from background. |
| Adipocytes appear strongly positive. | This conflicts with HPA's Not detected adipocyte result; nonspecific staining or endogenous activity is possible (HPA tissue IHC; general IHC practice). | Verify cell identity on the counterstain and examine a no-primary control. Seek independent target confirmation before reporting adipocyte CHRM2. |
| Signal is exclusively cytoplasmic or punctate. | HPA reports cytoplasmic staining, while UniProt places CHRM2 at membranes and notes receptor internalization (HPA tissue IHC; UniProt P08172). | Document the pattern without assigning a mechanism. Compare controls and an independent detection approach before calling the cytoplasmic signal specific. |
| A listed antibody produces an unexpected pattern. | Approved IHC status coexists with very low staining–RNA consistency and pending external verification (HPA antibodies; HPA reliability description). | Check the antibody identity and assay controls. If available, compare a second independently validated antibody or orthogonal evidence before accepting the cell-type assignment. |
| An IF/ICC result disagrees with the paraffin-section IHC result. | HPA gives a Membrane summary but no ICC-IF cell-line images, so the supplied evidence cannot resolve the difference (HPA subcellular). | Assess each assay with its own controls and record its compartment pattern. Avoid transferring an IHC intensity expectation to IF/ICC without separate validation. |
Experiment evidence
IHC & IF images showing CHRM2 staining
M02733 · IHCImmunohistochemical analysis of paraffin-embedded H. brain section using CHRM2. M02733 was diluted at 1:25 dilution. A undiluted biotinylate
M02733 · IHCImmunohistochemical analysis of paraffin-embedded H. heart section using CHRM2. M02733 was diluted at 1:25 dilution. A undiluted biotinylate
M02733 · IFFluorescent image of SH-SY5Y cells stained with CHRM2 Antibody. M02733 was diluted at 1:25 dilution. An Alexa Fluor® 488-conjugated goat ant
PA1325-1 · IHCAnti-Muscarinic Acetylcholine Receptor 2 antibody, PA1325-1, IHC(P) IHC(P): Rat Brain Tissue
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000181072-CHRM2/tissue/cerebral+cortex#imid_7514912
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000181072-CHRM2/tissue/colon#imid_7514838
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000181072-CHRM2/tissue/liver#imid_7514847
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000181072-CHRM2/tissue/kidney#imid_7514832
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CHRM2 IHC & IF
🧪Run cerebellum first and require staining in Purkinje cells (HPA: High in cerebellar Purkinje cells); adjacent cells not identified as positive should show only background. Run adipose tissue as the negative tissue and assess adipocytes (HPA: Not detected in adipocytes).
Positive control tissue: Adrenal gland (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 CHRM2; 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 either a host- and isotype-matched control for a monoclonal antibody or matched nonimmune host IgG for a polyclonal antibody; use CHRM2 knockout tissue as a biological negative where available. For chromogenic detection, check endogenous peroxidase and endogenous biotin background because the selected brain-section example uses a biotinylated secondary and DAB (M02733 tissue-IHC caption).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported in the supplied evidence, and the fixative in the selected paraffin-section caption is unreported (M02733 tissue-IHC caption). Antigen-retrieval dependence is unreported, so compare retrieval conditions empirically; the supplied evidence does not establish that frozen sections or IF are easier (HPA subcellular: no ICC-IF cell-line images). In cerebellum, check for background from the biotin-based detection used in the selected brain-section example (M02733 tissue-IHC caption).
HPA tissue IHC evidence for CHRM2
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Very 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 |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Appendix | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | High | Protein (IHC) | HPA → |
| Cerebellum | Purkinje cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CHRM2 IHC Tips
Troubleshoot CHRM2 staining in paraffin sections by checking retrieval, tissue preservation, compartment, detection background, and scoring controls (UniProt P08172; standard IHC practice).
How should I optimize antigen retrieval for CHRM2 in paraffin sections?
Start CHRM2 paraffin IHC with citrate pH 6.0 HIER at 95–98 °C for 20 min (page retrieval rule: cytoplasmic / membrane antigen). Allow slides to cool in buffer, then compare a retrieved section with a matched section without retrieval to assess signal and tissue preservation (standard IHC practice). CHRM2 crosses the membrane 7 times, so exposure of a particular epitope may depend on processing (UniProt P08172 topology; standard IHC practice). If staining remains weak, compare a fallback retrieval condition on serial sections while holding antibody concentration and DAB development constant (standard IHC practice).
How can I assess fixation effects on CHRM2 paraffin staining?
Target-specific fixation sensitivity is unknown: the selected CHRM2 tissue caption reports paraffin embedding but does not state a fixative (M02733 caption). Record the actual fixative and fixation duration for each specimen, then compare sections processed together before attributing signal differences to biology (standard IHC practice). Keep section thickness, citrate pH 6.0 retrieval at 95–98 °C for 20 min, and DAB development matched across comparisons (page retrieval rule; standard IHC practice). Assess morphology alongside staining because inadequate fixation can impair preservation and prolonged fixation can reduce epitope accessibility (standard IHC practice).
Should CHRM2 appear on membranes or in the cytoplasm?
CHRM2 is annotated at cell and postsynaptic membranes, and agonist-associated phosphorylation promotes receptor internalization (UniProt P08172 subcellular location). In paraffin sections, assess membrane-associated DAB against cellular morphology and record intracellular staining separately within identified cells (UniProt P08172 location; standard IHC practice). Its cytoplasmic segment spans residues 208–387, while 7 helices traverse the membrane; processing may affect epitope access and apparent staining pattern (UniProt P08172 topology; standard IHC practice). HPA reports ubiquitous cytoplasmic staining with very low concordance to RNA expression, so independently validate diffuse cytoplasmic signal before assigning it to CHRM2 (HPA tissue IHC).
Which CHRM2 epitope features matter when staining looks inconsistent?
Obtain the antibody’s immunogen or mapped epitope: the selected tissue caption gives staining conditions but no epitope coordinates (M02733 caption). CHRM2 has 7 transmembrane spans, extracellular residues 1–20, and a cytoplasmic segment at 208–387 (UniProt P08172 topology). Glycosylation sites at residues 2, 3, 6, 9 and cytoplasmic phosphorylation sites could influence staining if the epitope overlaps them (UniProt P08172 PTMs; standard IHC practice). No alternative isoforms are annotated; investigate discordant paraffin staining with epitope mapping and a specificity control before proposing a splice variant (UniProt P08172 isoforms; standard IHC practice).
How should I check CHRM2 localisation by multiplex immunofluorescence?
The selected antibody caption documents paraffin chromogenic IHC and provides no IF fixation or permeabilisation conditions (M02733 caption). For multiplex IF, pair CHRM2 with a validated neuronal marker in caudate sections and include single-label controls for spectral bleed-through (HPA: high in caudate neuronal cells; standard IF practice). Consider a far-red fluorophore when an unstained section shows stronger autofluorescence in shorter-wavelength channels (standard IF practice). For a mapped cytoplasmic epitope, titrate permeabilisation to preserve membrane morphology; an extracellular epitope may require less permeabilisation (UniProt P08172 topology; standard IF practice). Compare compartment patterns with the separately scored chromogenic sections (standard IF practice).
How can I trace diffuse or edge-heavy CHRM2 DAB background?
The selected paraffin image used M02733 at 1:25, an undiluted biotinylated polyvalent secondary, and DAB, giving several steps to check (M02733 caption; standard IHC practice). Run no-primary and secondary-only sections, block endogenous peroxidase before DAB, and assess endogenous biotin for this biotin-based detection workflow (M02733 caption; standard IHC practice). If staining remains diffuse, titrate the primary from 1:25 and adjust DAB development while keeping citrate pH 6.0 retrieval constant (M02733 caption; page retrieval rule; standard IHC practice). Evaluate tissue edges and damaged regions separately before assigning diffuse signal to CHRM2 (standard IHC practice).
How should I score CHRM2 chromogenic staining across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring: CHRM2 is membrane-associated but can internalize after agonist response (UniProt P08172 subcellular location). Report percent positive cells and an H-score based on preset intensity categories, with thresholds established from control sections (standard IHC practice). Normalize positive counts or stained area to evaluable cell number or tissue area in mm², and report membrane-associated and cytoplasmic staining separately (standard IHC practice). Match section thickness, retrieval, imaging conditions, and DAB timing across samples; document any deviation from the caption’s 1:25 dilution (M02733 caption; standard IHC practice). Exclude folds, necrosis, and edge zones using preset criteria (standard IHC practice).
How do I distinguish true CHRM2 staining from artefact?
A plausible positive is cell-associated and compatible with membrane localisation, although intracellular redistribution can follow receptor activation (UniProt P08172 subcellular location). Reported high neuronal staining in caudate and cerebral cortex offers context, but HPA’s very low RNA concordance calls for independent validation (HPA tissue IHC). Signal in adipocytes, reported as undetected, or uniform edge-heavy DAB deserves scrutiny; necrosis and endogenous peroxidase can also mimic positives (HPA: adipocytes not detected; standard IHC practice). Compare no-primary and peroxidase-block controls, then verify suspected positives with an independent specificity assay before interpreting intensity as receptor abundance (standard IHC practice).
Boster reagents
Best CHRM2 / Muscarinic acetylcholine receptor M2 IHC Antibodies
Validated anti-CHRM2 options show IHC in human brain and heart paraffin sections and rat brain tissue, plus IF in SH-SY5Y cells (catalog image captions: M02733, PA1325-1).
Real IHC data
Anti-CHRM2 Antibody
Cat # M02733
Real IHC data
Anti-Muscarinic Acetylcholine Receptor 2/CHRM2 Antibody ®
Cat # PA1325-1
M02733 has IHC images from human brain and heart paraffin sections and an IF image from SH-SY5Y cells; its listed reactivity is human and mouse (M02733 image captions; catalog reactivity). PA1325-1 has a rat brain IHC(P) image and listed human, mouse and rat reactivity (PA1325-1 image caption; catalog reactivity).
Which to pick: For tissue IHC, choose mouse monoclonal M02733 for the documented human brain or heart paraffin-section examples, or rabbit PA1325-1 for the documented rat brain IHC(P) example (M02733 catalog host and IHC captions; PA1325-1 catalog host and IHC caption). For IF/ICC planning, M02733 lists IF and shows SH-SY5Y staining (M02733 catalog applications and IF caption). For human, mouse and rat coverage, PA1325-1 lists all three species (PA1325-1 catalog reactivity); neither SKU’s IHC caption reports the fixative (M02733 and PA1325-1 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P08172 (ACM2_HUMAN, Muscarinic acetylcholine receptor M2).
- Human Protein Atlas. CHRM2 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. CHRM2 subcellular location (ICC-IF): Membrane.
- Human Protein Atlas. CHRM2 antibody validation summary (2 antibodies).
- Muscarinic cholinergic receptor (M2) plays a crucial role in the development of myopia in mice. Disease models & mechanisms 2013 — PMC3759334.
- Effect of Tiotropium Bromide on Airway Inflammation and Programmed Cell Death 5 in a Mouse Model of Ovalbumin-Induced Allergic Asthma. Canadian respiratory journal 2019 — PMC6791200.
- Chemical genetic activation of the cholinergic basal forebrain hippocampal circuit rescues memory loss in Alzheimer's disease. Alzheimer's research & therapy 2022 — PMC9006585.
- PubMed PMID:3037705 — UniProt-cited evidence.
- PubMed PMID:3443095 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CHRM2 (Muscarinic acetylcholine receptor M2). Expected localisation Cytoplasmic tissue staining (HPA tissue IHC); membrane expected (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: M02733 at 1:25 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Adipose tissue. IHC workflow; specimen preparation unspecified. Protocol steps: 1. Sample: Paraffin-embedded H. brain tissue; fixative not specified (datasheet M02733); 2. Fixation: Image fixative and duration unreported (datasheet M02733); 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-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Mouse monoclonal (clone 1424CT461.78.60) anti-CHRM2, 1:25 (datasheet M02733); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Keep fixation consistent across sections (standard IHC practice; not target-specific); Caveat: Staining has very low consistency with RNA expression (HPA tissue IHC); Isoform / epitope: No isoforms; extracellular versus cytoplasmic epitopes matter (UniProt); Regulation: Agonist-driven internalization (UniProt). Evidence: UniProt P08172, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Datasheet M02733, PMC6791200. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CHRM2/immunohistochemistry-ihc-guide.