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CPLX1 / Complexin-1 · IHC design guide
Design Immunohistochemistry for CPLX1
Plan chromogenic CPLX1 IHC in paraffin sections using cytoplasmic staining in neural and neuroendocrine tissues and synaptic staining in brain as expected readouts (HPA tissue IHC). The guide uses the catalog antibody’s documented IHC workflow and flags the possibility of signal from more than one gene product (datasheet A05254; HPA tissue IHC).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
CPLX1 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 in neural and neuroendocrine tissues; synaptic in brain (HPA tissue IHC) | |
| Staining pattern | Neuronal and neuroendocrine cytoplasm; brain synapses (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A05254) | |
| Positive control | Adrenal gland+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 | Signal may include products of more than one gene (HPA tissue IHC) | |
| Regulation | Increased in Parkinson substantia nigra (UniProt) | |
| Isoform / epitope | No isoforms; single cytosolic chain, no ectodomain (UniProt) |
Section 1
Recommended CPLX1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol (datasheet A05254) is followed by two published CPLX1 protocols for chromogenic staining of paraffin sections (PMC6944231; PMC12283636).
| Sample | Paraffin-embedded rat brain tissue; fixative not specified (datasheet A05254) |
| Fixation | Image fixative and duration unreported (datasheet A05254); 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 A05254); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A05254) |
| Primary antibody | Rabbit anti-CPLX1, 1:50 (datasheet A05254) |
| Primary incubation | Overnight at 4 °C (datasheet A05254) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A05254) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | CPLX1-positive staining in medullary cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in nervous and neuroendocrine tissues. Synaptic positivity in brain. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A05254). The published protocols report citrate pH 6, and one also reports Tris-EDTA pH 9 (PMC6944231; PMC12283636).
Section 2
What Is the Expected CPLX1 Staining Pattern?
CPLX1 is a cytosolic protein enriched at presynaptic release sites in mature neurons; it has no transmembrane segment (UniProt O14810). In paraffin IHC, expect cytoplasmic staining in neuronal and neuroendocrine cells, with synaptic positivity in brain (HPA tissue IHC). HPA rates the tissue pattern Supported, with medium agreement between staining and RNA data, and cautions that the profile includes antibodies targeting proteins from more than one gene (HPA tissue IHC).
| Cytoplasmic staining in cerebral cortical neurons and cerebellar granular-layer cells, with staining in synaptic regions. | This fits HPA's High staining in both cell groups and its reported synaptic positivity in brain (HPA tissue IHC). Presynaptic enrichment provides a localization rationale, but an individual positive cell cannot be identified as inhibitory from staining alone (UniProt O14810). |
| Cytoplasmic staining in adrenal medullary cells or pancreatic endocrine cells. | These are useful comparisons outside brain: HPA reports High staining in adrenal medullary cells and Medium staining in pancreatic endocrine cells (HPA tissue IHC). Judge intensity within the identified cell population; other cells on the section need not match it. |
| Strong, predominantly nuclear staining in cells expected to be positive. | Nuclear dominance conflicts with the reported cytosolic and presynaptic localization (UniProt O14810; HPA tissue IHC). Treat it as a localization warning, then compare the same compartment in a known-positive tissue and a primary-antibody omission control before interpreting it as CPLX1. |
| Prominent staining in an HPA-listed negative cell population, such as adipocytes or bronchial respiratory epithelium. | HPA reports CPLX1 as Not detected in those cell populations (HPA tissue IHC). Unexpected staining warrants checks for background, endogenous detection activity or antibody cross-reactivity; the multi-gene antibody caution makes target attribution particularly uncertain (HPA tissue IHC). |
| Diffuse stain across the section, or no signal in cerebral cortical neurons. | Neither result gives a convincing cell-restricted positive pattern: HPA reports High cortical neuronal staining (HPA tissue IHC). Use an omission control to assess diffuse signal, and check section and detection controls before treating absent cortical staining as biological absence (general IHC practice). |
💡Expected CPLX1 appearanceA convincing positive shows cytoplasmic neuronal staining with synaptic positivity in brain, including High cortical neuronal staining; broad staining of HPA-negative cells or dominant nuclear staining calls for control-based review (HPA tissue IHC; UniProt O14810).
| Cell and tissue selection | HPA reports High staining in cortical neurons, cerebellar granular-layer cells and adrenal medullary cells; hippocampal neurons and pancreatic endocrine cells are Medium (HPA tissue IHC). Use those distinctions when choosing a comparison section and judging intensity. |
| Compartment and protein topology | CPLX1 is cytosolic, occurs in the perikaryon and presynapse, and lacks a transmembrane segment (UniProt O14810). Score cytoplasmic and synaptic staining separately; a crisp cell-surface rim is not the localization predicted by this record. |
| Strength of target attribution | HPA calls its tissue IHC reliability Supported but notes medium agreement with RNA and antibodies targeting proteins from more than one gene (HPA tissue IHC). The reported pattern supports interpretation, yet an unexpected positive population needs independent validation. |
| IF/ICC: What pattern can be used for comparison? | HPA describes mainly vesicular localization, marked uncertain, from antibodies targeting proteins from multiple genes (HPA subcellular ICC-IF). Treat it as a cautious comparison only; the IHC tissue profile reports cytoplasmic and synaptic staining (HPA tissue IHC). |
| Situation | Likely cause | Next action |
|---|---|---|
| No stain in a cerebral cortex positive-control section | Cortical neurons are reported High, so a blank result may reflect the section, staining workflow or detection system rather than expected tissue distribution (HPA tissue IHC; general IHC practice). | Confirm neuronal morphology and section integrity; check primary incubation, antigen retrieval and chromogen controls using the validated IHC workflow (general IHC practice). Target-specific fixation sensitivity is unreported in the supplied sources. |
| Diffuse chromogen across tissue and empty spaces | A field-wide deposit lacks the cell restriction in HPA's neuronal and neuroendocrine profile; nonspecific detection or excess background is possible (HPA tissue IHC; general IHC practice). | Compare a primary-antibody omission control, then review blocking, wash steps and detection reagent exposure (general IHC practice). Do not score diffuse deposit as CPLX1 solely because a positive cell group is present. |
| Strong signal in adipocytes or bronchial respiratory epithelium | HPA lists these cell populations as Not detected, and flags a multi-gene antibody limitation for its tissue profile (HPA tissue IHC). The stain therefore needs a specificity check. | Compare a known-positive section and omission control in the same run; review whether signal follows cells, edges or deposits (general IHC practice). Report the unexpected population separately rather than folding it into the expected CPLX1 pattern. |
| Nuclear staining dominates while neuronal cytoplasm is faint | That distribution differs from the cytosolic, perikaryal and presynaptic locations recorded for CPLX1 (UniProt O14810). A nuclear pattern alone is insufficient evidence of the expected IHC result. | Recheck compartment boundaries against the counterstain and compare with a cortical neuronal positive control (HPA tissue IHC; general IHC practice). If nuclear dominance persists, flag localization as discordant and seek independent target validation. |
| A pancreatic section appears weaker than an adrenal section | HPA reports Medium staining in pancreatic endocrine cells and High staining in adrenal medullary cells (HPA tissue IHC). A weaker pancreatic signal can therefore fit the observed tissue profile. | Identify the endocrine or medullary cells before comparing intensity, and score each cell population within its own section (HPA tissue IHC; general IHC practice). Avoid calling the pancreatic section failed solely because it is lighter. |
| Cerebellar staining is present, but its cell identity is unclear | HPA's High cerebellar call concerns cells in the granular layer; UniProt says CPLX1 is expressed mainly by inhibitory neurons in cerebellum (HPA tissue IHC; UniProt O14810). These statements do not identify every stained cell. | Use anatomy and counterstain to record the stained layer and compartment (general IHC practice). Describe the observed cells without assigning inhibitory identity from CPLX1 staining alone; investigate a different distribution with appropriate controls. |
Experiment evidence
IHC & IF images showing CPLX1 staining
A05254 · IHCIHC analysis of CPLX1 using anti-CPLX1 antibody (A05254). CPLX1 was detected in a paraffin-embedded section of rat brain tissue. Heat mediat
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000168993-CPLX1/tissue/cerebral+cortex#imid_20693944
HPA · Adrenal glandHuman Protein Atlas IHC, Adrenal gland. Source: https://www.proteinatlas.org/ENSG00000168993-CPLX1/tissue/adrenal+gland#imid_1266
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000168993-CPLX1/tissue/colon#imid_20693862
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000168993-CPLX1/tissue/liver#imid_20693871
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA071191; green = CPLX1, blue = nucleus, red = microtubules.
HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA071191; green = CPLX1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for CPLX1 IHC & IF
🧪Run cerebral cortex first: neuronal cells should stain strongly (HPA: High in cerebral cortex neuronal cells). Use adipose tissue as the negative tissue (HPA: Not detected in adipocytes); on the cortex slide, assess neighboring non-neuronal cells as candidate internal background controls rather than assuming they are CPLX1-negative (HPA: staining assigned to neuronal cells).
Positive control tissue: Adrenal gland (Medullary cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CPLX1 in A-549, Hep-G2, U2OS, with annotated localisation: Vesicles (uncertain) (HPA subcellular).
Technical controls: Include no-primary (secondary-only), a concentration-matched rabbit IgG isotype control with clonality matched if known, and CPLX1-knockout tissue as a biological negative (caption: rabbit primary; standard IHC practice). Block endogenous peroxidase for chromogenic detection in the positive tissue (caption: HRP/DAB detection; standard IHC practice).
⚠️Feasibility: No target-specific fixation window or fixation effect is reported, and the selected A05254 paraffin-section caption does not state a fixative (caption: fixative unreported). The demonstrated IHC procedure uses heat retrieval in EDTA at pH 8.0; retrieval dependence has not been established by a comparison without retrieval (caption: EDTA retrieval). The supplied evidence does not establish that frozen sections or IF are easier; brain autofluorescence should be checked separately if IF is pursued (HPA: High in cerebral cortex neuronal cells; standard IF practice).
HPA tissue IHC evidence for CPLX1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Medium consistency between antibody staining and RNA expression data. Caution, targets protein from more than one gene.). 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 | Medullary cells | High | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
| Caudate | Neuronal cells | Medium | Protein (IHC) | HPA → |
| Hippocampus | Neuronal cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Appendix | Endocrine cells | Not detected | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Not detected | Protein (IHC) | HPA → |
| Breast | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced CPLX1 IHC Tips
Troubleshoot CPLX1 staining in paraffin sections by checking retrieval, expected cellular patterns, controls, and scoring before interpreting chromogenic signal.
What retrieval should I try first for weak CPLX1 staining in paraffin sections?
Start with heat-mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A05254). The catalog antibody produced CPLX1 staining in rat brain after this retrieval, followed by 1:200 primary antibody overnight at 4°C (datasheet A05254). If signal is weak, check that sections were fully deparaffinised and compare retrieval duration on adjacent sections while keeping the antibody dilution and DAB development constant (standard IHC practice). Use the same tissue region and include a no-primary control, because stronger DAB alone cannot establish that a retrieval change improved specific staining (standard IHC practice).
Could fixation explain weak or uneven CPLX1 staining?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state the fixative (datasheet A05254). Record the fixative, fixation duration, section thickness, and storage history for each specimen so a weak slide can be compared with a consistently processed positive section (standard IHC practice). Assess fixation and retrieval changes on adjacent sections while holding the 1:200 antibody dilution and detection steps constant (datasheet A05254; standard IHC practice). Uneven staining near tissue edges can reflect processing or reagent access, so compare well-preserved interior regions before treating the pattern as biological variation (standard IHC practice).
Where should convincing CPLX1 chromogenic staining appear?
Look for cytoplasmic neuronal staining and synaptic signal in brain; CPLX1 is reported in the cytosol, perikaryon, and presynapse (UniProt O14810 localisation; HPA: synaptic positivity in brain). In mature neurons, enrichment at synaptic release sites supports a regional neuropil pattern alongside cellular staining (UniProt O14810 localisation). Cerebral cortical neuronal cells and cerebellar granular-layer cells are listed as strongly positive, while hippocampal neuronal cells have medium staining (HPA tissue IHC). Compare cellular and neuropil compartments separately, because diffuse nuclear-only DAB or staining restricted to damaged edges does not match the reported localisation (UniProt O14810 localisation; standard IHC practice).
How should an unknown antibody epitope affect CPLX1 troubleshooting?
The supplied record describes a 134-amino-acid CPLX1 chain and annotates no isoforms, glycosylation sites, or modified residues (UniProt O14810 record). It gives no antibody epitope, so those annotations cannot establish which sequence the catalog antibody recognises or how fixation affects access (UniProt O14810 record; datasheet A05254). Keep retrieval and the 1:200 dilution matched when comparing tissues, and seek the antibody's immunogen or epitope documentation before attributing discordant staining to an isoform (datasheet A05254; standard IHC practice). An independent antibody recognising a documented different epitope can help investigate specificity if discordance persists (standard IHC practice).
How can I investigate CPLX1 localisation with a separate IF experiment?
For IF, multiplex CPLX1 with a marker of the expected neuronal cell population and inspect cellular versus synaptic regions in separate channels (UniProt O14810 localisation; standard IF practice). Choose fluorophores after checking tissue autofluorescence in an unstained section, and place the weaker signal in a channel with lower background (standard IF practice). CPLX1 is cytosolic and has no transmembrane segment, so permeabilise sufficiently to let antibody reach intracellular antigen while preserving fine processes (UniProt O14810 topology and localisation; standard IF practice). Optimise fixation, permeabilisation, and antibody concentration for IF separately; the 1:200 overnight condition comes from paraffin-section chromogenic IHC (datasheet A05254).
How can I reduce diffuse DAB background without losing synaptic signal?
Start by comparing the stained section with a no-primary control and reviewing whether DAB development was stopped before background obscured fine structures (standard IHC practice). The catalog paraffin-section example used 10% goat serum, a peroxidase-conjugated secondary antibody, and DAB detection (datasheet A05254). Check that blocking, washes, and an endogenous peroxidase block are effective before lowering the 1:200 primary antibody concentration (datasheet A05254; standard IHC practice). If background persists, compare shorter primary incubation or a dilution series on adjacent sections against expected neuronal cytoplasmic and synaptic staining (UniProt O14810 localisation; standard IHC practice).
What is a defensible way to quantify CPLX1 IHC across sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define comparable anatomical regions before scoring and use identical imaging, counterstain, and DAB development settings across sections (standard IHC practice). An H-score can summarize the percentage of cells at each staining intensity, while positive-cell percentage measures cellular staining and positive-area density per mm² can capture neuropil signal (standard IHC practice). Report cellular and synaptic compartments separately because both are plausible CPLX1 locations in brain (UniProt O14810 localisation; HPA: synaptic positivity in brain). Normalise cell-based scores to the number of eligible cells and area-based measures to sampled tissue area, excluding folds, edges, and damaged regions by prespecified rules (standard IHC practice).
How do I distinguish true CPLX1 positivity from chromogenic artefact?
A credible result should fit CPLX1's cytoplasmic, perikaryal, or presynaptic localisation and the reported neuronal and synaptic brain pattern (UniProt O14810 localisation; HPA tissue IHC). Strong nuclear-only staining, signal confined to cut edges or necrotic tissue, and staining that remains in a no-primary control warrant investigation before interpretation (standard IHC practice). Check endogenous enzyme activity and DAB deposition with appropriate controls, then compare the same compartment in a well-preserved region of the section (standard IHC practice). Treat tissue-pattern agreement as supporting evidence rather than proof of antibody specificity, since the HPA tissue profile carries a multiple-gene antibody caution (HPA tissue IHC reliability description).
Boster reagents
Best CPLX1 / Complexin-1 IHC Antibodies
A05254 has real IHC data from paraffin-embedded rat brain (image caption); its catalog lists IF/ICC applications and Human, Mouse, and Rat reactivity (catalog: applications and reactivity).
Real IHC data
Anti-CPLX1 Antibody
Cat # A05254
A05254 is the only SKU shown and has an IHC image from a paraffin-embedded rat brain section (image caption). IF/ICC are listed applications, but no IF image is supplied (catalog: applications and IF image alts).
Which to pick: For tissue IHC, choose A05254: its rat brain image documents heat retrieval in EDTA at pH 8.0 and primary antibody at 1:200 overnight at 4°C (A05254 image caption); the fixative is unreported (A05254 image caption). For IF/ICC or work across Human, Mouse, and Rat, A05254 is the listed option because it is polyclonal and includes those applications and species (catalog: dilution_raw, applications, reactivity); the supplied figure documents rat brain IHC only (A05254 image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O14810 (CPLX1_HUMAN, Complexin-1).
- Human Protein Atlas. CPLX1 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. CPLX1 subcellular location (ICC-IF): Mainly localized to vesicles. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. CPLX1 antibody validation summary (2 antibodies).
- The Proteome of the Dentate Terminal Zone of the Perforant Path Indicates Presynaptic Impairment in Alzheimer Disease. Molecular & cellular proteomics : MCP 2020 — PMC6944231.
- CPLX1 is a novel prognostic biomarker in CRC correlating with immunotherapy resistance and ferroptosis. Frontiers in immunology 2025 — PMC12283636.
- The schizophrenia risk gene product miR-137 alters presynaptic plasticity. Nature neuroscience 2015 — PMC4506960.
- PubMed PMID:7553862 — UniProt-cited evidence.
- PubMed PMID:24722188 — UniProt-cited evidence.
- PubMed PMID:21697133 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for CPLX1 (Complexin-1). Expected localisation Cytoplasmic in neural and neuroendocrine tissues; synaptic in brain (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A05254). Validated IHC antibody: A05254 at 1:50 (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Medullary cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded rat brain tissue; fixative not specified (datasheet A05254); 2. Fixation: Image fixative and duration unreported (datasheet A05254); 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 A05254); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A05254); 8. Primary antibody: Rabbit anti-CPLX1, 1:50 (datasheet A05254); 9. Primary incubation: Overnight at 4 °C (datasheet A05254). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Signal may include products of more than one gene (HPA tissue IHC); Isoform / epitope: No isoforms; single cytosolic chain, no ectodomain (UniProt); Regulation: Increased in Parkinson substantia nigra (UniProt). Evidence: UniProt O14810, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to vesicles. Caution: Based on antibodies targeting…, Datasheet A05254, PMC6944231, PMC12283636. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/CPLX1/immunohistochemistry-ihc-guide.