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GCLC / Glutamate--cysteine ligase catalytic subunit · IHC design guide
Design Immunohistochemistry for GCLC
Plan paraffin-section GCLC IHC around variable cytoplasmic staining, with high signal reported in fallopian-tube glandular cells and bladder urothelial cells (HPA tissue IHC). Start with the IHC-validated antibody at 2–5 μg/ml (datasheet A01722).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
GCLC 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 | Variable cytoplasmic staining (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining varies across tissue cell types (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A01722) | |
| Positive control | Fallopian tube+4 more · see all | |
| Negative control | Ovary |
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 (HPA tissue IHC) | |
| Regulation | Liver-enhanced RNA expression (HPA RNA) | |
| Isoform / epitope | No isoforms annotated; one 1–637 chain (UniProt) |
Section 1
Recommended GCLC IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published chromogenic GCLC IHC methods for hepatocellular carcinoma and animal-model liver tissue (PMC6603424; PMC7434920).
| Sample | Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A01722) |
| Fixation | Image fixative and duration unreported (datasheet A01722); 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 A01722); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A01722) |
| Primary antibody | Rabbit anti-GCLC, 2-5 μg/ml (datasheet A01722) |
| Primary incubation | Overnight at 4 °C (datasheet A01722) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A01722) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | GCLC-positive staining in glandular cells of fallopian tube (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression at variable levels. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet: A01722); neither published chromogenic protocol specifies retrieval (PMC6603424; PMC7434920).
Section 2
What Is the Expected GCLC Staining Pattern?
In paraffin-section IHC, expect variable cytoplasmic GCLC staining, including high staining in fallopian-tube glandular cells and urinary-bladder urothelial cells (HPA tissue IHC). HPA rates the tissue pattern Approved but reports low consistency between antibody staining and RNA expression (HPA tissue IHC). GCLC has no transmembrane segment or signal peptide, consistent with an intracellular pattern (UniProt P48506 topology and processing).
| Cytoplasmic stain in fallopian-tube glandular cells or bladder urothelial cells. | This matches the reported high-intensity IHC pattern in those cell types (HPA tissue IHC). Judge the positive cells within their tissue context; adjacent cells need not stain equally because HPA describes variable cytoplasmic expression (HPA tissue IHC). |
| Predominantly nuclear stain in paraffin-section IHC. | This departs from HPA's cytoplasmic tissue-IHC profile (HPA tissue IHC). Check controls and staining distribution before calling it an artefact: HPA separately reports approved nucleoplasmic and nucleolar localization in ICC-IF (HPA subcellular ICC-IF). |
| Strong stain in ovarian stromal cells. | HPA reports GCLC as not detected in those cells (HPA tissue IHC). Consider nonspecific antibody binding or endogenous detection activity (standard chromogenic IHC practice); the stain alone cannot identify which cause applies. |
| Color spread broadly across tissue, with little cell-specific contrast. | That distribution is harder to reconcile with cell-associated cytoplasmic expression (HPA tissue IHC). Review blocking, washes and chromogen development as general background controls (standard chromogenic IHC practice); broad color does not establish GCLC expression. |
| No stain in an otherwise interpretable fallopian-tube or bladder section. | A complete absence conflicts with the reported high staining in the specified glandular or urothelial cells (HPA tissue IHC). Check that those cells are present, then review the antibody and detection controls (standard IHC practice). |
💡Expected GCLC appearanceCall a positive IHC result when cytoplasmic color is strong in fallopian-tube glandular or bladder urothelial cells (HPA tissue IHC); dominant nuclear color or diffuse color without cell-specific contrast warrants control review (HPA tissue IHC; standard chromogenic IHC practice).
| Tissue and cell selection (HPA tissue IHC) | High staining is reported in fallopian-tube glandular and bladder urothelial cells; ovarian stromal cells are not detected (HPA tissue IHC). Compare the named cell populations, since HPA also reports medium and low staining elsewhere (HPA tissue IHC). |
| Evidence strength (HPA tissue IHC; HPA antibody validation) | The tissue profile is Approved, yet antibody staining has low consistency with RNA expression (HPA tissue IHC). Three listed antibodies have Approved IHC status; that does not by itself establish an Enhanced result (HPA antibody validation). |
| Cellular compartment and assay (HPA tissue IHC; HPA subcellular ICC-IF) | Tissue IHC is described as cytoplasmic, whereas ICC-IF is mainly nucleoplasmic with additional nucleolar and cytosolic localization (HPA tissue IHC; HPA subcellular ICC-IF). Interpret each observation in its own assay context. |
| Protein topology and processing (UniProt P48506) | The sequence has no transmembrane segment, signal peptide or propeptide; the annotated chain spans residues 1–637 (UniProt P48506 topology and processing). These facts support an intracellular expectation but specify neither an antibody epitope nor antigen-retrieval conditions. |
| Situation | Likely cause | Next action |
|---|---|---|
| Known-positive glandular or urothelial cells have no color (HPA tissue IHC). | The expected cells may be absent from the section, or the IHC workflow may have failed (standard IHC practice). | Confirm cell identity on the counterstain; run a documented positive control and inspect primary-antibody and detection steps (standard IHC practice). |
| Only weak color appears in a reported high-staining cell population (HPA tissue IHC). | Assay conditions may limit visible signal; HPA's high designation does not define a dilution or retrieval recipe (HPA tissue IHC; standard IHC practice). | Use the catalog antibody's IHC-P instructions and control tissue to assess retrieval, dilution and development (standard IHC practice). |
| The whole section develops brown color without useful cell boundaries. | Nonspecific binding or endogenous enzyme activity can contribute to chromogenic background (standard IHC practice). | Check blocking, washes and enzyme-control conditions; compare with a no-primary control (standard chromogenic IHC practice). |
| Ovarian stromal cells stain strongly despite HPA's not-detected result (HPA tissue IHC). | Possible cross-reactivity or endogenous detection activity needs testing; staining alone cannot distinguish them (standard IHC practice). | Compare a no-primary control and a known-positive section, then assess whether staining is cell-associated (standard IHC practice). |
| Tissue IHC shows dominant nuclear staining rather than cytoplasmic staining (HPA tissue IHC). | The pattern differs from HPA tissue IHC, although nuclear localization is reported in ICC-IF (HPA tissue IHC; HPA subcellular ICC-IF). | Review tissue controls and antibody specificity before interpreting the nuclear IHC signal as GCLC (standard IHC practice). |
| Why can ICC-IF show nuclear GCLC when tissue IHC is cytoplasmic? | HPA reports mainly nucleoplasmic ICC-IF localization, plus nucleoli and cytosol, alongside a cytoplasmic tissue-IHC profile (HPA subcellular ICC-IF; HPA tissue IHC). | Record the assay used and interpret each pattern against its corresponding HPA evidence; use the separate IF/ICC guide for that assay (HPA tissue IHC; HPA subcellular ICC-IF). |
Experiment evidence
IHC & IF images showing GCLC staining
A01722 · IHCIHC analysis of GCLC using anti-GCLC antibody (A01722). GCLC was detected in a paraffin-embedded section of human colon cancer tissue. Heat
A01722 · IHCIHC analysis of GCLC using anti-GCLC antibody (A01722). GCLC was detected in a paraffin-embedded section of human colon cancer tissue. Heat
A01722 · IFIF analysis of GCLC using anti-GCLC antibody (A01722). GCLC was detected in a paraffin-embedded section of human liver cancer tissue. Heat m
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000001084-GCLC/tissue/cerebral+cortex#imid_10195065
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000001084-GCLC/tissue/colon#imid_10194975
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000001084-GCLC/tissue/liver#imid_10194992
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000001084-GCLC/tissue/kidney#imid_10194977
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036360; green = GCLC, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA036360; green = GCLC, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for GCLC IHC & IF
🧪Run fallopian tube first and expect staining in its glandular cells (HPA: High in fallopian-tube glandular cells). Use ovary as the negative tissue, assessing its stromal cells (HPA: Not detected in ovarian stroma cells); on the positive slide, treat other morphologically distinct cells as internal background references only if they are unstained, because HPA does not identify a negative cell population there (HPA: fallopian-tube glandular-cell row).
Positive control tissue: Fallopian tube (Glandular cells, HPA High)
Negative control tissue: Ovary (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show GCLC in A-431, U-251MG, U2OS, with annotated localisation: Nucleoplasm (approved) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control, a concentration-matched rabbit IgG isotype control for the rabbit primary (selected A01722 caption: rabbit anti-GCLC), and GCLC-knockout material as a biological negative where available (standard IHC practice). For chromogenic detection, quench endogenous peroxidase and inspect the no-primary slide for background before scoring DAB signal (selected A01722 caption: peroxidase/DAB detection; standard IHC practice).
⚠️Feasibility: The selected paraffin-section caption uses heat-mediated EDTA retrieval at pH 8.0, but does not establish that retrieval is required; its fixative is unreported, and no target-specific fixation window or fixation effect is supplied (selected A01722 caption). Paraffin IHC is documented in that caption, while the supplied IF evidence consists of ICC images in A-431, U-251MG and U2OS; neither frozen sections nor IF can be judged easier from these sources (selected A01722 caption; HPA: ICC-IF image cell lines). No fallopian-tube-specific artefact is reported in the supplied evidence (HPA: fallopian-tube row; selected A01722 caption).
HPA tissue IHC evidence for GCLC
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Low 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 |
|---|---|---|---|---|
| Fallopian tube | Glandular cells | High | Protein (IHC) | HPA → |
| Urinary bladder | Urothelial cells | High | Protein (IHC) | HPA → |
| Adipose tissue | Adipocytes | Medium | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced GCLC IHC Tips
Troubleshoot GCLC staining in paraffin sections by checking retrieval, compartment, controls, and cell-specific scoring (A01722 caption; HPA tissue IHC; HPA subcellular).
What retrieval should I start with if GCLC staining is weak?
Start with heat-mediated retrieval in EDTA buffer at pH 8.0 (datasheet A01722). The selected paraffin-section image used that retrieval before incubation with 2 μg/ml antibody overnight at 4°C, so repeat those documented conditions before changing several variables together (A01722 caption). If staining remains weak, compare retrieval duration on adjacent sections while keeping antibody concentration, detection, and development consistent (standard IHC practice). Include a tissue region with documented GCLC staining, such as fallopian tube glandular cells, and assess both signal and tissue preservation; excessive heat can damage morphology and complicate scoring (HPA: High in fallopian tube glandular cells; standard IHC practice).
Could fixation explain weak GCLC staining in my paraffin sections?
The selected image documents a paraffin-embedded section but does not state its fixative, so target-specific GCLC sensitivity to fixation is unknown (A01722 caption). Record the actual fixative, fixation duration, and processing history for the sections under comparison; uneven fixation can change antigen accessibility across a section (standard IHC practice). Run adjacent sections with the documented EDTA pH 8.0 retrieval and 2 μg/ml primary antibody to limit other differences while assessing the effect of processing (datasheet A01722; A01722 caption; standard IHC practice). Do not infer fixation tolerance from GCLC tissue staining, its lack of a transmembrane segment, or its listed modified residues (HPA tissue IHC; UniProt P48506 topology and modified residues).
How should I judge nuclear and cytoplasmic GCLC staining?
In chromogenic tissue IHC, cytoplasmic staining is a documented pattern, with intensity varying across cell types (HPA tissue IHC). Cell imaging also places GCLC mainly in the nucleoplasm, with additional nucleolar and cytosolic localisation, so a nuclear signal warrants assessment rather than automatic rejection (HPA subcellular). Score nuclear and cytoplasmic compartments separately, using morphology and a matched no-primary control to judge whether either signal follows an interpretable cellular pattern (standard IHC practice). UniProt supplies no subcellular annotation and reports no transmembrane segment; those facts alone cannot establish which compartment a particular paraffin-section stain detects specifically (UniProt P48506 subcellular annotation and topology).
Could an isoform or processing event explain patchy GCLC staining?
The supplied GCLC record lists 0 isoforms and a single chain spanning residues 1–637, with no signal peptide or propeptide processing annotation (UniProt P48506 isoforms and processing). It also lists N-terminal acetylation and phosphorylation sites at residues 5 and 8, but the antibody epitope is unspecified here (UniProt P48506 modified residues; A01722 caption). Thus, patchy paraffin-section staining cannot be assigned to a particular isoform, cleavage product, or modification on this evidence (UniProt P48506; A01722 caption). Check whether the pattern follows cell boundaries, tissue preservation, and the no-primary control before interpreting it as biological variation (standard IHC practice).
How can I compare GCLC immunofluorescence with this chromogenic IHC result?
Use chromogenic IHC to identify the relevant cells first: fallopian tube glandular cells and urinary bladder urothelial cells have high reported tissue staining (HPA tissue IHC). For a separate IF experiment, multiplex GCLC with an independently validated marker for the expected cell type, then choose fluorophores whose emission can be distinguished from tissue autofluorescence and each other (standard IF practice). GCLC has no transmembrane segment, while cell imaging reports nucleoplasmic, nucleolar, and cytosolic localisation; permeabilisation is therefore needed to assess these intracellular compartments, although this antibody’s epitope accessibility is unspecified (UniProt P48506 topology; HPA subcellular; standard IF practice; A01722 caption). Compare compartment patterns with controls in each detection method rather than equating fluorescence intensity with DAB intensity (standard IHC/IF practice).
What should I change when GCLC DAB staining looks diffuse?
The selected paraffin-section example used 10% goat serum blocking, 2 μg/ml primary antibody overnight at 4°C, and a peroxidase-linked secondary followed by DAB (A01722 caption). First compare a no-primary section and inspect whether diffuse color appears in tissue compartments lacking a convincing cellular pattern (standard IHC practice). Apply an endogenous peroxidase block and check secondary-antibody background; these are general chromogenic IHC controls, not evidence of a GCLC-specific requirement (standard IHC practice). If the no-primary control is clean, optimise primary concentration, washing, and DAB development while retaining a reference section in each run (standard IHC practice).
How should I quantify variable GCLC staining across cell populations? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring because tissue IHC reports variable cytoplasmic GCLC expression, while cell imaging also reports nuclear localisation (HPA tissue IHC; HPA subcellular). For chromogenic sections, report the percentage of positive target cells and an intensity-weighted H-score, or positive-cell density per mm² when cell counts and area are more suitable (standard IHC practice). Normalise positive counts to the number of evaluable target cells, or density to analysed viable tissue area, using the same threshold and DAB development conditions across sections (standard IHC practice). Document excluded necrotic and damaged regions and report nuclear and cytoplasmic scores separately if both patterns occur (standard IHC practice).
How can I distinguish true GCLC staining from an artefact?
Look for reproducible staining within intact cells and compare its compartment with documented cytoplasmic tissue staining and nucleoplasmic, nucleolar, or cytosolic cell-imaging patterns (HPA tissue IHC; HPA subcellular; standard IHC practice). A plausible positive population includes fallopian tube glandular cells or urinary bladder urothelial cells, whereas ovarian stromal cells are reported as not detected in the supplied tissue profile (HPA tissue IHC). Discount isolated section-edge color, necrotic areas, and signal persisting without primary antibody; check endogenous peroxidase when DAB deposits appear unrelated to cells (standard IHC practice). Treat apparent agreement cautiously because the tissue IHC profile is approved but has low consistency with RNA expression data (HPA tissue IHC reliability).
Boster reagents
Best GCLC / Glutamate--cysteine ligase catalytic subunit IHC Antibodies
A01722 has IHC images from paraffin sections of human colon and liver cancers and an IF image from human liver cancer (catalog image captions); its listed reactivity is human, mouse and rat (catalog).
Real IHC data
Anti-GCLC Antibody ®
Cat # A01722
A01722 will render with its own IHC figure from a paraffin-embedded human colon cancer section (A01722 IHC image caption). Its IF image shows a paraffin-embedded human liver cancer section (A01722 IF image caption).
Which to pick: For tissue IHC, choose A01722 or PB9201: each has an IHC image from a human paraffin section, and neither caption reports the fixative (respective IHC image captions). For IF, choose A01722 for the documented human tissue example; M01722-2 lists IF but has no IF image, and ICC validation is not listed for either (catalog applications and image captions). A01722 lists human, mouse and rat reactivity across its applications, but its pictured IHC and IF examples are human, so validate staining in mouse or rat tissue before use (catalog reactivity and A01722 image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P48506 (GSH1_HUMAN, Glutamate--cysteine ligase catalytic subunit).
- Human Protein Atlas. GCLC tissue IHC expression (reliability: Approved).
- Human Protein Atlas. GCLC subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the nucleoli and cytosol..
- Human Protein Atlas. GCLC antibody validation summary (3 antibodies).
- High GCLC level in tumor tissues is associated with poor prognosis of hepatocellular carcinoma after curative resection. Journal of Cancer 2019 — PMC6603424.
- Computational repositioning of dimethyl fumarate for treating alcoholic liver disease. Cell death & disease 2020 — PMC7434920.
- Neuronal glutathione loss leads to neurodegeneration involving gasdermin activation. Scientific reports 2023 — PMC9859798.
- Characterization of the proteome of stable and unstable carotid atherosclerotic plaques using data-independent acquisition mass spectrometry. Journal of translational medicine 2024 — PMC10921703.
- PubMed PMID:1350904 — UniProt-cited evidence.
- PubMed PMID:14574404 — UniProt-cited evidence.
- PubMed PMID:15489334 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for GCLC (Glutamate--cysteine ligase catalytic subunit). Expected localisation Variable cytoplasmic staining (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A01722). Validated IHC antibody: A01722 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Fallopian tube (Glandular cells); negative Ovary. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human colon cancer tissue; fixative not specified (datasheet A01722); 2. Fixation: Image fixative and duration unreported (datasheet A01722); 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 A01722); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A01722); 8. Primary antibody: Rabbit anti-GCLC, 2-5 μg/ml (datasheet A01722); 9. Primary incubation: Overnight at 4 °C (datasheet A01722). Watch for: Fixation: Keep fixation conditions consistent across sections. (standard IHC practice; not target-specific); Caveat: Antibody staining has low consistency with RNA (HPA tissue IHC); Isoform / epitope: No isoforms annotated; one 1–637 chain (UniProt); Regulation: Liver-enhanced RNA expression (HPA RNA). Evidence: UniProt P48506, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the nucleoplasm. In addition localized to the…, Datasheet A01722, PMC6603424, PMC7434920. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/GCLC/immunohistochemistry-ihc-guide.