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AGT / Angiotensinogen · IHC design guide
Design Immunohistochemistry for AGT
This paraffin IHC guide uses high proximal tubule cell-body staining and medium cytotrophoblast staining as AGT reference patterns (HPA tissue IHC). It covers fixation and interpretation of a secreted plasma protein (UniProt; HPA tissue IHC), alongside the catalog antibody’s 2–5 μg/ml IHC range (datasheet A02103-4).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
AGT 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 | Plasma positivity; proximal tubule cell bodies (HPA tissue IHC) | |
| Staining pattern | High proximal tubule cell-body staining; medium cytotrophoblast staining (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02103-4) | |
| Positive control | Kidney+1 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific) | |
| Caveat | Secreted plasma AGT may make tissue staining differ from RNA levels (UniProt; HPA tissue IHC) | |
| Regulation | Stimulus-dependent regulation unreported (UniProt; HPA tissue IHC) | |
| Isoform / epitope | 0 isoforms; mature chain spans residues 25–476 (UniProt) |
Section 1
Recommended AGT IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published angiotensinogen staining methods for rat liver and kidney (datasheet A02103-4; PMC3560510; PMC2974762).
| Sample | Paraffin-embedded human lung adenocarcinoma tissue; fixative not specified (datasheet A02103-4) |
| Fixation | Image fixative and duration unreported (datasheet A02103-4); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A02103-4); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02103-4) |
| Primary antibody | Rabbit anti-AGT, 2-5 μg/ml (datasheet A02103-4) |
| Primary incubation | Overnight at 4 °C (datasheet A02103-4) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A02103-4) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | AGT-positive staining in proximal tubules (cell body) of kidney (HPA tissue IHC: High). HPA tissue profile: Positivity in plasma. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet A02103-4); the published rat protocols provide tissue and detection details (PMC3560510; PMC2974762).
Section 2
What Is the Expected AGT Staining Pattern?
AGT is secreted, has no transmembrane segment, and is expressed by the liver and released into plasma (UniProt P01019). On tissue sections, expect staining in kidney proximal tubule cell bodies and placenta cytotrophoblasts (HPA: High in proximal tubules; Medium in cytotrophoblasts). HPA rates its tissue IHC pattern Supported, while cautioning that protein location and tissue RNA expression may differ for a secreted protein (HPA tissue IHC).
| Clear proximal tubule cell-body staining in kidney, with less intense cytotrophoblast staining in placenta. | This matches the reported relative levels: High in kidney proximal tubules and Medium in placental cytotrophoblasts (HPA tissue IHC). Assess the cells identified by the counterstain before assigning a positive result (standard IHC practice). |
| Predominantly nuclear staining, or a crisp membrane outline presented as the expected AGT pattern. | Treat this as discordant and check controls: UniProt describes a secreted protein without a transmembrane segment, and HPA reports proximal tubule cell-body staining (UniProt P01019; HPA tissue IHC). The HPA subcellular summary says Membrane, Secreted but provides no main location or ICC-IF images (HPA subcellular). |
| Strong staining in adipocytes or adrenal glandular cells while the expected positive cells are weak. | Consider cross-reactivity or endogenous chromogenic activity; neither is established by that slide alone (standard IHC practice). These cell populations are reported as Not detected, so compare them with a positive kidney section and a no-primary control (HPA tissue IHC; standard IHC practice). |
| Diffuse color across tissue, including spaces outside identifiable cells. | A secreted, plasma-associated target makes extracellular signal possible, but diffuse color alone cannot identify AGT (UniProt P01019; HPA: positivity in plasma). Review tissue morphology, wash and blocking conditions, and the no-primary control before calling specific staining (standard IHC practice). |
| No signal in kidney proximal tubules on a run intended to detect AGT. | This conflicts with the reported High proximal tubule level (HPA tissue IHC). Confirm that the section contains recognizable tubules, then assess the run controls and IHC-validated antibody conditions; an absent signal does not by itself establish absent AGT (standard IHC practice). |
💡Expected AGT appearanceCall a section positive when identifiable kidney proximal tubule cell bodies stain strongly, with Medium cytotrophoblast staining as a second reference; isolated nuclear or widespread staining in reported negative cell types is suspect (HPA tissue IHC; UniProt P01019 topology).
| Tissue source versus deposited protein | Liver-enriched RNA and liver expression do not require liver to be the only protein-positive site: AGT enters plasma, and HPA observes kidney and placental staining (UniProt P01019; HPA tissue IHC). Interpret tissue staining as protein presence, without inferring local synthesis from IHC alone (standard IHC practice). |
| Secretory processing and topology | AGT has a signal peptide at residues 1–24 and a mature chain at 25–476, with no transmembrane segment (UniProt P01019). Because the antibody epitope is unspecified here, these annotations cannot predict whether processing affects its staining (UniProt P01019; supplied antibody record). |
| Tissue IHC validation | The listed antibody, CAB025798, has Supported IHC status; HPA also rates the tissue pattern Supported (HPA antibodies; HPA tissue IHC). That supports comparison with reported tissues, but does not establish an Enhanced, independently reproduced pattern (HPA antibodies; HPA validation definition). |
| Plasma-associated distribution | HPA explicitly describes positivity in plasma, consistent with UniProt's secreted annotation (HPA tissue IHC; UniProt P01019). Evaluate extracellular color alongside defined cell staining and controls; plasma-associated signal alone cannot identify which nearby cells produced AGT (standard IHC practice). |
| Situation | Likely cause | Next action |
|---|---|---|
| Kidney proximal tubules show no signal. | Possible run failure or insufficient detection; the expected level is High (HPA tissue IHC; standard IHC practice). | Check morphology, positive and no-primary controls, retrieval and detection steps, then repeat using documented conditions for the IHC-validated antibody (standard IHC practice). |
| The whole section has diffuse chromogenic color. | Possible background or endogenous detection activity; AGT is also plasma-associated (standard IHC practice; HPA tissue IHC). | Inspect the no-primary control, blocking, washes and endogenous enzyme quenching; score identifiable cells separately from extracellular color (standard IHC practice). |
| Nuclei dominate the positive signal. | This is discordant with secreted AGT and the reported proximal tubule cell-body pattern (UniProt P01019; HPA tissue IHC). | Verify compartment against the counterstain and no-primary control; reassess antibody and detection specificity before scoring (standard IHC practice). |
| Adipocytes stain strongly. | Cross-reactivity or background is possible because HPA reports adipocytes as Not detected in sampled tissues (HPA tissue IHC; standard IHC practice). | Compare kidney on the same run, inspect no-primary staining and require cell-specific morphology before assigning AGT positivity (HPA tissue IHC; standard IHC practice). |
| Placenta is weaker than kidney. | That ordering matches Medium cytotrophoblast versus High proximal tubule levels (HPA tissue IHC). | Score the identified cell populations separately; investigate only if the positive controls or expected cells fail to stain (HPA tissue IHC; standard IHC practice). |
| Can this IHC pattern serve as an ICC-IF expectation? | No ICC status or ICC-IF images are supplied for CAB025798; HPA lists no main subcellular location (HPA antibodies; HPA subcellular). | Use the separate IF/ICC guide for assay-specific validation; treat HPA's Membrane, Secreted summary as context, not a verified image pattern for this antibody (HPA subcellular; HPA antibodies). |
Experiment evidence
IHC & IF images showing AGT staining
A02103-4 · IHCIHC analysis of Angiotensinogen/AGT using anti-Angiotensinogen/AGT antibody (A02103-4). Angiotensinogen/AGT was detected in a paraffin-embed
A02103-4 · IHCIHC analysis of Angiotensinogen/AGT using anti-Angiotensinogen/AGT antibody (A02103-4). Angiotensinogen/AGT was detected in a paraffin-embed
A02103-3 · IHCIHC analysis of Angiotensinogen/AGT using anti-Angiotensinogen/AGT antibody (A02103-3). Angiotensinogen/AGT was detected in a paraffin-embed
A02103-2 · IFICC staining Angiotensinogen in Hela cells (green). The nuclear counter stain is DAPI (blue). Cells were fixed in paraformaldehyde, permeabi
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000135744-AGT/tissue/cerebral+cortex#imid_18740864
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000135744-AGT/tissue/colon#imid_18740782
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000135744-AGT/tissue/liver#imid_18740791
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000135744-AGT/tissue/kidney#imid_18740792
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for AGT IHC & IF
🧪Run kidney first: proximal tubule cell bodies should stain (HPA: High in kidney proximal tubules). Use adipose tissue as the negative: adipocytes should show no specific staining (HPA: Not detected in adipocytes); neighboring cells outside the proximal tubules provide an internal background comparison (HPA: High staining is assigned to proximal tubules).
Positive control tissue: Kidney (Proximal tubules (cell body), 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 AGT; derive a cell-line control from the positive tissue's cell type (Proximal tubules (cell body)) and confirm it by RNA or western blot first.
Technical controls: Include a no-primary, secondary-only slide and a concentration-matched rabbit IgG isotype control for the rabbit catalog antibody (selected-SKU caption: rabbit primary and goat anti-rabbit secondary); AGT-knockout tissue, if available, provides a biological specificity control (standard IHC practice). For kidney, assess endogenous peroxidase before interpreting DAB staining and account for proximal-tubule autofluorescence if using IF (standard IHC/IF practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported in the supplied evidence; the selected-SKU paraffin-section caption does not state a fixative (selected-SKU caption: fixative unreported). Heat retrieval in EDTA at pH 8.0 was used for the captioned lung adenocarcinoma section, but an AGT-specific retrieval dependency is unreported (selected-SKU caption: heat retrieval in EDTA, pH 8.0). The supplied evidence does not establish that frozen sections or IF are easier; in kidney, proximal-tubule autofluorescence can complicate IF interpretation (HPA: no ICC-IF cell-line images; standard IF practice).
HPA tissue IHC evidence for AGT
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — At least one protein variant secreted, tissue location of RNA and protein might differ and correlation is complex.). 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 |
|---|---|---|---|---|
| Kidney | Proximal tubules (cell body) | High | Protein (IHC) | HPA → |
| Placenta | Cytotrophoblasts | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | 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 → |
Section 3
Advanced AGT IHC Tips
Troubleshoot AGT staining in paraffin sections by checking retrieval, controls, cellular distribution, and how the signal is scored.
What retrieval should I start with for AGT staining in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 (datasheet A02103-4). The selected paraffin-section image used that retrieval before 2 μg/ml primary antibody overnight at 4°C, so keep those conditions together for the first comparison (datasheet A02103-4). If staining is weak, compare a shorter and longer heating period on matched sections while keeping the buffer and detection steps fixed (standard IHC practice). Include a known positive section and a no-primary control in the same run, then assess whether added heating improves cell-associated signal without increasing diffuse or edge staining (standard IHC practice).
How should I troubleshoot possible fixation effects on AGT IHC?
Target-specific fixation sensitivity is unknown: the selected image describes a paraffin-embedded section but does not state its fixative (datasheet A02103-4). Record each specimen's fixative and fixation duration, then compare matched sections processed with the same pH 8.0 EDTA retrieval and 2 μg/ml primary antibody conditions (datasheet A02103-4; standard IHC practice). If signal varies between specimens, inspect tissue preservation and section adherence before changing retrieval or antibody concentration (standard IHC practice). Keep a reference section in each staining run so differences in processing can be assessed separately from changes in chromogenic development (standard IHC practice).
Where should convincing AGT signal appear in tissue sections?
Interpret AGT staining with its secreted biology in mind: UniProt identifies AGT as secreted, with no transmembrane segment, and HPA reports plasma positivity (UniProt P01019; HPA: positivity in plasma). HPA reports high staining in kidney proximal tubule cell bodies and medium staining in placental cytotrophoblasts, providing cell-level reference patterns (HPA: kidney proximal tubules high; placenta cytotrophoblasts medium). Compare cell-associated staining with adjacent extracellular or vascular signal on the same section, rather than assigning every deposit to local synthesis (UniProt P01019; standard IHC practice). Use tissue morphology and a matched negative control to judge whether the distribution is reproducible (standard IHC practice).
Could AGT processing or glycosylation change what this antibody detects?
AGT has no annotated isoforms in the supplied record, but its signal peptide spans residues 1–24 and the angiotensinogen chain spans 25–476 (UniProt P01019). Four glycosylation sites are annotated at residues 38, 161, 295, and 319, so epitope accessibility may depend on which region the antibody recognizes (UniProt P01019; standard IHC practice). The selected caption does not identify the antibody epitope, so a negative section cannot distinguish absent AGT from an inaccessible epitope (datasheet A02103-4; standard IHC practice). Check epitope documentation before interpreting discordant specimens, and compare staining under the documented retrieval conditions (standard IHC practice; datasheet A02103-4).
How can IF help clarify ambiguous AGT IHC staining?
For a separate IF experiment, pair AGT with a validated marker of the expected cell type, such as a proximal tubule marker when assessing kidney, and check whether the signals occupy the same cells (HPA: kidney proximal tubules high; standard IF practice). Choose spectrally separated fluorophores and place the weaker channel away from prominent tissue autofluorescence; include single-stain controls for multiplex interpretation (standard IF practice). AGT is secreted and has no transmembrane segment, so select permeabilisation according to whether the chosen epitope is being assessed in an intracellular pool or an accessible extracellular pool (UniProt P01019; standard IF practice). Confirm IF findings against tissue morphology and the chromogenic IHC distribution (standard IF/IHC practice).
What should I check when AGT DAB staining looks diffuse?
First compare the stained section with a no-primary control and inspect vascular spaces, tissue edges, and damaged areas for signal that does not follow intact cells (standard IHC practice). AGT is secreted and HPA reports plasma positivity, so extracellular or vascular colour alone does not establish local cellular expression (UniProt P01019; HPA: positivity in plasma). The selected protocol used 10% goat serum blocking, an anti-rabbit peroxidase secondary, and DAB development (datasheet A02103-4). Check peroxidase blocking and DAB development on matched sections, then adjust primary incubation only after confirming that the detection system itself is clean (standard IHC practice).
How should I score AGT staining across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cellular compartment and tissue region before scoring, because secreted AGT can also appear in plasma-associated spaces (UniProt P01019; HPA: positivity in plasma; standard IHC practice). For a cell-based comparison, record percentage-positive cells and an H-score from 0–300 within the same morphologically defined population (standard IHC practice). Normalise counts to the number of eligible cells, or normalise positive-cell density to sampled area in mm², while excluding vessels and damaged regions according to prespecified rules (standard IHC practice). Keep retrieval, 2 μg/ml primary concentration, imaging, and DAB development consistent across sections before comparing scores (datasheet A02103-4; standard IHC practice).
How can I distinguish genuine AGT positivity from staining artefact?
Look for reproducible cell-associated staining in an anatomically appropriate population, with high kidney proximal tubule cell-body staining and medium placental cytotrophoblast staining as supplied references (HPA: kidney proximal tubules high; placenta cytotrophoblasts medium). AGT is secreted and plasma positive, so staining confined to vascular contents does not by itself show production by nearby cells (UniProt P01019; HPA: positivity in plasma). Treat strong section-edge colour, necrotic deposits, and signal reproduced in a no-primary control as possible artefacts (standard IHC practice). If colour follows an endogenous enzyme-rich area, review the peroxidase block and repeat the comparison under matched DAB development conditions (standard IHC practice).
Boster reagents
Best AGT / Angiotensinogen IHC Antibodies
Human AGT antibodies have IHC images from paraffin-embedded lung adenocarcinoma and pancreatic cancer sections, plus an ICC image from HeLa cells (catalog image captions; catalog reactivity: Human).
Real IHC data
Anti-Angiotensinogen/AGT Antibody ®
Cat # A02103-4
Real IHC data
Anti-Angiotensinogen/AGT Antibody ®
Cat # A02103-3
Real IF data
Anti-Angiotensinogen AGT Antibody
Cat # A02103-2
A02103-4 shows IHC staining in human lung adenocarcinoma and rectum adenocarcinoma paraffin sections; A02103-3 shows IHC staining in a human pancreatic cancer paraffin section (catalog IHC image captions). A02103-2 shows ICC staining in HeLa and HepG2 cells and is listed for IF/ICC (catalog IF image captions; catalog applications).
Which to pick: For human tissue IHC, choose A02103-4 for the documented lung or rectum adenocarcinoma examples, or A02103-3 for the pancreatic cancer example; each SKU’s own caption reports EDTA retrieval at pH 8.0 and an overnight 2 μg/ml incubation, but does not report the fixative (catalog IHC image captions). For IF/ICC, choose A02103-2: it is listed for IF/ICC at 1:50–1:200 and has HeLa and HepG2 ICC images (catalog applications, dilution and IF image captions). No cross-species choice is supported: every listed SKU has Human reactivity only (catalog reactivity).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P01019 (ANGT_HUMAN, Angiotensinogen).
- Human Protein Atlas. AGT tissue IHC expression (reliability: Supported).
- Human Protein Atlas. AGT subcellular location (ICC-IF): Membrane, Secreted.
- Human Protein Atlas. AGT antibody validation summary (1 antibodies).
- Expression of angiotensinogen during hepatic fibrogenesis and its effect on hepatic stellate cells. Medical science monitor : international medical journal of experimental and clinical research 2011 — PMC3560510.
- Mismatch repair deficiency does not mediate clinical resistance to temozolomide in malignant glioma. Clinical cancer research : an official journal of the American Association for Cancer Research 2008 — PMC2830553.
- Glomerular angiotensinogen is induced in mesangial cells in diabetic rats via reactive oxygen species--ERK/JNK pathways. Hypertension research : official journal of the Japanese Society of Hypertension 2010 — PMC2974762.
- Evidence of Augmented Intrarenal Angiotensinogen Associated With Glomerular Swelling in Gestational Hypertension and Preeclampsia: Clinical Implications. Journal of the American Heart Association 2019 — PMC6662362.
- PubMed PMID:6089875 — UniProt-cited evidence.
- PubMed PMID:2924688 — UniProt-cited evidence.
- PubMed PMID:1692023 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for AGT (Angiotensinogen). Expected localisation Plasma positivity; proximal tubule cell bodies (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02103-4). Validated IHC antibody: A02103-4 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Kidney (Proximal tubules (cell body)); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung adenocarcinoma tissue; fixative not specified (datasheet A02103-4); 2. Fixation: Image fixative and duration unreported (datasheet A02103-4); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: EDTA pH 8.0 (datasheet A02103-4); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02103-4); 8. Primary antibody: Rabbit anti-AGT, 2-5 μg/ml (datasheet A02103-4); 9. Primary incubation: Overnight at 4 °C (datasheet A02103-4). Watch for: Fixation: Keep paraffin-section fixation consistent across samples. (standard IHC practice; not target-specific); Caveat: Secreted plasma AGT may make tissue staining differ from RNA levels (UniProt; HPA tissue IHC); Isoform / epitope: 0 isoforms; mature chain spans residues 25–476 (UniProt); Regulation: Stimulus-dependent regulation unreported (UniProt; HPA tissue IHC). Evidence: UniProt P01019, HPA tissue IHC · Supported, HPA ICC-IF · Membrane, Secreted, Datasheet A02103-4, PMC3560510, PMC2974762. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/AGT/immunohistochemistry-ihc-guide.