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HSD3B1 / 3 beta-hydroxysteroid dehydrogenase/Delta 5-->4-isomerase type 1 · IHC design guide
Design Immunohistochemistry for HSD3B1
Use placental syncytiotrophoblasts as a positive tissue control (HPA tissue IHC). For paraffin-section IHC, start with 0.5–1 μg/mL of the IHC-validated antibody A02856-2 (datasheet A02856-2).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
HSD3B1 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 placenta and adrenal gland (HPA tissue IHC) | |
| Staining pattern | Cytoplasmic staining in placental syncytiotrophoblasts (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A02856-2) | |
| Positive control | Placenta+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 fixation consistent across sections. (standard IHC practice; not target-specific) | |
| Caveat | Cross-reactivity possible: antibody targets more than one gene (HPA tissue IHC) | |
| Regulation | Placenta high; adrenal zona fasciculata low (HPA tissue IHC) | |
| Isoform / epitope | No isoforms annotated; epitope sidedness unassigned (UniProt) |
Section 1
Recommended HSD3B1 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is paired with published HSD3B1 staining of human aldosterone-producing adenoma and rat testis (PMC4821076; PMC5643909).
| Sample | Paraffin-embedded human placenta tissue; fixative not specified (datasheet A02856-2) |
| Fixation | Image fixative and duration unreported (datasheet A02856-2); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A02856-2); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A02856-2) |
| Primary antibody | Rabbit anti-HSD3B1, 0.5-1μg/ml (datasheet A02856-2) |
| Primary incubation | Overnight at 4 °C (datasheet A02856-2) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A02856-2) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | HSD3B1-positive staining in syncytiotrophoblasts of placenta (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in placenta and adrenal gland. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA at pH 8.0 for the catalog antibody (datasheet: A02856-2). The rat testis study used citrate at pH 6.0 (PMC5643909).
Section 2
What Is the Expected HSD3B1 Staining Pattern?
HSD3B1 is an endoplasmic reticulum and mitochondrial membrane protein with one transmembrane segment at residues 288–308 (UniProt P14060 topology). In paraffin IHC, expect cytoplasmic staining strongest in placental syncytiotrophoblasts, with weaker staining reported in adrenal zona fasciculata cells (HPA: High in syncytiotrophoblasts; Low in zona fasciculata). HPA rates its tissue profile Supported but cautions that the antibodies can target proteins from more than one gene (HPA: tissue reliability).
| Placental syncytiotrophoblasts stain strongly in the cytoplasm, while nearby structures provide visible contrast (HPA: High in syncytiotrophoblasts; cytoplasmic profile). | This matches the strongest documented tissue result and the expected view of a membrane-associated protein at routine IHC resolution (HPA: High in syncytiotrophoblasts; UniProt P14060 subcellular location). Score the named cell population rather than averaging stain across the whole section (general IHC practice). |
| A dominant nuclear or nucleolar deposit replaces cytoplasmic staining in placental syncytiotrophoblasts (HPA: cytoplasmic tissue profile). | Treat this as a localization discrepancy and check the chromogen, counterstain and antibody controls (general IHC practice). Nucleolar localization is uncertain in HPA ICC-IF and does not establish a nucleolar IHC pattern (HPA: subcellular location, uncertain). |
| Strong staining appears chiefly in cells listed as undetected, such as adipocytes or appendix glandular cells (HPA: Not detected in those cells). | Question specificity before calling those cells HSD3B1 positive: HPA cautions that its antibodies may target more than one gene (HPA: tissue reliability). A positive chromogen deposit can also reflect endogenous detection activity; assess it with an appropriate detection control (general IHC practice). |
| Color spreads across stroma, section edges or many unrelated cell types without a defined cytoplasmic pattern (HPA: cytoplasmic tissue profile). | Interpret the spread as background until controls support cellular staining (general IHC practice). Examine blocking, washes and detection conditions; the HPA tissue profile alone cannot identify which workflow step produced the background (HPA: tissue profile; general IHC practice). |
| Placental syncytiotrophoblasts show no convincing stain on a slide intended as a positive control (HPA: High in syncytiotrophoblasts). | An absent signal conflicts with the reported positive tissue pattern, but does not by itself establish absent protein (HPA: High in syncytiotrophoblasts). Review tissue identity, antibody and detection controls, and the general IHC workflow before interpreting the sample (general IHC practice). |
💡Expected HSD3B1 appearanceCall a positive result when placental syncytiotrophoblasts show High cytoplasmic staining; dominant nuclear staining or equally strong staining in HPA-undetected cells warrants review (HPA: tissue IHC profile).
| Positive tissue and cell selection (HPA: tissue IHC profile). | Placental syncytiotrophoblasts offer the clearest reported positive comparison (High); testicular spermatogonia are Medium and adrenal zona fasciculata cells are Low (HPA: tissue IHC). A weak adrenal result should therefore be judged against its reported level, not against placental intensity (HPA: tissue IHC). |
| Membrane topology and epitope access (UniProt P14060 topology). | HSD3B1 has a transmembrane segment at 288–308 and is assigned to endoplasmic reticulum and mitochondrial membranes (UniProt P14060). The supplied record gives no antibody epitope, so topology cannot establish which side of either membrane an antibody recognizes or prescribe antigen retrieval (UniProt P14060 topology; supplied antibody data). |
| Antibody interpretation (HPA: tissue reliability; antibody validation). | The tissue profile is Supported, and the three listed antibodies have Supported IHC status (HPA: tissue reliability; HPA043261, HPA043264, HPA044028). HPA also reports possible binding to proteins from more than one gene; agreement with the expected cells strengthens interpretation but cannot, alone, prove target specificity (HPA: tissue reliability). |
| IF/ICC Q&A: should nucleolar or ciliary fluorescence count as the expected result? (HPA: subcellular ICC-IF). | Endoplasmic reticulum localization is Supported in ICC-IF; nucleolar, primary cilium and other listed locations are Uncertain, with a multiple-gene antibody caution (HPA: subcellular ICC-IF). Assess any such fluorescence in its own controls; those observations do not redefine the cytoplasmic paraffin IHC expectation (HPA: tissue IHC; subcellular ICC-IF). |
| Situation | Likely cause | Next action |
|---|---|---|
| Placental syncytiotrophoblasts are unstained (HPA: High in syncytiotrophoblasts). | The expected positive control has failed, or the intended cell population is absent from the examined area (HPA: High in syncytiotrophoblasts; general IHC practice). | Confirm the cells on the counterstained section, then check antibody application, detection reagents and control slides before judging other tissue (general IHC practice). |
| Placental staining is present but faint (HPA: High in syncytiotrophoblasts). | A weak run is possible, although intensity cannot be attributed to a specific step from the HPA profile alone (HPA: tissue IHC; general IHC practice). | Compare with a run control and review antibody dilution, antigen retrieval and detection conditions as general paraffin IHC variables; no HSD3B1-specific retrieval condition is supplied (general IHC practice; supplied evidence). |
| Adrenal zona fasciculata looks weaker than placenta (HPA: Low in zona fasciculata; High in syncytiotrophoblasts). | That difference is consistent with the reported tissue levels and is not, by itself, a failed assay (HPA: tissue IHC). | Judge adrenal cells against the reported Low pattern and verify the placental positive control before increasing detection intensity (HPA: tissue IHC; general IHC practice). |
| Chromogen appears across many cells or in cell-free areas (HPA: cytoplasmic tissue profile). | Nonspecific background or endogenous detection activity is possible; appearance alone cannot assign the cause (general IHC practice). | Inspect a no-primary control, blocking and wash steps, and the chromogenic detection system; compare any remaining cellular stain with the HPA cell pattern (general IHC practice; HPA: tissue IHC). |
| Adipocytes or appendix glandular cells stain strongly (HPA: Not detected in those cells). | The result conflicts with HPA tissue observations, and HPA flags possible binding to proteins from more than one gene (HPA: tissue IHC; tissue reliability). | Check controls and examine the same run's placental pattern; seek independent specificity evidence before assigning the unexpected stain to HSD3B1 (general IHC practice; HPA: tissue reliability). |
| Nuclear or nucleolar staining dominates a paraffin section (HPA: cytoplasmic tissue profile). | Localization conflicts with the reported IHC pattern; uncertain nucleolar ICC-IF localization cannot resolve that conflict (HPA: tissue IHC; subcellular ICC-IF). | Review counterstain, detection controls and the antibody's IHC validation, then score cytoplasmic staining separately from the discrepant deposit (general IHC practice; HPA: antibody validation). |
Experiment evidence
IHC & IF images showing HSD3B1 staining
A02856-2 · IHCIHC analysis of HSD3B1 using anti-HSD3B1 antibody (A02856-2). HSD3B1 was detected in paraffin-embedded section of human placenta tissue. Hea
A02856-2 · IFIF analysis of HSD3B1 using anti-HSD3B1 antibody (A02856-2). HSD3B1 was detected in immunocytochemical section of U20S cells. Enzyme antigen
M02856 · IFIF analysis of HSD3B1 using anti-HSD3B1 antibody (M02856). HSD3B1 was detected in an immunocytochemical section of U20S cells. Enzyme antige
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000203857-HSD3B1/tissue/cerebral+cortex#imid_19831477
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000203857-HSD3B1/tissue/colon#imid_19831395
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000203857-HSD3B1/tissue/liver#imid_19831396
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000203857-HSD3B1/tissue/kidney#imid_19831397
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA044028; green = HSD3B1, blue = nucleus, red = microtubules.
HPA · ICC-IF CACO-2Human Protein Atlas immunofluorescence, CACO-2, antibody HPA044028; green = HSD3B1, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for HSD3B1 IHC & IF
🧪Run placenta first: syncytiotrophoblasts should stain (HPA: High in placental syncytiotrophoblasts). Use appendix glandular cells as a negative tissue (HPA: Not detected); on the placenta slide, assess non-syncytiotrophoblast areas as internal negative areas rather than assuming every cell will stain (HPA: High in syncytiotrophoblasts).
Positive control tissue: Placenta (Syncytiotrophoblasts, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show HSD3B1 in CACO-2, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), with annotated localisation: Nucleoli (uncertain), Endoplasmic reticulum (supported), Primary cilium (uncertain) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit isotype control alongside a biological specificity control such as HSD3B1 knockout tissue or a validated peptide block (datasheet: rabbit primary; standard IHC practice). For placental DAB staining with a biotin-based detection system, block endogenous peroxidase and check for endogenous biotin background (selected-SKU IHC caption: biotinylated secondary, streptavidin–biotin detection and DAB; standard IHC practice).
⚠️Feasibility: The selected-SKU paraffin-section caption does not report a fixative, and the supplied evidence reports no HSD3B1-specific fixation window or fixation effect (selected-SKU IHC caption: fixative not stated). Heat retrieval in EDTA at pH 8.0 produced placental staining, but whether retrieval is required has not been established (selected-SKU IHC caption: EDTA retrieval and placental staining). The evidence does not establish whether frozen sections or IF are easier; with the caption’s biotin-based placental IHC method, endogenous biotin is a background concern to check (selected-SKU IHC caption: biotin-based detection; standard IHC practice).
HPA tissue IHC evidence for HSD3B1
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Supported — Caution, targets protein from more than one gene. Presumed off target binding observed and disregarded. Paired antibodies with high similarity supports the expression profile.). 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 |
|---|---|---|---|---|
| Placenta | Syncytiotrophoblasts | High | Protein (IHC) | HPA → |
| Testis | Spermatogonia 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 | Glandular 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 HSD3B1 IHC Tips
Troubleshooting HSD3B1 staining in paraffin sections starts with the validated retrieval conditions, compartment-aware controls, and cautious interpretation of antibody specificity.
How should I optimize antigen retrieval when placental HSD3B1 staining is weak?
Start with heat-mediated retrieval in EDTA pH 8.0 for paraffin sections (datasheet A02856-2). The catalog antibody produced a placental IHC image after this retrieval, followed by 10% goat serum blocking and 1 μg/ml primary antibody overnight at 4°C (caption A02856-2). If staining remains weak, vary heating duration and cooling conditions on matched sections while keeping detection and exposure to DAB consistent (standard IHC practice). Judge improvement by signal in placental syncytiotrophoblasts and by background in cells expected to be unstained, since strong staining alone does not establish specificity (HPA: High in placental syncytiotrophoblasts; HPA: presumed off-target binding).
Could tissue fixation be causing weak or uneven HSD3B1 staining?
Target-specific fixation sensitivity is unknown because the selected placental caption calls the specimen paraffin-embedded but does not state its fixative (caption A02856-2). Record fixative, fixation duration, section age and processing history for each block before comparing staining across samples (standard IHC practice). Run matched sections through the same EDTA pH 8.0 retrieval and 1 μg/ml primary-antibody condition used in the catalog example, then compare tissue preservation and background alongside signal (caption A02856-2; standard IHC practice). If fixation histories differ, interpret differences cautiously and use a similarly processed placental control; neither tissue-expression data nor membrane topology demonstrates a particular fixation effect (HPA: placental expression; UniProt P14060 topology).
Where should HSD3B1 staining appear, and how should nuclear signal be assessed?
Expect predominantly cytoplasmic staining in placental IHC, especially in syncytiotrophoblasts (HPA: cytoplasmic expression; HPA: High in placental syncytiotrophoblasts). This pattern is compatible with HSD3B1 at endoplasmic reticulum and mitochondrial membranes, with a predicted transmembrane segment at residues 288–308 (UniProt P14060 subcellular location and topology). Review nuclear or nucleolar staining separately from cytoplasmic staining: the reported nucleolar location is uncertain and comes from antibodies that may recognize proteins from multiple genes (HPA subcellular: nucleoli uncertain; HPA subcellular caution). Compare the suspect pattern with a placental control, negative-control slide and cell morphology before treating it as HSD3B1-specific signal (standard IHC practice).
How can I assess epitope accessibility and possible cross-reactivity?
Map the antibody's documented immunogen against HSD3B1 before interpreting weak staining or a surprising compartment, because no epitope sequence is supplied here (standard antibody-validation practice). UniProt lists a 373-aa precursor, a chain spanning residues 2–373, one transmembrane segment at 288–308, and no annotated isoforms or glycosylation sites (UniProt P14060). Those annotations do not establish which side of a membrane contains this antibody's epitope or whether retrieval exposes it (UniProt P14060 topology; standard IHC practice). Seek an independently validated antibody with a documented, distinct epitope and compare cellular patterns, particularly because the tissue profile carries a multi-gene targeting and presumed off-target caution (HPA: Supported reliability description).
How should I adapt the IHC findings for multiplex HSD3B1 immunofluorescence?
Treat IF as a separate validation experiment: the supplied EDTA pH 8.0 retrieval and 1 μg/ml primary condition document paraffin-section chromogenic IHC, not an IF protocol (caption A02856-2). Pair HSD3B1 with a validated syncytiotrophoblast marker and inspect whether their signals occur in the expected cells, using placental sections as a reference (HPA: High in placental syncytiotrophoblasts; standard IF practice). Select spectrally separated fluorophores, favoring a longer-wavelength channel when tissue autofluorescence interferes, and include single-label and no-primary controls (standard IF practice). Because the epitope's membrane-facing side is unspecified, titrate permeabilisation and compare signal distribution with the expected endoplasmic reticulum or mitochondrial pattern (UniProt P14060 subcellular location and topology; standard IF practice).
How do I troubleshoot diffuse brown staining in placental sections?
First check a no-primary slide, tissue edges and areas with poor morphology; these reveal detection background and processing artifacts that can resemble weak HSD3B1 signal (standard IHC practice). The catalog example used 10% goat serum, a biotinylated goat anti-rabbit secondary, a streptavidin-biotin complex and DAB, so assess both nonspecific binding and biotin-related background in that workflow (caption A02856-2; standard IHC practice). Use an appropriate endogenous-peroxidase block, verify blocking and washing, and shorten DAB development if the whole section darkens (standard chromogenic IHC practice). Retain a placental positive control while optimizing: genuine interpretation requires the expected cellular and cytoplasmic pattern, particularly given the reported off-target caution (HPA: placental syncytiotrophoblasts and cytoplasmic expression; HPA: reliability description).
What should I quantify when comparing HSD3B1 across paraffin sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the scored compartment and cell population before reading slides: placental syncytiotrophoblast cytoplasm is an evidence-supported reference, while other populations need their own validation (HPA: High in placental syncytiotrophoblasts; HPA: cytoplasmic expression). For chromogenic IHC, report the percentage of positive target cells and an intensity-weighted H-score, with fixed scoring thresholds and observer blinding where feasible (standard IHC practice). Normalize counts to the number of eligible, intact target cells, or report positive-cell density per mm² of annotated viable tissue when cell counting is impractical (standard IHC practice). Keep retrieval, primary concentration, DAB development and image settings consistent, and record excluded necrotic or edge regions (caption A02856-2; standard IHC practice).
How can I distinguish genuine HSD3B1 positivity from staining artifacts?
A convincing placental result places staining in syncytiotrophoblasts with a predominantly cytoplasmic pattern, matching the tissue evidence and membrane-associated biology (HPA: High in placental syncytiotrophoblasts and cytoplasmic expression; UniProt P14060 subcellular location). Question isolated nuclear or nucleolar staining because that reported location is uncertain and the underlying antibodies may target proteins from multiple genes (HPA subcellular: nucleoli uncertain; HPA subcellular caution). Exclude section edges, necrotic areas and diffusely brown no-primary controls before scoring, and check endogenous-peroxidase blocking when DAB background is widespread (standard chromogenic IHC practice). Concordance with a second, independently validated antibody strengthens attribution, especially because presumed off-target binding was observed in the tissue-profile assessment (HPA: Supported reliability description).
Boster reagents
Best HSD3B1 / 3 beta-hydroxysteroid dehydrogenase/Delta 5-->4-isomerase type 1 IHC Antibodies
Two anti-HSD3B1 antibodies have image data: human placenta paraffin-section IHC for A02856-2 and cell-based IF for A02856-2 and M02856 (catalog image captions).
Real IHC data
Anti-HSD3B1 Antibody ®
Cat # A02856-2
Real IF data
Anti-HSD3B1 Antibody ® (monoclonal, 3F3C/7F9C8)
Cat # M02856
A02856-2 will render with IHC staining of a human placenta paraffin section (A02856-2 image caption; catalog: Human reactivity). M02856 will render with IF staining of U20S cells (M02856 image caption; catalog: IF/ICC applications).
Which to pick: For tissue IHC, choose A02856-2: its rabbit antibody has a paraffin-section IHC image and a listed working range of 0.5–1 μg/ml (catalog: host, applications and IHC dilution; A02856-2 image caption). For IF/ICC, either SKU has cell-based IF image data; M02856 is the mouse monoclonal choice when catalog-listed Human, Mouse and Rat reactivity matters, though its pictured IF experiment is in U20S cells (catalog: clone, reactivity and applications; M02856 image caption). The A02856-2 IHC caption reports a paraffin section but does not report its fixative (A02856-2 image caption).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P14060 (3BHS1_HUMAN, 3 beta-hydroxysteroid dehydrogenase/Delta 5-->4-isomerase type 1).
- Human Protein Atlas. HSD3B1 tissue IHC expression (reliability: Supported).
- Human Protein Atlas. HSD3B1 subcellular location (ICC-IF): Mainly localized to the nucleoli, endoplasmic reticulum and primary cilium. In addition localized to the microtubules, cytokinetic bridge and primary cilium transition zone. Caution: Based on antibodies targeting proteins from multiple genes..
- Human Protein Atlas. HSD3B1 antibody validation summary (3 antibodies).
- Effect of selenium-enriched Ganoderma lucidum powder in testicular tissues, oxidative stress biomarkers and antiapoptotic capacities in roosters. Poultry science 2026 — PMC12723041.
- 3β-Hydroxysteroid dehydrogenase isoforms in human aldosterone-producing adenoma. Molecular and cellular endocrinology 2015 — PMC4821076.
- A Short-Term Exposure to Tributyltin Blocks Leydig Cell Regeneration in the Adult Rat Testis. Frontiers in pharmacology 2017 — PMC5643909.
- Immunohistochemical evaluation of Sertoli and Leydig cells of scotch cat testis (Felis catus) at diverse ages. Open veterinary journal 2026 — PMC13313884.
- PubMed PMID:2779585 — UniProt-cited evidence.
- PubMed PMID:2139411 — UniProt-cited evidence.
- PubMed PMID:2082186 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for HSD3B1 (3 beta-hydroxysteroid dehydrogenase/Delta 5-->4-isomerase type 1). Expected localisation Cytoplasmic in placenta and adrenal gland (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A02856-2). Validated IHC antibody: A02856-2 at 0.5-1μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Placenta (Syncytiotrophoblasts); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human placenta tissue; fixative not specified (datasheet A02856-2); 2. Fixation: Image fixative and duration unreported (datasheet A02856-2); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: EDTA pH 8.0 (datasheet A02856-2); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A02856-2); 8. Primary antibody: Rabbit anti-HSD3B1, 0.5-1μg/ml (datasheet A02856-2); 9. Primary incubation: Overnight at 4 °C (datasheet A02856-2). Watch for: Fixation: Keep fixation consistent across sections. (standard IHC practice; not target-specific); Caveat: Cross-reactivity possible: antibody targets more than one gene (HPA tissue IHC); Isoform / epitope: No isoforms annotated; epitope sidedness unassigned (UniProt); Regulation: Placenta high; adrenal zona fasciculata low (HPA tissue IHC). Evidence: UniProt P14060, HPA tissue IHC · Supported, HPA ICC-IF · Mainly localized to the nucleoli, endoplasmic reticulum and primary…, Datasheet A02856-2, PMC4821076, PMC5643909. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/HSD3B1/immunohistochemistry-ihc-guide.