This website uses cookies to ensure you get the best experience on our website.
- Table of Contents
PI16 / Peptidase inhibitor 16 · IHC design guide
Design Immunohistochemistry for PI16
Plan PI16 chromogenic IHC in paraffin sections around extracellular staining, mainly in prostate, urinary bladder and testis (HPA tissue IHC). Start with the IHC-validated antibody at 2–5 μg/ml (datasheet A09887-1), and interpret cellular staining in light of PI16 secretion (UniProt).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
PI16 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 | Extracellular staining in tissues (HPA tissue IHC) | |
| Staining pattern | Extracellular signal mainly in prostate, bladder and testis (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A09887-1) | |
| Positive control | Fallopian tube+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A09887-1) | |
| Caveat | Secreted PI16 may stain away from producing cells (HPA tissue IHC) | |
| Regulation | Staining varies across tissues (HPA tissue IHC) | |
| Isoform / epitope | Two isoforms; epitope coverage is unspecified (UniProt) |
Section 1
Recommended PI16 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by two published chromogenic PI16 protocols for paraffin sections (PMC7541153; PMC7616500).
| Sample | Paraffin-embedded mouse intestine tissue; fixative not specified (datasheet A09887-1) |
| Fixation | Image fixative and duration unreported (datasheet A09887-1); 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 A09887-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A09887-1) |
| Primary antibody | Rabbit anti-PI16, 2-5μg/ml (datasheet A09887-1) |
| Primary incubation | Overnight at 4 °C (datasheet A09887-1) |
| Detection | Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A09887-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | PI16-positive staining in ciliated cells (ciliary rootlets) of fallopian tube (HPA tissue IHC: High). HPA tissue profile: Extracellular positivity in several tissues, mainly in prostate, urinary bladder and testis. Additional cytoplasmic or membranous expression in a few tissues including retina. No signal in the no-primary control. |
💡Decision noteStart with heat-mediated EDTA pH 8.0 retrieval for the catalog antibody (datasheet: A09887-1); a published limb protocol uses boiled citrate pH 6.0 (PMC7616500).
Section 2
What Is the Expected PI16 Staining Pattern?
PI16 is secreted and has no transmembrane segment (UniProt Q6UXB8 topology). In paraffin sections, expect mainly extracellular staining, particularly in prostate, urinary bladder and testis (HPA: Approved tissue IHC). HPA also records staining at ciliary rootlets in fallopian tube cells and in testis peritubular cells (HPA: High and Medium, respectively). Interpret cell boundaries cautiously because secreted protein can appear away from its producing cells (HPA: reliability description).
| Extracellular signal in prostate, urinary bladder or testis, with tissue structures still discernible. | This fits the predominant reported distribution (HPA: Approved tissue IHC). Extracellular deposits need not identify the cells that produced PI16: it is secreted, and HPA warns that RNA and protein locations may differ (UniProt Q6UXB8; HPA: reliability description). |
| Strong staining at fallopian tube ciliary rootlets, or moderate staining of testis peritubular cells. | These are specifically annotated positives (HPA: High in fallopian tube ciliated-cell rootlets; Medium in testis peritubular cells). Do not dismiss every cell-associated signal as artefact: HPA also reports limited cytoplasmic or membranous staining in some tissues (HPA: tissue profile). |
| Dominant nuclear staining, especially where the expected extracellular or rootlet pattern is absent. | A nuclear pattern is outside the reported PI16 locations (UniProt Q6UXB8: secreted; HPA: tissue and subcellular profiles). Treat it as suspect and review negative controls, counterstain, and whether the apparent signal follows recognizable tissue structures (general IHC practice). |
| Prominent staining in cells annotated as negative, such as bone-marrow hematopoietic cells or adipocytes. | That conflicts with these specific HPA annotations (HPA: Not detected in bone-marrow hematopoietic cells and adipocytes). Check for cross-reactivity or endogenous detection activity before assigning PI16 expression; an annotated negative cell population does not establish that its entire tissue is PI16-free (general IHC practice; HPA: tissue annotations). |
| Diffuse brown haze across positive and negative areas, or no signal in a known-positive structure. | Neither pattern alone establishes PI16 distribution. Haze may reflect background from blocking, detection or counterstain; absent signal may reflect a failed staining run (general IHC practice). Compare a positive structure and an annotated negative cell population on the same run before interpreting biology (HPA: tissue annotations). |
💡Expected PI16 appearanceCall a result positive when the chromogenic signal follows mainly extracellular structures in HPA-reported tissues, or the High fallopian tube ciliary-rootlet or Medium testis peritubular pattern; dominant nuclear staining or uniform haze is suspect (HPA: tissue IHC; UniProt Q6UXB8: secreted).
| Secretion and processing | PI16 has a signal peptide at residues 1–27 and a mature chain at 28–463, with no transmembrane segment (UniProt Q6UXB8). Extracellular staining can therefore extend beyond a producing cell; cell-associated exceptions must be judged against the HPA images (HPA: tissue profile). |
| Antibody validation | The listed antibody, HPA076574, is Approved for IHC; the supplied record does not mark it Enhanced (HPA: antibody record). Use its reported pattern as a comparison, while treating unexpected staining as a finding to verify rather than confirmed PI16 (HPA: tissue IHC). |
| Regional variation | HPA reports High fallopian tube rootlet staining, Medium testis peritubular staining, and Low staining in several glandular or tubular populations (HPA: tissue IHC). Score the named cell or structure, rather than assigning one intensity to the whole organ (HPA: tissue annotations). |
| Processing, variants and epitope | UniProt lists 2 isoforms and glycosylation sites at 114, 403 and 409 (UniProt Q6UXB8). The supplied sources give no antibody epitope or PI16-specific antigen-retrieval result; they cannot predict which variant or processed form this stain detects. |
| IF/ICC: what pattern is established? | HPA summarizes PI16 as membrane-associated and secreted but supplies no ICC-IF image or main-location annotation (HPA: subcellular record). An IF/ICC pattern therefore cannot be confirmed from this record; assess that application using its separate guide (HPA: subcellular record). |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining at a fallopian tube ciliary-rootlet positive control. | The expected High structure is missing (HPA: fallopian tube tissue IHC); the run or tissue preparation may need review (general IHC practice). | Confirm the section contains identifiable ciliated rootlets, then review the IHC antibody and detection steps against the run controls (HPA: fallopian tube annotation; general IHC practice). |
| Prostate or testis shows little extracellular signal, but some cells stain. | Cellular signal alone may not reproduce the predominant extracellular profile (HPA: tissue IHC); secreted PI16 can be spatially separated from producing cells (UniProt Q6UXB8; HPA: reliability description). | Examine preserved extracellular structures and compare the same run with an annotated positive structure; do not infer production from a stained cell alone (HPA: tissue IHC; general IHC practice). |
| Brown staining is widespread, including annotated negative cells. | This may reflect nonspecific binding or endogenous detection activity (general IHC practice); HPA lists adipocytes and bone-marrow hematopoietic cells as Not detected (HPA: tissue IHC). | Review appropriate reagent-omission and detection controls, blocking, and wash steps; reassess whether signal tracks the HPA-described compartments (general IHC practice; HPA: tissue profile). |
| A strong nuclear pattern dominates the section. | That compartment does not match the secreted PI16 annotation or HPA's reported tissue pattern (UniProt Q6UXB8; HPA: tissue and subcellular profiles). | Check counterstain and detection controls, then compare the pattern with positive and annotated negative structures before scoring it as PI16 (general IHC practice; HPA: tissue annotations). |
| A low-staining glandular population appears negative. | A weak expected signal can be hard to distinguish from the run background (general IHC practice); HPA rates endometrial glandular cells Low (HPA: endometrium tissue IHC). | First verify a stronger annotated positive in the run; score the glandular population only when localized signal is distinguishable from background (HPA: tissue IHC; general IHC practice). |
| Changing antigen retrieval seems to change the result. | The supplied UniProt and HPA records do not establish PI16-specific retrieval or fixation sensitivity; a change in staining alone cannot identify its mechanism. | Compare retrieval conditions with matched positive and negative controls and preserved morphology, and report the condition used without attributing the difference to PI16 epitope masking (general IHC practice). |
Experiment evidence
IHC & IF images showing PI16 staining
A09887-1 · IHCIHC analysis of PI-16/PI16 using anti-PI-16/PI16 antibody (A09887-1). PI-16/PI16 was detected in paraffin-embedded section of mouse intestin
A09887-1 · IHCIHC analysis of PI-16/PI16 using anti-PI-16/PI16 antibody (A09887-1). PI-16/PI16 was detected in paraffin-embedded section of rat intestine
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000164530-PI16/tissue/cerebral+cortex#imid_20500275
HPA · RetinaHuman Protein Atlas IHC, Retina. Source: https://www.proteinatlas.org/ENSG00000164530-PI16/tissue/retina#imid_1104
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000164530-PI16/tissue/colon#imid_20500201
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000164530-PI16/tissue/liver#imid_20500210
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for PI16 IHC & IF
🧪Run fallopian tube first and look for staining in ciliated cells at the ciliary rootlets (HPA: High in fallopian tube ciliated cells). Use adipose tissue adipocytes as a negative reference (HPA: Not detected in adipocytes); on the positive slide, assess background in neighboring cells outside the ciliary-rootlet staining pattern without assuming those cells are PI16-negative.
Positive control tissue: Fallopian tube (Ciliated cells (ciliary rootlets), 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 PI16; derive a cell-line control from the positive tissue's cell type (Ciliated cells (ciliary rootlets)) and confirm it by RNA or western blot first.
Technical controls: Include no-primary (secondary-only) and concentration-matched rabbit IgG isotype controls alongside a PI16-knockout tissue control if available (selected IHC caption: rabbit primary antibody). For chromogenic detection, control endogenous peroxidase and assess endogenous biotin because the selected IHC caption uses a biotinylated secondary, streptavidin–biotin complex and DAB (selected IHC caption: detection method).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the exact selected-SKU tissue-IHC caption does not state a fixative (selected IHC caption: fixative not stated). The demonstrated paraffin-section procedure used heat retrieval in EDTA at pH 8.0; a comparison with frozen sections or IF is unreported (selected IHC caption: retrieval; HPA subcellular: no ICC-IF image cell lines listed). In fallopian tube, score the specified ciliary-rootlet pattern and check for nonspecific staining with the controls, especially when using biotin-based DAB detection (HPA: High in ciliated cells at ciliary rootlets; selected IHC caption: biotin-based DAB detection).
HPA tissue IHC evidence for PI16
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — At least one protein variant secreted, tissue location of RNA and protein might differ and correlation is complex. Antibody staining in cells/structures not annotated, view images.). 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 | Ciliated cells (ciliary rootlets) | High | Protein (IHC) | HPA → |
| Bronchus | Ciliated cells (ciliary rootlets) | Medium | Protein (IHC) | HPA → |
| Endometrium | Glandular cells | Low | Protein (IHC) | HPA → |
| Epididymis | Glandular cells | Medium | Protein (IHC) | HPA → |
| Retina | Nerve fiber layer | 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 → |
| Caudate | Glial cells | Not detected | Protein (IHC) | HPA → |
| Cerebellum | Cells in granular layer | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced PI16 IHC Tips
Use compartment, tissue context and matched controls to troubleshoot PI16 staining in paraffin sections; the antibody’s tissue caption does not identify a fixative.
Which retrieval condition should I start with for weak PI16 staining?
Start with heat-mediated retrieval in EDTA pH 8.0 (datasheet A09887-1). The selected paraffin-section example used this condition before 10% goat-serum blocking and 2 µg/ml primary antibody overnight at 4°C (caption A09887-1). If staining is weak, compare retrieval duration on matched sections while keeping detection and exposure constant; excessive heating can damage morphology (standard IHC practice). Assess both extracellular signal and tissue structure, because PI16 is secreted (UniProt Q6UXB8 subcellular location) and extracellular positivity is reported across several tissues (HPA: tissue IHC profile); a gain in diffuse background alone does not establish better retrieval.
How should I troubleshoot fixation when PI16 signal varies between blocks?
PI16-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state its fixative (caption A09887-1). Record the fixative, fixation duration and processing history for each block, then compare matched sections using the same EDTA pH 8.0 retrieval and staining run (datasheet A09887-1; standard IHC practice). Check whether weak signal tracks a processing batch, tissue damage or loss of morphology before changing antibody concentration (standard IHC practice). Keep the 2 µg/ml overnight primary incubation as a documented starting point (caption A09887-1), but do not assign a PI16-specific fixation effect without a controlled comparison.
Where should convincing PI16 staining appear in tissue sections?
Interpret staining in its tissue context: PI16 is secreted and has no transmembrane segment (UniProt Q6UXB8 subcellular location and topology). Extracellular positivity is reported in several tissues, mainly prostate, urinary bladder and testis, with cytoplasmic or membranous expression in a few tissues (HPA: tissue IHC profile). In testis, assess the reported medium peritubular-cell staining against surrounding extracellular signal (HPA: testis tissue IHC). Do not require every positive structure to show a crisp cell boundary; compare anatomical distribution, morphology and a matched negative control before attributing intracellular or diffuse staining to PI16 (standard IHC practice).
Could PI16 isoforms or processing explain inconsistent staining?
PI16 has 2 listed isoforms, a signal peptide at residues 1–27, and a mature chain spanning 28–463 (UniProt Q6UXB8 isoforms and processing). Its SCP domain spans residues 37–165, with glycosylation sites at 114, 403 and 409 (UniProt Q6UXB8 domains and glycosylation). The supplied caption does not identify the antibody epitope or establish isoform-specific detection (caption A09887-1). When sections disagree, request the mapped immunogen or epitope and compare anatomically matched samples under identical retrieval conditions; neither glycosylation nor splicing alone proves why a section stained weakly (standard IHC interpretation).
How can I examine PI16 by multiplex immunofluorescence?
Treat IF as a separate assay: the supplied A09887-1 example documents chromogenic paraffin-section IHC, without an IF result or a stated fixative (caption A09887-1). Pair PI16 with a validated marker for the expected compartment or cell population, such as peritubular cells in testis, and confirm the marker’s specificity independently (HPA: testis tissue IHC; standard IF practice). Select spectrally separated fluorophores, favour a far-red PI16 channel when tissue autofluorescence obscures shorter wavelengths, and include single-stain and unstained controls (standard IF practice). Because PI16 is secreted with no transmembrane segment, choose permeabilisation only after establishing whether the antibody’s epitope is extracellular or intracellular; its side is not supplied (UniProt Q6UXB8 topology; standard IF practice).
How do I distinguish PI16 signal from chromogenic background?
Inspect a no-primary control alongside the PI16 section to identify background from the secondary and detection system (standard IHC practice). The documented tissue example used biotinylated anti-rabbit secondary, a streptavidin–biotin complex and DAB, so assess endogenous biotin and peroxidase contributions when using that workflow (caption A09887-1; standard IHC practice). Apply an appropriate peroxidase block, limit DAB development and titrate primary antibody from the documented 2 µg/ml condition if diffuse staining persists (caption A09887-1; standard IHC practice). Preserve plausible extracellular signal while judging background: PI16 is secreted, and extracellular tissue positivity is reported (UniProt Q6UXB8 subcellular location; HPA: tissue IHC profile).
How should I score PI16 when much of the stain is extracellular? ⚠ ANSWER MARKED FOR VERIFICATION
Define anatomical regions and exclude folds, necrosis and section edges before scoring matched slides (standard IHC practice). For extracellular PI16, measure positive stained area or optical-density signal per mm² of viable tissue, using the same threshold and DAB acquisition settings across groups (UniProt Q6UXB8 subcellular location; HPA: tissue IHC profile; standard IHC practice). If scoring a specified cell population, report % positive cells or an H-score and normalise to eligible cells in that compartment (standard IHC practice). Keep extracellular and cellular scores separate, and record background from a no-primary control so changing tissue composition does not masquerade as altered PI16 abundance (standard IHC practice).
What makes a PI16-positive section credible rather than artefactual?
A credible result has reproducible anatomical signal with preserved morphology and low no-primary background (standard IHC practice). For PI16, extracellular staining fits the reported tissue profile, while medium peritubular-cell staining in testis offers a specific cellular comparison (HPA: tissue IHC profile; HPA: testis tissue IHC). Treat isolated nuclear staining, staining confined to cut edges or necrotic regions, and DAB deposition that survives omission of primary antibody as suspect (UniProt Q6UXB8 subcellular location; standard IHC practice). Review morphology and controls before calling a discordant compartment positive, since the reported secreted protein can occupy extracellular space beyond the cells producing it (UniProt Q6UXB8 subcellular location; standard IHC interpretation).
Boster reagents
Best PI16 / Peptidase inhibitor 16 IHC Antibodies
The catalog includes one anti-PI16 antibody with IHC images from paraffin sections of mouse and rat intestine and human bladder and pancreatic cancer tissue (catalog IHC captions).
Real IHC data
Anti-PI-16/PI16 Antibody ®
Cat # A09887-1
A09887-1 is listed for IHC and reactivity with human, mouse, rat and monkey (catalog applications; catalog reactivity). Its images document IHC in mouse and rat intestine and human bladder and pancreatic cancer paraffin sections; no IF image is supplied (catalog IHC captions; catalog IF images).
Which to pick: For tissue IHC, choose A09887-1 for the paraffin-section samples documented in its own captions; the fixative is unreported (catalog IHC captions). No listed SKU has IF/ICC validation or an IF image (catalog applications; catalog IF images). For cross-species IHC, A09887-1 has documented human, mouse and rat IHC examples; monkey is listed as reactive but has no monkey IHC image (catalog IHC captions; catalog reactivity).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry Q6UXB8 (PI16_HUMAN, Peptidase inhibitor 16).
- Human Protein Atlas. PI16 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. PI16 subcellular location (ICC-IF): Membrane, Secreted.
- Human Protein Atlas. PI16 antibody validation summary (1 antibodies).
- PI16 attenuates response to sorafenib and represents a predictive biomarker in hepatocellular carcinoma. Cancer medicine 2020 — PMC7541153.
- PI16 is a shear stress and inflammation-regulated inhibitor of MMP2. Scientific reports 2016 — PMC5171773.
- Single cell approaches to address adipose tissue stromal cell heterogeneity. The Biochemical journal 2020 — PMC7875330.
- A human embryonic limb cell atlas resolved in space and time. Nature 2024 — PMC7616500.
- PubMed PMID:12975309 — UniProt-cited evidence.
- PubMed PMID:16303743 — UniProt-cited evidence.
- PubMed PMID:14702039 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for PI16 (Peptidase inhibitor 16). Expected localisation Extracellular staining in tissues (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A09887-1). Validated IHC antibody: A09887-1 at 2-5μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Fallopian tube (Ciliated cells (ciliary rootlets)); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded mouse intestine tissue; fixative not specified (datasheet A09887-1); 2. Fixation: Image fixative and duration unreported (datasheet A09887-1); 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 A09887-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A09887-1); 8. Primary antibody: Rabbit anti-PI16, 2-5μg/ml (datasheet A09887-1); 9. Primary incubation: Overnight at 4 °C (datasheet A09887-1). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A09887-1); Caveat: Secreted PI16 may stain away from producing cells (HPA tissue IHC); Isoform / epitope: Two isoforms; epitope coverage is unspecified (UniProt); Regulation: Staining varies across tissues (HPA tissue IHC). Evidence: UniProt Q6UXB8, HPA tissue IHC · Approved, HPA ICC-IF · Membrane, Secreted, Datasheet A09887-1, PMC7541153, PMC7616500. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/PI16/immunohistochemistry-ihc-guide.