RAB3IP / Rab-3A-interacting protein · IHC design guide

Design Immunohistochemistry for RAB3IP

Plan paraffin-section RAB3IP IHC around the reported cytoplasmic tissue pattern (HPA tissue IHC). Compare staining with appropriate controls because the tissue IHC profile has uncertain reliability (HPA tissue IHC).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
Immunohistochemistry protocol sheet for RAB3IP (IHC for RAB3IP): expected localisation Predominantly cytoplasmic tissue staining (HPA tissue IHC), antibody A09085-3, validated IHC image, and IHC protocol steps
Printable RAB3IP IHC protocol sheet — expected localisation Predominantly cytoplasmic tissue staining (HPA tissue IHC), antibody A09085-3, controls and protocol steps. Open the full RAB3IP IHC guide →

RAB3IP 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 Predominantly cytoplasmic tissue staining (HPA tissue IHC)
Staining pattern Cytoplasmic staining across several tissue types (HPA tissue IHC)
Antigen retrieval EDTA pH 8.0 HIER, heat-mediated (datasheet A09085-3)
Positive control ⓘ Appendix+4 more · see all
Negative control ⓘ Adipose tissue+4 more · see all
Important caveats
Reasons your staining may differ from the expected pattern.
Fixation Keep fixation consistent across sections (standard IHC practice; not target-specific). Selected-image fixative and duration unreported (datasheet A09085-3); verify before use.
Caveat Low consistency between IHC staining and RNA expression (HPA tissue IHC)
Regulation PKC redistributes RAB3IP to lamellipodia (UniProt)
Isoform / epitope 8 isoforms; check epitope coverage (UniProt)
Section 1

Recommended RAB3IP IHC & IF Protocols

The catalog antibody’s IHC-P protocol (datasheet A09085-3) is paired with a published RAB3IP IHC protocol for human gastric cancer tissue (PMC6277663).

Recommended immunohistochemistry (IHC-P) protocol parameters
SampleParaffin-embedded human ovarian serous adenocarcinoma tissue; fixative not specified (datasheet A09085-3)
FixationImage fixative and duration unreported (datasheet A09085-3); verify before use.
Sectioning4–5 µm sections on charged slides (standard)
DeparaffinisationXylene, graded ethanol series to water (standard)
Antigen retrievalHeat retrieval: EDTA pH 8.0 (datasheet A09085-3); 20 min, 95–100 °C (standard)
Peroxidase block3% H2O2, 10 min, room temperature (standard)
Blocking10% goat serum (datasheet A09085-3)
Primary antibodyRabbit anti-RAB3IP, 2-5 μg/ml (datasheet A09085-3)
Primary incubationOvernight at 4 °C (datasheet A09085-3)
DetectionHRP-conjugated secondary, DAB chromogen (datasheet A09085-3)
CounterstainHematoxylin, blue, dehydrate and mount (standard)
Expected resultRAB3IP-positive staining in glandular cells of appendix (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in several different tissue types. No signal in the no-primary control.
💡Decision noteStart with heat-mediated EDTA retrieval at pH 8.0 (datasheet A09085-3); the published protocol does not specify retrieval (PMC6277663).
Section 2

What Is the Expected RAB3IP Staining Pattern?

In paraffin-section IHC, expect predominantly cytoplasmic RAB3IP staining in selected glandular cells and kidney tubule cells (HPA: tissue IHC; UniProt Q96QF0: predominantly cytoplasmic). RAB3IP has no transmembrane segment (UniProt Q96QF0: topology). Treat this as a provisional pattern: HPA rates its tissue IHC reliability Uncertain because antibody staining has low consistency with RNA expression data and awaits external verification (HPA: tissue IHC reliability).

What am I looking at on my slide?
Cytoplasmic staining in appendix, breast, or endometrial glandular cells, or kidney tubule cells.This matches cell populations scored High by HPA and its broader cytoplasmic tissue profile (HPA: tissue IHC). Judge the signal by the named cell type, not by staining anywhere in the organ. HPA's Uncertain reliability means agreement is supportive, not definitive proof of specificity (HPA: tissue IHC reliability).
Predominantly sharp plasma-membrane or extracellular staining, without convincing cytoplasmic staining.This is discordant with the expected tissue pattern (HPA: cytoplasmic tissue expression) and with a protein lacking a transmembrane segment (UniProt Q96QF0: topology). Consider nonspecific deposition or detection artefact. A small nuclear component alone is not disqualifying: UniProt describes predominantly cytoplasmic RAB3IP with a small nuclear proportion (UniProt Q96QF0: subcellular location).
Strong staining mainly in a cell population HPA scores as not detected, such as adipocytes.Adipocytes are scored Not detected (HPA: adipose-tissue IHC). Cross-reactivity or endogenous detection activity is possible; compare staining with a no-primary control and the expected positive cell population. A negative HPA score is a comparison point, not absolute proof of absence, given the uncertain tissue IHC reliability (HPA: tissue IHC reliability).
Diffuse chromogen across cells, stroma, or section edges, obscuring cell boundaries.An uninterpretable distribution cannot establish RAB3IP localization. Check no-primary and detection-only controls for nonspecific or endogenous detection activity, then reassess blocking and washing as general IHC practice. The expected tissue profile is cellular and cytoplasmic (HPA: tissue IHC); diffuse background should not be scored as target-positive.
No convincing signal in appendix glandular cells or kidney tubule cells.Both populations are scored High (HPA: appendix and kidney IHC), so an absent signal warrants a run check. Confirm tissue preservation, control performance, the antibody's IHC-P instructions, and detection reagents before interpreting the specimen as negative. HPA's Uncertain reliability limits any single tissue's value as a definitive positive control (HPA: tissue IHC reliability).
💡Expected RAB3IP appearanceCall a result provisionally positive when selected glandular cells or kidney tubule cells show clear, predominantly cytoplasmic staining consistent with HPA's High scores; broad extracellular or membrane-only color is suspect, and HPA rates the tissue pattern Uncertain (HPA: tissue IHC and reliability; UniProt Q96QF0: topology).
How each factor affects the staining
Tissue and cell selectionHPA scores glandular cells in appendix, breast, endometrium, epididymis, fallopian tube, and gallbladder, kidney tubule cells, and liver cholangiocytes as High (HPA: tissue IHC). It scores adipocytes and heart cardiomyocytes as Not detected (HPA: tissue IHC). Select the specific cell population when comparing slides; an organ-wide positive or negative label loses that distinction.
Source disagreement and IHC confidenceUniProt lists expression in heart and says RAB3IP is not detected in liver (UniProt Q96QF0: tissue specificity), while HPA scores cardiomyocytes Not detected and liver cholangiocytes High by IHC (HPA: tissue IHC). These statements concern different evidence and cell populations. Record the observed cell type and avoid resolving the discrepancy by assumption; HPA rates tissue IHC Uncertain (HPA: tissue IHC reliability).
Subcellular interpretationUniProt describes cytoplasmic, cytoskeletal, vesicular, centrosomal, lamellipodial, and minor nuclear localization (UniProt Q96QF0: subcellular location). HPA's tissue IHC summary is cytoplasmic (HPA: tissue IHC), so that is the principal paraffin-section comparison. Fine structures may be difficult to resolve in chromogenic sections; do not require a centrosomal or vesicular pattern for an IHC positive call.
Antibody evidenceHPA lists tissue IHC for HPA039794 as Uncertain and ICC for HPA057728 as Supported (HPA: antibody validation). Those ratings apply to different antibodies and applications. ICC support does not independently validate an IHC-P staining pattern. No target-specific fixation sensitivity or antigen-retrieval effect is supplied by UniProt or HPA; follow the IHC-validated antibody's documented workflow without claiming a RAB3IP-specific retrieval requirement.
IF/ICC Q&A: should the IHC compartment rule carry over?No single compartment rule should be transferred. HPA reports mainly nucleoplasmic staining, with additional cytosol and centrosome localization, in ICC-IF (HPA: subcellular ICC-IF); its tissue IHC summary is cytoplasmic (HPA: tissue IHC). Use the separate IF/ICC guide for that application. The HPA ICC rating is Supported for HPA057728 (HPA: antibody validation).
Why is my staining missing, weak or wrong?
SituationLikely causeNext action
Expected glandular or tubule cells are blank.The run may have insufficient detection, or the sampled cells may be absent; HPA scores these specified populations High (HPA: tissue IHC).Locate the named cells on a counterstained section, verify a run control, and check the IHC-validated antibody's instructions and detection reagents. Treat a persistent negative cautiously because HPA tissue IHC reliability is Uncertain (HPA: tissue IHC reliability).
Most structures, including stroma, carry similar color.Diffuse background may reflect nonspecific reagent binding or endogenous detection activity; it does not match HPA's cellular cytoplasmic summary (HPA: tissue IHC).Compare no-primary and detection-only controls. Reassess blocking, washes, and detection chemistry as general IHC practice, then score only signal that remains cell-associated and interpretable.
Only membrane rims or extracellular deposits stain.That distribution conflicts with cytoplasmic tissue IHC and the lack of a transmembrane segment (HPA: tissue IHC; UniProt Q96QF0: topology).Inspect the no-primary control and compare a specified HPA-positive cell population. Do not assign extracellular or membrane-only color to RAB3IP without independent support.
Nuclear staining accompanies cytoplasmic staining.A small nuclear fraction is described by UniProt (UniProt Q96QF0: subcellular location), while HPA tissue IHC emphasizes cytoplasm (HPA: tissue IHC).Record both compartments and the cell type. Check controls if nuclear staining dominates; do not automatically reject a limited nuclear component or import the mainly nucleoplasmic ICC-IF result as the paraffin IHC standard (HPA: subcellular ICC-IF).
Liver cholangiocytes stain, but a broad liver expectation suggests negativity.HPA scores cholangiocytes High by IHC (HPA: liver IHC), whereas UniProt says liver expression was not detected (UniProt Q96QF0: tissue specificity).Document that the signal is in cholangiocytes, compare controls, and report the source discrepancy. Avoid calling either result a universal liver pattern; HPA tissue IHC remains Uncertain (HPA: tissue IHC reliability).
A strongly stained cell population lies outside the expected HPA pattern.Cross-reactivity or endogenous detection activity is possible; HPA's tissue profile is cell-type specific and rated Uncertain (HPA: tissue IHC and reliability).Identify the stained cell type, compare no-primary and detection-only controls, and inspect a specified HPA-positive population in the same run. Report an unresolved signal as unverified rather than scoring it as RAB3IP.

Sample controls for RAB3IP IHC & IF

🧪Run appendix first: its glandular cells are High for RAB3IP (HPA: Appendix, glandular cells, High). Use adipose tissue as the negative tissue, where adipocytes are Not detected (HPA: Adipose tissue, adipocytes, Not detected); on the appendix slide, use only independently verified RAB3IP-negative cells as internal controls and expect background-level DAB staining in them.
Positive control tissue: Appendix (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show RAB3IP in A-431, CACO-2, U2OS, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit IgG isotype control, alongside a RAB3IP knockout biological negative where available (caption: rabbit primary antibody; standard IHC practice). Check endogenous peroxidase activity in appendix inflammatory cells and quench it before HRP/DAB detection (standard IHC practice).
⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A09085-3 paraffin-section caption does not state a fixative (A09085-3 caption: fixative unreported). Start IHC-P optimization with heat-mediated EDTA retrieval at pH 8.0, as used in the caption; whether retrieval is required is unreported (A09085-3 caption: EDTA pH 8.0). Frozen-section performance is unreported, while ICC-IF images exist for A-431, CACO-2 and U2OS; these data do not establish that frozen sections or IF are easier, and endogenous peroxidase in appendix inflammatory cells can complicate HRP/DAB interpretation (HPA: ICC-IF image cell lines; standard IHC practice).

HPA tissue IHC evidence for RAB3IP

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Uncertain — Low consistency between antibody staining and RNA expression data. Pending external verification.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.

Positive expression · recommended positive controls

TissueCell typeLevelEvidenceSource
Appendix Glandular cells High Protein (IHC) HPA →
Breast Glandular cells High Protein (IHC) HPA →
Endometrium Glandular cells High Protein (IHC) HPA →
Epididymis Glandular cells High Protein (IHC) HPA →
Fallopian tube Glandular cells High Protein (IHC) HPA →

Undetected expression · recommended negative controls

TissueCell typeLevelEvidenceSource
Adipose tissue Adipocytes Not detected Protein (IHC) HPA →
Adrenal gland 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 RAB3IP IHC Tips

Troubleshoot RAB3IP staining in paraffin sections by checking retrieval, cellular compartment, tissue context and controls before scoring chromogenic signal.

What retrieval should I try first when RAB3IP staining is weak in paraffin sections?
Start with heat-mediated retrieval in EDTA at pH 8.0 for this catalog antibody (datasheet A09085-3). The selected paraffin-section example used that retrieval before overnight primary incubation at 4°C, but it does not establish an optimal heating duration (datasheet A09085-3). If staining remains weak, compare a modestly adjusted heating duration on adjacent sections while holding antibody concentration, detection and DAB development constant (standard IHC practice). Include a tissue control and assess whether stronger signal preserves cellular detail; excessive retrieval can damage morphology or increase nonspecific staining (standard IHC practice).
Could fixation explain weak or uneven RAB3IP staining?
Target-specific fixation sensitivity is unknown: the selected RAB3IP paraffin-section caption does not state a fixative (datasheet A09085-3). Record each specimen’s fixative and fixation interval, then compare similarly processed sections before attributing an intensity difference to RAB3IP abundance (standard IHC practice). Check whether staining varies with section thickness, tissue preservation or distance from the specimen edge, and repeat staining with the same EDTA pH 8.0 retrieval (datasheet A09085-3; standard IHC practice). If processing histories differ, report that limitation alongside the scores; neither the tissue staining profile nor RAB3IP’s lack of a transmembrane segment establishes a fixation effect (HPA tissue IHC; UniProt Q96QF0 topology).
Which cellular compartments should count as plausible RAB3IP staining?
For paraffin-section IHC, assess cytoplasmic staining first: the tissue profile describes cytoplasmic expression across several tissue types (HPA tissue IHC). Nuclear signal warrants separate recording because UniProt describes a small nuclear fraction, while cell imaging reports mainly nucleoplasmic signal with additional cytosol and centrosome localisation (UniProt Q96QF0 subcellular; HPA subcellular). RAB3IP also associates with the cytoskeleton, vesicles and lamellipodia, so a discrete peripheral pattern can be plausible in an appropriate cellular context (UniProt Q96QF0 subcellular). Do not score diffuse extracellular DAB deposit as intracellular RAB3IP; inspect cell boundaries and compare a negative control on the same run (standard IHC practice).
How should isoforms and epitope uncertainty affect an unexpected staining pattern?
RAB3IP has 8 listed isoforms, but the supplied antibody caption does not identify its epitope or establish which isoforms it detects (UniProt Q96QF0 isoforms; datasheet A09085-3). Do not assign a particular nuclear or cytoplasmic IHC pattern to one isoform on staining alone (UniProt Q96QF0 isoforms; standard IHC interpretation). If an independently mapped epitope becomes available, check whether its sequence occurs in the isoforms relevant to the specimen before comparing scores (standard antibody validation practice). The record also lists multiple phosphorylated residues, including positions 163 and 165; without epitope information, their effect on recognition remains unknown (UniProt Q96QF0 modified residues; datasheet A09085-3).
How can IF help resolve ambiguous chromogenic RAB3IP localisation?
Use IF as a separate localisation check rather than assuming the paraffin-section IHC conditions transfer directly (datasheet A09085-3; standard IF practice). Multiplex RAB3IP with a validated marker for the cell population being scored, selecting spectrally separated fluorophores and a channel with low tissue autofluorescence (standard IF practice). RAB3IP has no transmembrane segment and is reported in cytosolic, nuclear and centrosomal compartments, so permeabilise as needed to expose intracellular epitopes while preserving morphology (UniProt Q96QF0 topology and subcellular; HPA subcellular; standard IF practice). Include single-label and secondary-only controls, and interpret any IF pattern alongside the chromogenic section’s cell identity and compartment (standard IF practice).
How can I distinguish weak RAB3IP staining from DAB background?
The selected example used 10% goat serum blocking, 2 μg/ml primary antibody overnight at 4°C, a peroxidase-conjugated secondary and DAB detection (datasheet A09085-3). For diffuse staining, compare a primary-omission control, check endogenous peroxidase blocking, and shorten DAB development if the control also darkens (standard IHC practice). If background persists only with primary antibody, titrate its concentration while keeping retrieval and detection consistent; the caption’s concentration is a tested starting point, not a universal optimum (datasheet A09085-3; standard IHC practice). Review pigment, folds and section edges under brightfield before calling faint cytoplasmic deposit positive (standard IHC practice).
How should I score RAB3IP across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, then report the percentage of positive cells and an intensity-based H-score within the same annotated region (standard IHC scoring practice). An H-score can sum the percentages at intensity grades 0–3, weighted by grade, yielding 0–300 (standard IHC scoring practice). Normalise counts to eligible intact cells, or report positive-cell density per mm² of viable annotated tissue when cell counts are impractical (standard IHC scoring practice). Keep retrieval, imaging and scoring thresholds consistent, and report nuclear and cytoplasmic scores separately because both compartments have localisation support (datasheet A09085-3; UniProt Q96QF0 subcellular; HPA subcellular).
What makes an apparent RAB3IP-positive cell convincing?
A convincing call has intracellular staining in an identifiable, preserved cell and agrees with an appropriate control processed in the same run (standard IHC practice). Cytoplasmic staining is consistent with the tissue profile; a nuclear component is biologically plausible but should be scored separately and checked against neighbouring morphology (HPA tissue IHC; UniProt Q96QF0 subcellular). Inspect glandular cells or kidney tubule cells as potential comparison populations, while remembering that the tissue-IHC profile carries Uncertain reliability (HPA tissue IHC). Exclude edge enhancement, necrotic debris and endogenous enzyme signal using morphology and omission controls before interpreting differences as RAB3IP expression (standard IHC practice).
Boster reagents

Best RAB3IP / Rab-3A-interacting protein IHC Antibodies

A09085-3 has IHC data from paraffin sections of human ovarian serous adenocarcinoma and liver cancer, plus mouse and rat kidney (catalog image captions); no IF/ICC data are supplied (catalog applications and images).

Real IHC data IHC analysis of RAB3IP using anti-RAB3IP antibody (A09085-3). RAB3IP was detected in a paraffin-embedded section of human ovarian serous adenocarcinoma tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH 8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 μg/ml rabbit anti-RAB3IP Antibody (A09085-3) overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit (Catalog # SV0002) with DAB as the chromogen.
Anti-RAB3IP Antibody ®
Cat # A09085-3

A09085-3 is listed for IHC in human, mouse and rat samples (catalog applications and reactivity). Its IHC captions show paraffin sections of human ovarian serous adenocarcinoma, human liver cancer, mouse kidney and rat kidney (catalog image captions).

Which to pick: Choose A09085-3 for paraffin-section IHC; its ovarian tissue caption documents EDTA pH 8.0 retrieval and 2 μg/ml primary antibody (A09085-3 IHC image caption). The fixative is unreported (catalog image captions), and IF/ICC is not listed or illustrated, so there is no payload-supported IF/ICC pick (catalog applications and images). For human, mouse or rat IHC, A09085-3 is the listed option with species-specific tissue captions; its host is rabbit and clonality is unreported (catalog reactivity, image captions and host/clone fields).

Each figure is that product's own IHC / IF validation image from its datasheet.

References

  1. UniProt Consortium. UniProt entry Q96QF0 (RAB3I_HUMAN, Rab-3A-interacting protein).
  2. Human Protein Atlas. RAB3IP tissue IHC expression (reliability: Uncertain).
  3. Human Protein Atlas. RAB3IP subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol and centrosome..
  4. Human Protein Atlas. RAB3IP antibody validation summary (2 antibodies).
  5. Protocol for in vivo elimination of avian auditory hair cells, multiplexed mRNA detection, immunohistochemistry, and S-phase labeling. STAR protocols 2024 — PMC11217775.
  6. Promotion of Cell Proliferation through Inhibition of Cell Autophagy Signalling Pathway by Rab3IP is Restrained by MicroRNA-532-3p in Gastric Cancer. Journal of Cancer 2018 — PMC6277663.
  7. Hypomethylation of long interspersed nuclear element-1 (LINE-1) leads to activation of proto-oncogenes in human colorectal cancer metastasis. Gut 2014 — PMC3884067.
  8. PubMed PMID:12007189 — UniProt-cited evidence.
  9. PubMed PMID:12221131 — UniProt-cited evidence.
  10. PubMed PMID:14702039 — UniProt-cited evidence.