BIK / Bcl-2-interacting killer · IHC design guide

Design Immunohistochemistry for BIK

Plan BIK paraffin IHC with the catalog antibody at 0.5–1 μg/ml (datasheet PB9755). Use gallbladder glandular cells as a high-staining reference and interpret intensity in light of the low consistency between antibody staining and RNA expression (HPA tissue IHC).

Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use

Immunohistochemistry protocol sheet for BIK (IHC for BIK): expected localisation Cytoplasmic staining in most tissues (HPA tissue IHC), antibody PB9755, validated IHC image, and IHC protocol steps — Printable **BIK** IHC protocol sheet — expected localisation Cytoplasmic staining in most tissues (HPA tissue IHC), antibody PB9755, controls and protocol steps. Open the full BIK IHC guide →

Navigate to: Protocols Staining pattern Controls Antibodies

BIK 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 staining in most tissues (HPA tissue IHC)
	Staining pattern	Cytoplasmic staining; high in gallbladder glandular cells (HPA tissue IHC)
	Antigen retrieval	Citrate pH 6 HIER, heat-mediated (datasheet PB9755)
	Positive control ⓘ	Gallbladder+4 more · see all
	Negative control ⓘ	Breast+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 PB9755)
	Caveat	Staining has low consistency with RNA expression (HPA tissue IHC)
	Regulation	Expression regulation is not annotated (UniProt)
	Isoform / epitope	One 1–160 chain; no isoforms or processing annotated (UniProt)

Section 1

Recommended BIK IHC & IF Protocols

The catalog antibody’s IHC-P protocol is paired with a published chromogenic BIK IHC protocol from breast tumor research (PMC1198222).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded Human Thyroid Cancer tissue; fixative not specified (datasheet PB9755)
Fixation	Image fixative and duration unreported (datasheet PB9755); verify before use.
Sectioning	4–5 µm sections on charged slides (standard)
Deparaffinisation	Xylene, graded ethanol series to water (standard)
Antigen retrieval	Heat retrieval: Citrate pH 6, 20 min (datasheet PB9755)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet PB9755)
Primary antibody	Rabbit anti-BIK, 0.5-1μg/ml (datasheet PB9755)
Primary incubation	Overnight at 4 °C (datasheet PB9755)
Detection	Streptavidin-biotin complex (SABC), DAB chromogen (datasheet PB9755)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	BIK-positive staining in glandular cells of gallbladder (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control.

💡Decision noteStart with heat-mediated citrate pH 6 retrieval (datasheet: PB9755); the published protocol used autoclaving in citrate pH 6.1 (PMC1198222).

Section 2

What Is the Expected BIK Staining Pattern?

BIK is a membrane-associated protein reported around the nuclear envelope and in cytoplasmic membranes; UniProt also places it in the endomembrane system and mitochondrial membrane (UniProt Q13323 topology, residues 136–156; subcellular location). In tissue IHC, expect mainly cytoplasmic staining, including high signal in gallbladder glandular cells and spleen red-pulp cells (HPA: tissue IHC). HPA rates its IHC evidence Approved, while reporting low consistency between staining and RNA expression (HPA: reliability).

What am I looking at on my slide?
Cytoplasmic staining in gallbladder glandular cells or spleen red-pulp cells.	This matches the reported high-staining cells (HPA: High in gallbladder glandular cells; High in spleen red-pulp cells). A rim near the nucleus can fit the reported membrane location (UniProt Q13323: around the nuclear envelope), but chromogenic IHC alone cannot identify a specific organelle.
A predominantly nuclear or sharply extracellular signal replaces the expected cytoplasmic pattern.	Treat this as a possible staining artefact and review morphology and controls. The reported locations are cytoplasmic or membrane associated (HPA: cytoplasmic expression in most tissues; UniProt Q13323: subcellular location); the supplied evidence does not support nuclear or extracellular BIK staining.
Strong staining appears in a cell population reported as undetected, such as breast adipocytes.	Consider antibody cross-reactivity or endogenous chromogen-generating activity before assigning the signal to BIK (HPA: Not detected in breast adipocytes). The HPA observation applies to that cell population; it does not establish that every cell in the tissue is negative.
Colour spreads across tissue structures and the slide background, obscuring cell boundaries.	Interpret localisation only after assessing this nonspecific background. Uneven blocking, incomplete washing, or detection background are general IHC possibilities; a diffuse field cannot be scored reliably against HPA's predominantly cytoplasmic tissue pattern (HPA: tissue IHC profile).
No staining is visible in gallbladder glandular cells or spleen red-pulp cells.	Check the assay and tissue morphology before calling the sample BIK negative: both populations are reported as high (HPA: tissue IHC). HPA's Approved rating carries a low staining-versus-RNA consistency caveat, so one absent result does not settle biological expression (HPA: reliability).

💡Expected BIK appearanceCall a positive result when discrete cytoplasmic staining is visible in the expected cells, preferably strong in gallbladder glandular cells or spleen red-pulp cells (HPA: High); dominant nuclear, extracellular, or diffuse slide-wide colour warrants control review (HPA: cytoplasmic profile; UniProt Q13323: subcellular location).

How each factor affects the staining
Membrane topology and apparent compartment	BIK has one transmembrane segment at residues 136–156 and is reported around the nuclear envelope and in cytoplasmic membranes (UniProt Q13323: topology; subcellular location). These annotations support membrane-associated cytoplasmic interpretation, but do not prove that a chromogenic deposit marks a particular organelle.
Choice of positive and comparison cells	Gallbladder glandular cells and spleen red-pulp cells are reported High; appendix and colon glandular cells are Medium (HPA: tissue IHC). Use the named cell population when comparing intensity. Cervix and rectum glandular cells are reported Not detected (HPA: tissue IHC), which does not make the entire tissue a negative control.
Strength of validation evidence	The listed antibody HPA051360 has Approved IHC status, and HPA reports low consistency between tissue staining and RNA expression (HPA: antibody validation; reliability). Interpret intensity as an observed staining pattern, not a precise measure of BIK abundance; the supplied status is not an Enhanced validation claim.
Processing and target variants	The record lists a single 1–160 chain, no signal peptide or propeptide, no annotated glycosylation sites, and no listed isoforms (UniProt Q13323: processing; glycosylation; isoforms). These annotations offer no supported basis for predicting a distinct shed, processed, or isoform-specific IHC pattern.
Can IF/ICC establish a finer location?	HPA summarises the ICC-IF location as Membrane, but gives no main location or cell-line images in the supplied record (HPA: subcellular). That entry cannot independently verify a precise organelle pattern; IF/ICC assay design belongs in its own guide.

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
The positive-control cells show no chromogen.	The assay may have failed, or the selected section may lack intact target cells; absence alone cannot resolve expression (HPA: High in gallbladder glandular cells and spleen red-pulp cells).	Confirm cell identity and section quality, then review primary-antibody, detection, and counterstain steps. If the run still fails, optimise antigen retrieval as a general IHC workflow step; BIK-specific retrieval and fixation effects are unreported here.
The strongest signal is nuclear.	This conflicts with the reported cytoplasmic and membrane-associated locations (HPA: tissue IHC profile; UniProt Q13323: subcellular location). Nonspecific primary or detection signal is possible.	Compare with a primary-omission control and examine whether the signal follows nuclei across unrelated cells. Review staining conditions before scoring it as BIK.
An HPA-undetected cell population stains strongly.	Cross-reactivity or endogenous detection activity may mimic target staining; the expected observation for that named population is absent (HPA: Not detected in breast adipocytes, for example).	Verify the cell type, inspect a primary-omission control, and assess endogenous detection background using standard chromogenic IHC controls. Avoid extending that cell-level HPA result to the whole tissue.
Diffuse colour makes cytoplasmic boundaries hard to judge.	General IHC background from blocking, washing, or detection can obscure the reported cytoplasmic pattern (HPA: tissue IHC profile). The supplied sources do not identify a BIK-specific cause.	Check background on a primary-omission section and review blocking, washes, and detection exposure. Score localisation only where cells remain distinguishable.
A medium-staining tissue appears weaker than a high-staining control.	This can agree with the reported levels: appendix or colon glandular cells are Medium, while gallbladder glandular cells are High (HPA: tissue IHC).	Compare the same named cell types across well-preserved sections and score intensity within each population. Investigate a discrepancy only after confirming morphology and run controls.
IHC intensity disagrees with an RNA-based expectation.	HPA explicitly reports low consistency between antibody staining and RNA expression despite its Approved IHC rating (HPA: reliability).	Report the observed cellular staining and the validation caveat together. Use the IHC-validated antibody's controls and an independent readout if a stronger expression claim is needed; do not infer a BIK-specific fixation effect.

Source: UniProt Q13323 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing BIK staining

IHC analysis of Bik using anti-Bik antibody (PB9755). Bik was detected in paraffin-embedded section of Human Thyroid Cancer Tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-Bik Antibody (PB9755) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

PB9755 · IHCIHC analysis of Bik using anti-Bik antibody (PB9755). Bik was detected in paraffin-embedded section of Human Thyroid Cancer Tissue. Heat med

IHC analysis of Bik using anti-Bik antibody (PB9755). Bik was detected in paraffin-embedded section of Mouse Spleen Tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-Bik Antibody (PB9755) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.

PB9755 · IHCIHC analysis of Bik using anti-Bik antibody (PB9755). Bik was detected in paraffin-embedded section of Mouse Spleen Tissue. Heat mediated an

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000100290-BIK/tissue/cerebral+cortex#imid_16326914

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000100290-BIK/tissue/colon#imid_16314049

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000100290-BIK/tissue/liver#imid_16314058

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000100290-BIK/tissue/kidney#imid_16314059

← swipe / scroll for more images →

Real IHC / IF data behind the staining-pattern expectations in this section.

Sample controls for BIK IHC & IF

🧪Run spleen first: cells in the red pulp should stain (HPA: High in spleen red-pulp cells). Use liver cholangiocytes as the negative tissue (HPA: Not detected in liver cholangiocytes); white-pulp cells on the spleen slide are candidate internal negatives and should show background-level staining if independently confirmed BIK-negative (HPA: positive signal assigned to red-pulp cells).

Positive control tissue: Gallbladder (Glandular cells, HPA High)

Negative control tissue: Breast (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA carries no ICC-IF cell line for BIK; derive a cell-line control from the positive tissue's cell type (Glandular cells) and confirm it by RNA or western blot first.

Technical controls: Include a no-primary, secondary-only control, a rabbit IgG isotype control matched to the primary antibody, and BIK-knockout tissue or a validated peptide-block control (selected-SKU caption: rabbit primary antibody; standard IHC practice). Quench endogenous peroxidase in spleen red pulp and check for endogenous biotin when using the caption’s streptavidin–biotin detection system (HPA: spleen red-pulp cells; selected-SKU caption: SABC with DAB; standard IHC practice).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected-SKU paraffin-section caption does not state the fixative (selected-SKU caption: fixative not stated). Citrate retrieval at pH 6 for 20 minutes is a starting point from the paraffin thyroid-cancer caption; its performance in spleen is unreported (selected-SKU caption: citrate retrieval). No supplied evidence establishes that frozen sections or IF are easier for BIK; in chromogenic spleen IHC, red-pulp endogenous peroxidase can mimic specific signal unless quenched (HPA: spleen red-pulp cells; standard IHC practice).

HPA tissue IHC evidence for BIK

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

Positive expression · recommended positive controls

Tissue	Cell type	Level	Evidence	Source
Gallbladder	Glandular cells	High	Protein (IHC)	HPA →
Spleen	Cells in red pulp	High	Protein (IHC)	HPA →
Appendix	Glandular cells	Medium	Protein (IHC)	HPA →
Bone marrow	Hematopoietic cells	Medium	Protein (IHC)	HPA →
Bronchus	Ciliated cells (tip of cilia)	Medium	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
Breast	Adipocytes	Not detected	Protein (IHC)	HPA →
Cerebral cortex	Endothelial cells	Not detected	Protein (IHC)	HPA →
Cervix	Glandular cells	Not detected	Protein (IHC)	HPA →
Hippocampus	Glial cells	Not detected	Protein (IHC)	HPA →
Liver	Cholangiocytes	Not detected	Protein (IHC)	HPA →

Section 3

Advanced BIK IHC Tips

Troubleshoot BIK staining in paraffin sections by checking retrieval, cellular pattern, controls and scoring before interpreting chromogenic signal.

What retrieval should I start with for BIK IHC in paraffin sections?

Start with heat-mediated citrate retrieval at pH 6 for 20 minutes (datasheet PB9755). The selected paraffin-section image used that retrieval, followed by 1 µg/mL catalog antibody overnight at 4°C and chromogenic detection (datasheet PB9755). If signal is weak, check that sections reached a consistent retrieval temperature, then vary heating time cautiously while keeping the citrate buffer at pH 6 (standard IHC practice). Run each adjustment alongside the original conditions and a control section, because changes in staining intensity alone cannot establish BIK specificity (standard IHC practice).

How should I troubleshoot fixation-related loss of BIK staining?

Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative or fixation duration (datasheet PB9755). Record the fixative, time before fixation and fixation duration for each specimen, then compare sections processed under matched conditions (standard IHC practice). If a batch stains weakly, repeat the established pH 6 citrate retrieval for 20 minutes before testing a limited retrieval-time adjustment (datasheet PB9755; standard IHC practice). Include a concurrently processed control tissue and inspect morphology, since variable tissue preservation can confound an apparent change in chromogenic signal (standard IHC practice).

Where should convincing BIK staining appear in tissue sections?

Evaluate cytoplasmic and membrane-associated staining: BIK is reported around the nuclear envelope and in cytoplasmic membranes, including the mitochondrion membrane (UniProt Q13323 localisation). Its 136–156 transmembrane segment supports membrane association, but routine chromogenic IHC cannot identify a particular organelle from a diffuse cytoplasmic signal (UniProt Q13323 topology; standard IHC practice). HPA reports cytoplasmic expression in most tissues and high staining in gallbladder glandular cells and spleen red-pulp cells (HPA tissue IHC). If staining is exclusively nuclear or confined to tissue edges, check morphology and controls before assigning it to BIK (standard IHC practice).

Could epitope placement explain inconsistent BIK IHC staining?

Check the catalog antibody's immunogen or epitope information before changing retrieval, because the supplied evidence does not locate its binding site (datasheet PB9755). The annotated BIK chain spans residues 1–160 and has one transmembrane segment at 136–156; no isoforms are annotated in the supplied record (UniProt Q13323). An epitope within or near the membrane segment may be less accessible after tissue processing, but that remains a hypothesis until the epitope is known (UniProt Q13323 topology; standard IHC practice). Compare matched sections across a small retrieval series and judge gains against background and preserved morphology (standard IHC practice).

How can IF help assess a questionable BIK IHC pattern?

On the separate IF/ICC guide page, consider multiplexing BIK with a marker identifying the expected cell population, such as glandular cells in a gallbladder sample (HPA tissue IHC; standard IF practice). Choose spectrally separated fluorophores and assess an unstained section first; a far-red channel may help when tissue autofluorescence obscures shorter wavelengths (standard IF practice). BIK is membrane-associated, but the supplied topology does not establish which side carries the catalog antibody's epitope (UniProt Q13323 topology; datasheet PB9755). Compare controlled permeabilisation conditions if a cytoplasm-facing epitope is suspected, and verify the pattern against single-label and no-primary controls (standard IF practice).

What should I check when DAB background obscures BIK staining?

The selected paraffin-section image used 10% goat serum blocking, a biotinylated secondary antibody, an avidin–biotin complex and DAB (datasheet PB9755). For excessive brown signal, inspect no-primary and secondary-only controls, confirm adequate washes, and check whether the antibody concentration exceeds the illustrated 1 µg/mL condition (datasheet PB9755; standard IHC practice). Block endogenous peroxidase before DAB development and assess whether the avidin–biotin system contributes background in the specimen (standard IHC practice). Score only cellular staining with interpretable morphology, excluding precipitate, folds and damaged edges from the assessment (standard IHC practice).

How should I score BIK IHC across specimens? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell compartment and eligible cell population before scoring, since HPA reports cytoplasmic BIK staining and differing levels among tissue cell types (HPA tissue IHC). For a consistent population, record the percentage of positive cells and intensity categories, then calculate an H-score from intensity-weighted percentages on a 0–300 scale (standard IHC practice). If cell density varies substantially, report positive cells per mm² of viable tissue and retain the total eligible-cell count as the denominator (standard IHC practice). Use matched processing, retrieval and exposure to DAB across specimens, and exclude necrotic or folded regions by a prespecified rule (standard IHC practice).

How can I distinguish a true BIK signal from an IHC artefact?

A plausible signal follows cytoplasmic or membrane-associated cellular staining rather than an exclusively nuclear pattern (UniProt Q13323 localisation; HPA tissue IHC). Compare the same cell type across controls: HPA reports high staining in gallbladder glandular cells but no detected staining in cervix glandular cells, while warning of low consistency between antibody staining and RNA data (HPA tissue IHC). Edge-restricted colour, necrotic areas and signal persisting without primary antibody warrant investigation as staining artefacts (standard IHC practice). Check endogenous peroxidase blocking and chromogen development, then seek concordance across controls before interpreting a biological difference in BIK expression (standard IHC practice).

Boster reagents

Best BIK / Bcl-2-interacting killer IHC Antibodies

PB9755 has real IHC images from paraffin sections of human thyroid cancer and mouse and rat spleen (catalog IHC captions). ICC is listed, but no IF image is supplied (catalog applications; IF image list).

Real IHC data

Anti-Bik Antibody ®

Cat # PB9755

View product →

PB9755 will render with its own IHC image from a paraffin section of human thyroid cancer (catalog IHC caption). Additional captions show mouse and rat spleen paraffin sections; the catalog lists ICC but supplies no IF image (catalog IHC captions; catalog applications; IF image list).

Which to pick: Choose PB9755 for tissue IHC: its paraffin-section caption documents citrate retrieval at pH 6 for 20 minutes and primary antibody at 1 μg/ml (catalog IHC caption). For IF/ICC, PB9755 lists ICC, but has no IF image; clonality is unreported (catalog applications; IF image list; catalog clone field). For cross-species IHC, PB9755 lists human, mouse, and rat reactivity with matching paraffin-section captions; the fixative is unreported (catalog reactivity; catalog IHC captions).

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

References

UniProt Consortium. UniProt entry Q13323 (BIK_HUMAN, Bcl-2-interacting killer).
Human Protein Atlas. BIK tissue IHC expression (reliability: Approved).
Human Protein Atlas. BIK subcellular location (ICC-IF): Membrane.
Human Protein Atlas. BIK antibody validation summary (1 antibodies).
A molecular analysis by gene expression profiling reveals Bik/NBK overexpression in sporadic breast tumor samples of Mexican females. BMC cancer 2005 — PMC1198222.
Induced autoimmunity against gonadal proteins affects gonadal development in juvenile zebrafish. PloS one 2014 — PMC4250200.
BIK drives an aggressive breast cancer phenotype through sublethal apoptosis and predicts poor prognosis of ER-positive breast cancer. Cell death & disease 2020 — PMC7289861.
Interrogation of gender disparity uncovers androgen receptor as the transcriptional activator for oncogenic miR-125b in gastric cancer. Cell death & disease 2021 — PMC8096848.
PubMed PMID:7478623 — UniProt-cited evidence.
PubMed PMID:8816500 — UniProt-cited evidence.
PubMed PMID:10500065 — UniProt-cited evidence.