DNAJB1 / DnaJ homolog subfamily B member 1 · IHC design guide

Design Immunohistochemistry for DNAJB1

Plan DNAJB1 paraffin-section IHC around nuclear and cytoplasmic tissue staining (HPA tissue IHC). This guide covers high-staining tissue references (HPA tissue IHC), the catalog antibody’s 2–5 μg/mL IHC range (datasheet A03100-1), and heat-shock redistribution (UniProt).

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

Immunohistochemistry protocol sheet for DNAJB1 (IHC for DNAJB1): expected localisation Nucleus and cytoplasm (HPA tissue IHC), antibody A03100-1, validated IHC image, and IHC protocol steps — Printable **DNAJB1** IHC protocol sheet — expected localisation Nucleus and cytoplasm (HPA tissue IHC), antibody A03100-1, controls and protocol steps. Open the full DNAJB1 IHC guide →

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DNAJB1 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	Nucleus and cytoplasm (HPA tissue IHC)
	Staining pattern	General nuclear and cytoplasmic staining in tissue cells (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet A03100-1)
	Positive control ⓘ	Adrenal gland+4 more · see all
	Negative control ⓘ	None in HPA (detected in all 45 tissues); use no-primary + isotype controls

Important caveats

Reasons your staining may differ from the expected pattern.

	Fixation	Keep paraffin-section fixation consistent (standard IHC practice; not target-specific); Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A03100-1)
	Caveat	Heat shock can shift signal toward nucleoli (UniProt)
	Regulation	Abundance changes are unreported (UniProt)
	Isoform / epitope	2 isoforms; epitope coverage is unspecified (UniProt)

Section 1

Recommended DNAJB1 IHC & IF Protocols

The catalog antibody protocol uses EDTA pH 8.0 heat retrieval (datasheet A03100-1). The published protocols below describe fish ovary frozen sections (PMC6485077) and liver sections (PMC11278829).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human gastric signet ring cell carcinoma tissue; fixative not specified (datasheet A03100-1)
Fixation	Image fixative and duration unreported (datasheet A03100-1); 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 A03100-1); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet A03100-1)
Primary antibody	Rabbit anti-DNAJB1, 2-5 μg/ml (datasheet A03100-1)
Primary incubation	Overnight at 4 °C (datasheet A03100-1)
Detection	Streptavidin-biotin complex (SABC), DAB chromogen (datasheet A03100-1)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	DNAJB1-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: General nuclear and cytoplasmic expression. No signal in the no-primary control.

💡Decision noteFor paraffin sections, start with EDTA pH 8.0 heat retrieval (datasheet A03100-1); the published citric acid method was used on frozen sections (PMC6485077).

Section 2

What Is the Expected DNAJB1 Staining Pattern?

DNAJB1 staining is expected in nuclei and cytoplasm in paraffin sections (HPA: general nuclear and cytoplasmic expression). High staining is reported in skin basal cells, bronchial respiratory epithelium and adrenal glandular cells (HPA: High in each). The tissue pattern has Enhanced reliability, reflecting agreement between staining and RNA data (HPA: Enhanced). DNAJB1 has no transmembrane segment (UniProt P25685 topology).

What am I looking at on my slide?
Nuclear and cytoplasmic staining in the expected cells.	This fits the reported tissue pattern (HPA: general nuclear and cytoplasmic expression). Judge intensity within the identified cell population; adjacent cells need not stain equally.
Strong membrane, extracellular or lumenal staining dominates.	Treat this as suspect staining: DNAJB1 lacks a transmembrane segment and signal peptide (UniProt P25685 topology and processing). Review the no-primary control and tissue morphology before assigning a cellular signal (general IHC practice).
Strong staining occurs mainly in cells outside the reported population.	Check for cross-reactivity or endogenous detection activity (general IHC practice). For example, skin basal cells are reported High (HPA: High in skin basal layer); staining elsewhere alone needs corroboration.
Diffuse color covers tissue and empty spaces.	This is consistent with assay background rather than a resolved nuclear and cytoplasmic pattern (general IHC practice; HPA: tissue IHC profile). Compare with a no-primary control and inspect whether cell boundaries remain interpretable.
No staining appears in a known-positive tissue.	First investigate assay failure or specimen variation; a negative slide does not establish DNAJB1 absence (general IHC practice). Bronchial respiratory epithelial cells are a reported High reference (HPA: High in bronchus).

💡Expected DNAJB1 appearanceA positive IHC result shows discernible nuclear and cytoplasmic staining in reported High cell populations, such as skin basal cells (HPA: tissue IHC); dominant membrane or extracellular color is suspect (UniProt P25685 topology; general IHC practice).

How each factor affects the staining
Cell population and comparator tissue	HPA reports High staining in several epithelial and glandular populations, but Low in heart muscle cardiomyocytes and small-intestine glandular cells (HPA: tissue IHC). Low is a weaker comparator, not a verified negative control.
Compartment and stress state	Routine tissue IHC is described as nuclear and cytoplasmic (HPA: tissue IHC). DNAJB1 can move toward nuclei, especially nucleoli, after heat shock (UniProt P25685 subcellular annotation); nucleolar enrichment alone does not establish that stress occurred.
Antibody validation	HPA063247 has Enhanced IHC validation; CAB017450 has Supported IHC validation (HPA: antibody records). These statuses support interpretation of their observed patterns, but do not validate every specimen or detection setup (general IHC practice).
Isoforms and epitope coverage	DNAJB1 has 2 recorded isoforms (UniProt P25685 isoforms). The supplied evidence does not map the catalog antibody's epitope or isoform coverage, so a weak result cannot be assigned to an isoform without further validation.
Detection background	Endogenous enzyme activity or nonspecific secondary binding can add chromogenic color (general IHC practice). A no-primary control helps separate detection background from the reported cellular pattern (HPA: tissue IHC profile; general IHC practice).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
High-reference tissue has little or no signal.	A detection or staining step may have failed; tissue handling may also differ (general IHC practice).	Confirm the expected cell population, then review antibody dilution, retrieval and detection against the catalog antibody's IHC-P protocol; no target-specific fixation sensitivity is supplied (HPA: High tissue IHC; general IHC practice).
Membrane or extracellular color overwhelms cellular staining.	The dominant compartment conflicts with the reported pattern (HPA: tissue IHC; UniProt P25685 topology).	Inspect morphology and a no-primary control, then reassess blocking and detection conditions (general IHC practice). Avoid scoring this color as DNAJB1 without independent support.
Color appears broadly across cells reported Low.	Background or cross-reactivity is possible, although Low does not mean absent (HPA: Low tissue IHC; general IHC practice).	Compare the no-primary control and a reported High cell population in the same run; score the cell type and compartment separately from overall slide color (HPA: tissue IHC; general IHC practice).
Tissue edges and empty spaces show diffuse color.	Poorly resolved chromogen or nonspecific detection may obscure the cellular pattern (general IHC practice).	Check a no-primary control, blocking and wash steps; interpret only staining that can be assigned to intact nuclei or cytoplasm (general IHC practice; HPA: tissue IHC profile).
A second antibody gives a different pattern.	HPA reports different IHC validation levels for HPA063247 and CAB017450 (HPA: antibody records); discordance alone does not identify which result is correct.	Compare the same tissue and cell population, include detection controls, and favor a reproducible pattern matching HPA tissue IHC and UniProt localisation (general IHC practice; HPA: tissue IHC; UniProt P25685 subcellular annotation).
What should an IF/ICC image show?	HPA reports mainly nucleoplasmic DNAJB1 with additional cytosolic localisation in ICC-IF (HPA: supported subcellular locations).	Use those compartments as an image interpretation reference; consult the separate IF/ICC guide for its workflow (HPA: ICC-IF subcellular profile).

Source: UniProt P25685 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing DNAJB1 staining

IHC analysis of Hsp40/DNAJB1 using anti-Hsp40/DNAJB1 antibody (A03100-1). Hsp40/DNAJB1 was detected in a paraffin-embedded section of human gastric signet ring cell carcinoma 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-Hsp40/DNAJB1 Antibody (A03100-1) 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.

A03100-1 · IHCIHC analysis of Hsp40/DNAJB1 using anti-Hsp40/DNAJB1 antibody (A03100-1). Hsp40/DNAJB1 was detected in a paraffin-embedded section of human

IF analysis of Hsp40/DNAJB1 using anti-Hsp40/DNAJB1 antibody (A03100-1). Hsp40/DNAJB1 was detected in an immunocytochemical section of A549 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 μg/mL rabbit anti-Hsp40/DNAJB1 Antibody (A03100-1) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (BA1127) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

A03100-1 · IFIF analysis of Hsp40/DNAJB1 using anti-Hsp40/DNAJB1 antibody (A03100-1). Hsp40/DNAJB1 was detected in an immunocytochemical section of A549

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000132002-DNAJB1/tissue/cerebral+cortex#imid_17680383

HPA · EsophagusHuman Protein Atlas IHC, Esophagus. Source: https://www.proteinatlas.org/ENSG00000132002-DNAJB1/tissue/esophagus#imid_17680339

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000132002-DNAJB1/tissue/colon#imid_17680309

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000132002-DNAJB1/tissue/liver#imid_17680310

HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody CAB017450; green = DNAJB1, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for DNAJB1 IHC & IF

🧪Run esophagus first and score its squamous epithelial cells, which show high DNAJB1 staining (HPA: High in esophageal squamous epithelial cells). HPA detects DNAJB1 in all 45 scored tissues, so use no-primary and isotype controls for the negative comparison; no cell population on the positive slide is established as target-negative, and any unstained cells should be recorded as lacking visible signal rather than assumed negative (HPA: no negative tissue rows).

Positive control tissue: Adrenal gland (Glandular cells, HPA High)

Negative control tissue: None in HPA: DNAJB1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show DNAJB1 in A-431, U-251MG, U2OS, A-549, PC-3, with annotated localisation: Nucleoplasm (supported) (HPA subcellular).

Technical controls: Include a no-primary, secondary-only slide, a matched rabbit IgG isotype control, and, if available, a comparably processed DNAJB1 knockout specimen as a biological negative (selected-SKU caption: rabbit primary; standard IHC practice). Quench endogenous peroxidase and check for endogenous biotin before interpreting DAB staining, because the documented detection uses a biotinylated secondary and streptavidin–biotin complex (selected-SKU caption: biotinylated secondary, SABC and DAB).

⚠️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 unreported). That caption documents heat retrieval in EDTA at pH 8.0, but does not establish that retrieval is required under other conditions (selected-SKU caption: EDTA retrieval). Neither frozen sections nor IF is established as easier by the supplied evidence; for the esophagus slide, inspect squamous edges and keratin debris for nonspecific chromogen deposits before scoring cell-associated staining (HPA: esophageal squamous epithelial cells; standard IHC practice).

HPA tissue IHC evidence for DNAJB1

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — High 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
Adrenal gland	Glandular cells	High	Protein (IHC)	HPA →
Bronchus	Respiratory epithelial cells	High	Protein (IHC)	HPA →
Cervix	Squamous epithelial cells	High	Protein (IHC)	HPA →
Esophagus	Squamous epithelial cells	High	Protein (IHC)	HPA →
Gallbladder	Glandular cells	High	Protein (IHC)	HPA →

Undetected expression · recommended negative controls

Tissue	Cell type	Level	Evidence	Source
None in HPA: DNAJB1 is detected in all 45 scored tissues. Use a no-primary (secondary-only) and an isotype control instead.

Section 3

Advanced DNAJB1 IHC Tips

Troubleshoot DNAJB1 staining in paraffin sections by checking retrieval, compartment patterns, controls, and cell specific scoring before interpreting signal.

What retrieval should I try when DNAJB1 staining is weak?

Start with heat mediated retrieval in EDTA at pH 8.0 for the catalog antibody (datasheet A03100-1). The selected paraffin section image used that retrieval, followed by 2 μg/ml primary antibody overnight at 4°C (caption A03100-1). If staining remains weak, compare retrieval duration or a different buffer on adjacent sections while keeping primary concentration and detection conditions constant (standard IHC practice). Check that stronger signal retains interpretable nuclear and cytoplasmic staining, since both compartments are reported for DNAJB1 (HPA tissue IHC). Record retrieval conditions alongside morphology and background; an increase in diffuse staining alone does not establish successful antigen recovery (standard IHC practice).

Could fixation explain absent or uneven DNAJB1 staining?

The selected image documents a paraffin section but does not state its fixative, so target specific fixation sensitivity is unknown (caption A03100-1). Record the fixative and fixation history for each specimen, and compare sections with matched processing before attributing a difference to DNAJB1 abundance (standard IHC practice). Inspect preserved morphology and staining across the section; weak signal limited to poorly preserved areas warrants a processing review (standard IHC practice). Use the documented EDTA pH 8.0 retrieval and 2 μg/ml primary antibody as reference conditions when troubleshooting the catalog antibody (caption A03100-1). Do not infer fixation tolerance from DNAJB1 localisation or phosphorylation (UniProt P25685).

How should I judge nuclear versus cytoplasmic DNAJB1 staining?

Evaluate nuclear and cytoplasmic signal separately: tissue IHC reports general expression in both compartments (HPA tissue IHC). Nucleoplasmic localisation is supported by cell imaging, with additional cytosolic localisation; UniProt also lists nucleoli, especially after heat shock (HPA subcellular; UniProt P25685). Compare the same cell type across matched sections, because a change in compartment balance can be obscured by a single overall intensity score (standard IHC practice). Check the counterstain and cell boundaries before calling a nuclear pattern, and assess nearby negative reagent controls for nonspecific deposition (standard IHC practice). A nucleolar pattern should be interpreted with the specimen’s stress context rather than assumed from chromogenic intensity alone (UniProt P25685; standard IHC practice).

Can this stain distinguish DNAJB1 isoforms or phosphorylation states?

DNAJB1 has 2 annotated isoforms, a J domain at residues 2–70, and a reported phosphothreonine at residue 307 (UniProt P25685). The supplied catalog image does not identify the antibody epitope, so its coverage of each isoform or phosphorylation state cannot be established from this IHC result (caption A03100-1). Check the antibody’s documented immunogen and epitope information before designing an isoform specific interpretation (standard IHC practice). Keep retrieval and detection matched when comparing sections (standard IHC practice).

How can IF help resolve ambiguous DNAJB1 localisation?

Use IF/ICC as a separate localisation check: cell imaging supports mainly nucleoplasmic DNAJB1 with additional cytosolic signal (HPA subcellular). Multiplex with a marker for the cell type being evaluated, and choose fluorophores in channels with low measured tissue autofluorescence (standard IF practice). DNAJB1 has no annotated transmembrane segment, so optimise permeabilisation for intracellular nuclear and cytosolic access while preserving cell boundaries (UniProt P25685; standard IF practice). Include single label and secondary only controls to assess channel bleed through and background before comparing compartments (standard IF practice). The paraffin section image documents a chromogenic IHC method; it supplies no IF fixation or antibody validation conditions (caption A03100-1).

What should I check when DAB obscures specific staining?

Compare a section without primary antibody with the stained section to identify detection related background (standard IHC practice). The selected workflow used 10% goat serum blocking, a biotinylated secondary, a streptavidin biotin complex, and DAB (caption A03100-1). Check endogenous peroxidase blocking and, with this detection system, consider whether endogenous biotin contributes to signal in the specimen (standard IHC practice). Compare background after adjusting primary antibody exposure around the documented 2 μg/ml overnight at 4°C condition, without changing retrieval simultaneously (caption A03100-1; standard IHC practice). Score DNAJB1 only where nuclear or cytoplasmic signal is distinguishable from diffuse precipitate and tissue pigment (HPA tissue IHC; standard IHC practice).

How should I score DNAJB1 across heterogeneous sections? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and region before scoring, then record nuclear and cytoplasmic staining separately because both are reported for DNAJB1 (HPA tissue IHC). An H score can combine the percentage of cells at each intensity grade; alternatively, report percentage positive cells or positive cell density per mm² (standard IHC practice). Normalise density to the analysed tissue area and compare matched cell populations, since cellular composition can shift an overall section score (standard IHC practice). Set positivity thresholds against appropriate negative controls and apply the same threshold, counterstain assessment, and exclusion rules across sections (standard IHC practice). Report retrieval and detection conditions with the scores (standard IHC practice).

How can I distinguish DNAJB1 signal from artefact?

A credible pattern may include nuclear and cytoplasmic staining, consistent with reported tissue IHC; nucleoplasmic and cytosolic localisation also have cell imaging support (HPA tissue IHC; HPA subcellular). Compare the stained cell type with an expected positive population, such as respiratory epithelial cells in bronchus, while accounting for variation within the specimen (HPA tissue IHC). Treat staining confined to section edges or necrotic areas as suspect, and inspect a primary omitted control for detection artefacts (standard IHC practice). Residual endogenous peroxidase can mimic DAB signal, so review the peroxidase block and controls before calling a weak positive (standard IHC practice). Interpret prominent nucleolar staining in light of possible heat shock related redistribution (UniProt P25685).

Boster reagents

Best DNAJB1 / DnaJ homolog subfamily B member 1 IHC Antibodies

A03100-1 has IHC data from human paraffin sections (IHC caption) and IF/ICC data from A549 cells (IF caption); catalog reactivity lists human, mouse and rat (catalog: reactivity).

Real IHC data

Anti-Hsp40/DNAJB1 Antibody ®

Cat # A03100-1

View product →

A03100-1 will render with IHC data from a paraffin section of human gastric signet ring cell carcinoma tissue (IHC caption). A03100-1 is also listed for IF/ICC, with an image of A549 cells (catalog: applications; IF caption).

Which to pick: Choose A03100-1 for paraffin-section IHC: its image caption documents EDTA retrieval at pH 8.0 and 2 μg/ml primary antibody; the fixative is unreported (IHC caption). For IF/ICC, the same SKU has an A549 cell image using 5 μg/ml primary antibody (IF caption). For mouse or rat work, A03100-1 lists those species as reactive, but the supplied IHC and IF images show human samples; no clone is specified (catalog: reactivity and clone; IHC/IF captions).

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

References

UniProt Consortium. UniProt entry P25685 (DNJB1_HUMAN, DnaJ homolog subfamily B member 1).
Human Protein Atlas. DNAJB1 tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. DNAJB1 subcellular location (ICC-IF): Mainly localized to the nucleoplasm. In addition localized to the cytosol..
Human Protein Atlas. DNAJB1 antibody validation summary (2 antibodies).
Molecular cloning and characteristics of DnaJa1and DnaJb1 in Coilia nasus: possible function involved in oogenesis during spawning migration. BMC developmental biology 2019 — PMC6485077.
A Prognostic Prediction Model Developed Based on Four CpG Sites and Weighted Correlation Network Analysis Identified DNAJB1 as a Novel Biomarker for Pancreatic Cancer. Frontiers in oncology 2020 — PMC7477361.
CRISPR/Cas9 Engineering of Adult Mouse Liver Demonstrates That the Dnajb1-Prkaca Gene Fusion Is Sufficient to Induce Tumors Resembling Fibrolamellar Hepatocellular Carcinoma. Gastroenterology 2017 — PMC5801691.
Diagnosis and Molecular Characterization of Potential RNA Binding Protein Involved in the Pathogenesis of Liver Ischemia Reperfusion Injury. Journal of inflammation research 2024 — PMC11278829.
PubMed PMID:1754405 — UniProt-cited evidence.
PubMed PMID:8250930 — UniProt-cited evidence.
PubMed PMID:8975727 — UniProt-cited evidence.