CTSF / Cathepsin F · IHC design guide

Design Immunohistochemistry for CTSF

Plan CTSF chromogenic IHC in paraffin sections using hepatocytes or cortical neurons as positive controls (HPA tissue IHC). This guide covers granular cytoplasmic staining (HPA tissue IHC), lysosomal context (UniProt), and an antibody starting range of 2–5 μg/ml (datasheet A06600-1).

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

Immunohistochemistry protocol sheet for CTSF (IHC for CTSF): expected localisation Granular cytoplasm in tissue; lysosomal by annotation (HPA tissue IHC; UniProt), antibody A06600-1, validated IHC image, and IHC protocol steps — Printable **CTSF** IHC protocol sheet — expected localisation Granular cytoplasm in tissue; lysosomal by annotation (HPA tissue IHC; UniProt), antibody A06600-1, controls and protocol steps. Open the full CTSF IHC guide →

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CTSF 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	Granular cytoplasm in tissue; lysosomal by annotation (HPA tissue IHC; UniProt)
	Staining pattern	Granular cytoplasm in most tissues; neurons and hepatocytes high (HPA tissue IHC)
	Antigen retrieval	EDTA pH 8.0 HIER, heat-mediated (datasheet A06600-1)
	Positive control ⓘ	Cerebellum+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 paraffin-section fixation consistent. (standard IHC practice; not target-specific)
	Caveat	Colon glands lack staining despite moderate colon expression (HPA tissue IHC; UniProt)
	Regulation	Tissue-dependent; high in brain and testis (UniProt)
	Isoform / epitope	No annotated isoforms; processing requires epitope mapping (UniProt)

Section 1

Recommended CTSF IHC & IF Protocols

The catalog antibody protocol is accompanied by published CTSF IHC methods for lung tumour and brain metastasis tissue and human skin (PMC9174239; PMC9886782).

Recommended immunohistochemistry (IHC-P) protocol parameters
Sample	Paraffin-embedded human stomach cancer tissue; fixative not specified (datasheet A06600-1)
Fixation	Image fixative and duration unreported (datasheet A06600-1); 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 A06600-1); 20 min, 95–100 °C (standard)
Peroxidase block	3% H2O2, 10 min, room temperature (standard)
Blocking	10% goat serum (datasheet A06600-1)
Primary antibody	Rabbit anti-CTSF, 2-5 μg/ml (datasheet A06600-1)
Primary incubation	Overnight at 4 °C (datasheet A06600-1)
Detection	HRP-conjugated secondary, DAB chromogen (datasheet A06600-1)
Counterstain	Hematoxylin, blue, dehydrate and mount (standard)
Expected result	CTSF-positive staining in cells in molecular layer of cerebellum (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression with a granular pattern in most tissues. No signal in the no-primary control.

💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A06600-1); the lung tumour and brain metastasis study also reports EDTA retrieval (PMC9174239).

Section 2

What Is the Expected CTSF Staining Pattern?

CTSF localizes to lysosomes and has no transmembrane segment (UniProt Q9UBX1 localization/topology). In paraffin IHC, expect granular cytoplasmic staining, including strong signal in hepatocytes, cortical and hippocampal neurons, and testicular Leydig cells (HPA: tissue IHC). HPA rates the tissue IHC profile Enhanced while describing only medium consistency between staining and RNA data (HPA: reliability).

What am I looking at on my slide?
Discrete cytoplasmic granules in hepatocytes or cortical neurons, with stronger staining than adjacent negative cells (HPA: High in hepatocytes and cortical neurons).	This matches the reported tissue pattern and lysosomal location (HPA: granular cytoplasm; UniProt Q9UBX1: lysosome). Granularity alone cannot prove lysosomal identity or antibody specificity; assess the cell type and controls together (general IHC interpretation).
Predominantly nuclear staining or a continuous cell-edge outline replaces cytoplasmic granules (HPA: granular cytoplasmic tissue pattern).	Treat this as discordant with the expected IHC pattern and check controls and staining conditions (HPA: tissue IHC; general IHC practice). HPA also reports plasma-membrane IF signal with uncertain support, so an isolated edge signal is not decisive evidence of CTSF localization (HPA: subcellular IF).
Strong signal appears in adipocytes or bone-marrow hematopoietic cells (HPA: Not detected in these cell types).	Consider nonspecific antibody binding or detection-system activity, especially when the expected positive cells also stain poorly (general IHC interpretation). HPA's cell-specific observations are comparison points, not proof that every cell in those tissues lacks CTSF (HPA: tissue IHC).
Uniform haze covers cells and extracellular spaces, obscuring granules (HPA: expected granular cytoplasmic pattern).	This prevents a confident positive call (general IHC interpretation). Review primary-antibody concentration, blocking, washing, and chromogen development with a no-primary control; these are general IHC checks, not documented CTSF-specific sensitivities (general IHC practice).
No granular signal is visible in adequately sampled hepatocytes or cortical neurons (HPA: High in both cell types).	An absent expected positive weakens the run's interpretability (general IHC interpretation). Check tissue preservation, retrieval, primary-antibody conditions, and detection with controls; the supplied sources do not identify a CTSF-specific fixation or retrieval failure mode (general IHC practice).

💡Expected CTSF appearanceCall a section positive when appropriate cells show distinct, preferably strong cytoplasmic granules—such as hepatocytes or cortical neurons—while diffuse haze, dominant nuclear staining, or strong adipocyte staining raises concern for an artefact (HPA: tissue IHC; UniProt Q9UBX1: lysosome; general IHC interpretation).

How each factor affects the staining
Cellular location and topology (UniProt Q9UBX1: lysosome; no transmembrane segment).	The expected tissue readout is intracellular granularity, consistent with HPA's cytoplasmic pattern (HPA: tissue IHC). A chromogenic granule cannot identify its organelle by itself; assess morphology and controls (general IHC interpretation).
Proteolytic processing and antibody epitope (UniProt Q9UBX1: signal peptide 1–19, propeptide 20–270, mature chain 271–484).	Different epitopes could recognize different processing states, but the supplied record gives no epitope for the catalog antibody (UniProt Q9UBX1: processing; supplied antibody data). Do not predict which form produces tissue staining from these boundaries alone.
Tissue and cell-type differences (HPA: tissue IHC; UniProt Q9UBX1: tissue specificity).	HPA reports High hepatocyte staining but Not detected in colon glandular cells and ovarian stroma (HPA: tissue IHC). UniProt reports moderate colon and high ovary expression (UniProt Q9UBX1: tissue specificity); tissue-level expression and selected-cell IHC observations should be interpreted at their respective scales.
Antibody-specific validation (HPA: antibody records and tissue IHC reliability).	CAB002141 has Enhanced IHC status, whereas HPA031431 has Supported ICC status without an IHC rating in the supplied list (HPA: antibodies). HPA's Enhanced tissue profile includes medium staining–RNA consistency; neither label transfers automatically to a different catalog antibody (HPA: reliability; general assay interpretation).
IF/ICC Q: Should vesicles appear? (HPA: subcellular IF).	A: Vesicles are the supported main IF location; endoplasmic-reticulum and plasma-membrane locations are uncertain (HPA: subcellular IF). This helps interpret images from an independently validated IF assay but does not establish an IHC compartment beyond the reported granular cytoplasm (HPA: tissue IHC).

Why is my staining missing, weak or wrong?
Situation	Likely cause	Next action
Expected positive tissue has no staining (HPA: High in hepatocytes or cortical neurons).	The run may have failed at retrieval, primary incubation, or detection, but no CTSF-specific failure step is identified (general IHC practice; supplied sources).	Check a known-positive section in the same run, then review retrieval and reagent performance; avoid declaring the sample negative until its positive control works (general IHC practice).
Granules are weak in a reported High cell type (HPA: tissue IHC).	Weak detection or tissue condition is possible; HPA's High category does not set a required intensity for every section (general IHC interpretation; HPA: tissue IHC).	Compare matched positive and no-primary controls, and optimize antibody dilution or detection within the validated IHC workflow (general IHC practice).
Signal fills nuclei or traces cell borders more than cytoplasmic granules (HPA: tissue IHC).	The pattern conflicts with the reported lysosomal and granular distribution; its precise cause cannot be assigned from appearance alone (UniProt Q9UBX1: lysosome; HPA: tissue IHC).	Recheck morphology, positive tissue, and no-primary control; repeat with the IHC-validated antibody if the discordance persists (general IHC practice).
Cells reported as Not detected stain as strongly as expected positives (HPA: adipocytes or bone-marrow hematopoietic cells).	Nonspecific binding or endogenous detection activity is possible; an HPA negative call remains cell-type specific (general IHC interpretation; HPA: tissue IHC).	Compare a no-primary control; if the detection chemistry uses an enzyme, check its endogenous-activity block, then reassess antibody conditions (general IHC practice).
Diffuse brown background obscures cell boundaries and granules (HPA: granular cytoplasmic pattern).	Excess primary antibody, incomplete washing, or prolonged chromogen development can raise background (general chromogenic IHC practice).	Inspect no-primary background, then adjust blocking, dilution, washing, and development using the validated IHC workflow (general IHC practice).
Colon or ovary appears inconsistent with UniProt tissue expression (UniProt Q9UBX1: tissue specificity).	UniProt's tissue-level expression and HPA's Not detected calls for colon glandular cells and ovarian stroma describe different scopes (UniProt Q9UBX1; HPA: tissue IHC).	Score the identified cell population and report the observed compartment and intensity; use HPA's cell-specific pattern as the comparison, with its medium RNA consistency caveat (HPA: tissue IHC/reliability).

Source: UniProt Q9UBX1 Human Protein Atlas tissue Human Protein Atlas subcellular

Experiment evidence

IHC & IF images showing CTSF staining

IHC analysis of Cathepsin F/CTSF using anti-Cathepsin F/CTSF antibody (A06600-1). Cathepsin F/CTSF was detected in a paraffin-embedded section of human stomach cancer 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-Cathepsin F/CTSF Antibody (A06600-1) 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.

A06600-1 · IHCIHC analysis of Cathepsin F/CTSF using anti-Cathepsin F/CTSF antibody (A06600-1). Cathepsin F/CTSF was detected in a paraffin-embedded secti

HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000174080-CTSF/tissue/cerebral+cortex#imid_720632

HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000174080-CTSF/tissue/colon#imid_720550

HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000174080-CTSF/tissue/liver#imid_720567

HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000174080-CTSF/tissue/kidney#imid_720552

HPA · ICC-IF A-549Human Protein Atlas immunofluorescence, A-549, antibody HPA031431; green = CTSF, blue = nucleus, red = microtubules.

← swipe / scroll for more images →

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

Sample controls for CTSF IHC & IF

🧪Run liver first: hepatocytes should stain (HPA: High in hepatocytes). Use adipose tissue as the negative tissue; adipocytes should show no specific staining (HPA: Not detected in adipocytes). On the liver slide, non-hepatocyte areas can serve as background checks, but their CTSF status is not established by the supplied HPA row.

Positive control tissue: Cerebellum (Cells in molecular layer, HPA High)

Negative control tissue: Adipose tissue (HPA Not detected)

ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show CTSF in A-549, SK-MEL-30, U2OS, with annotated localisation: Vesicles (supported) (HPA subcellular).

Technical controls: Include no-primary (secondary-only) and matched rabbit IgG isotype controls (caption: rabbit primary antibody); use CTSF knockout tissue as a biological negative if available (standard IHC practice). Check for endogenous peroxidase activity in liver before interpreting DAB signal, and block endogenous biotin if using biotin-based detection (standard IHC practice).

⚠️Feasibility: A target-specific fixation window or fixation effect is unreported, and the selected A06600-1 tissue-IHC caption does not state a fixative (caption: fixative not stated). The caption reports heat-mediated retrieval in EDTA at pH 8.0 for a paraffin section; it does not establish whether retrieval is required under other conditions (caption: EDTA retrieval). Frozen-section or tissue IF ease is unreported; HPA ICC-IF supports vesicular localization in A-549, SK-MEL-30 and U2OS cells (HPA subcellular: vesicles supported).

HPA tissue IHC evidence for CTSF

Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium 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
Cerebellum	Cells in molecular layer	High	Protein (IHC)	HPA →
Cerebral cortex	Neuronal cells	High	Protein (IHC)	HPA →
Hippocampus	Neuronal cells	High	Protein (IHC)	HPA →
Liver	Hepatocytes	High	Protein (IHC)	HPA →
Seminal vesicle	Glandular cells	High	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 →
Colon	Glandular cells	Not detected	Protein (IHC)	HPA →
Kidney	Cells in glomeruli	Not detected	Protein (IHC)	HPA →

Section 3

Advanced CTSF IHC Tips

Troubleshoot CTSF staining in paraffin sections by checking retrieval, compartment, cell type, controls, and scoring before interpreting chromogenic signal.

Which retrieval conditions should I start with for CTSF in paraffin sections?

Start with heat mediated antigen retrieval in EDTA at pH 8.0 for paraffin sections (datasheet A06600-1). The selected CTSF tissue image used that condition before overnight incubation at 4°C with 2 μg/ml primary antibody (datasheet A06600-1). If staining is weak, adjust heating time in small increments while keeping section thickness, detection, and exposure to DAB consistent across the comparison. Include a known positive section, such as liver hepatocytes (HPA: High in hepatocytes), and inspect tissue morphology after each adjustment. Stronger color alone is insufficient if granular cytoplasmic detail or tissue structure deteriorates (HPA: granular cytoplasmic pattern).

Could fixation explain weak or uneven CTSF staining?

Target specific sensitivity to fixation is unknown: the selected paraffin tissue caption does not state a fixative (datasheet A06600-1: fixative not stated). Record the fixative, fixation duration, tissue thickness, and processing history for each block before comparing CTSF intensity. For a controlled check, stain sections from comparably processed blocks in the same run using EDTA retrieval at pH 8.0 (datasheet A06600-1). Review morphology alongside signal, because damaged or poorly preserved tissue can make a chromogenic pattern difficult to score. Do not infer a CTSF fixation effect from its lysosomal location or glycosylation sites (UniProt Q9UBX1: lysosome; five glycosylation sites).

What CTSF staining pattern should I expect within positive cells?

Expect predominantly granular cytoplasmic staining in tissue sections (HPA: granular cytoplasmic expression in most tissues). A vesicular pattern is also supported by cell imaging, while endoplasmic reticulum and plasma membrane locations are uncertain (HPA subcellular: vesicles supported; additional locations uncertain). This fits a lysosomal protein without a transmembrane segment (UniProt Q9UBX1: lysosome; topology). Assess granules within intact cells rather than treating diffuse extracellular DAB or isolated nuclear color as equivalent evidence. Compare the pattern with a positive reference such as neuronal cells in cerebral cortex (HPA: High in cortical neuronal cells), and keep counterstaining light enough to see intracellular detail.

How might CTSF processing affect what this antibody detects?

CTSF is produced with a signal peptide at residues 1–19, a propeptide at 20–270, and a cathepsin F chain at 271–484 (UniProt Q9UBX1: processing). The record lists five glycosylation sites and no annotated isoforms (UniProt Q9UBX1: glycosylation; isoforms). Check the antibody's documented immunogen or mapped epitope before deciding whether staining should represent precursor, processed protein, or both; the selected tissue caption does not identify that epitope (datasheet A06600-1: tissue caption). For discordant blocks, compare cellular pattern and retrieval under matched conditions rather than attributing every intensity difference to processing. An epitope in a cleaved region could change what is visible, but that possibility requires epitope evidence.

How can I check a CTSF pattern by multiplex immunofluorescence?

Use IF as a separate validation experiment alongside the paraffin IHC result, pairing CTSF with a validated marker for the expected cell type; neuronal cells in cerebral cortex are a documented high staining population (HPA: High in cortical neuronal cells). Choose spectrally separated fluorophores and place the weaker signal in a channel with low tissue autofluorescence, checking unstained and single stain controls first. If the epitope is intracellular, optimise permeabilisation to admit antibody while preserving vesicle detail; CTSF is lysosomal and lacks a transmembrane segment (UniProt Q9UBX1: location; topology). Compare puncta within marked cells with secondary only controls and with the chromogenic granular pattern (HPA: granular cytoplasmic expression).

What should I adjust when CTSF DAB staining is diffuse?

First inspect a no primary control for background from secondary detection and endogenous peroxidase; use a peroxidase blocking step where needed as standard chromogenic IHC practice. The selected tissue image used 10% goat serum blocking, 2 μg/ml primary antibody, and DAB detection (datasheet A06600-1). Keep incubation, washes, DAB development, and section handling consistent while changing one variable at a time. Diffuse staining that obscures granular cytoplasmic detail is less convincing for CTSF (HPA: granular cytoplasmic expression in most tissues). Compare the suspected background with cell specific staining in a positive reference, such as liver hepatocytes (HPA: High in hepatocytes).

How should I quantify CTSF staining across paraffin samples? ⚠ ANSWER MARKED FOR VERIFICATION

Define the cell population and region before scoring; for example, hepatocytes have high reported staining, whereas colon glandular cells are not detected in the HPA tissue panel (HPA: High in hepatocytes; Not detected in colon glandular cells). Score the percentage of positive cells and intensity with an H-score, or measure positive cell density per mm² when cell counts are more relevant. Normalise positive counts to the number of eligible cells, or stained area to the viable tissue area, using the same rule for every section. Exclude folds, torn edges, necrosis, and nonspecific DAB deposits by a predefined review rule. Report compartment and granular pattern alongside the numerical score (HPA: granular cytoplasmic expression).

How do I distinguish genuine CTSF staining from artefact?

Give greatest weight to granular cytoplasmic signal within intact cells, consistent with tissue staining and lysosomal localisation (HPA: granular cytoplasmic expression; UniProt Q9UBX1: lysosome). Check whether stained cells match the tissue context: liver hepatocytes and cortical neuronal cells are reported high, while adipocytes are not detected in the HPA panel (HPA: High in hepatocytes and cortical neuronal cells; Not detected in adipocytes). Treat staining confined to cut edges, necrotic zones, or extracellular deposits as suspect, and compare it with a no primary control. Rule out endogenous peroxidase contribution with an appropriate peroxidase block as standard chromogenic IHC practice. Interpret an isolated membrane or nuclear pattern cautiously because vesicles are the supported cell imaging location (HPA subcellular: vesicles supported).

Boster reagents

Best CTSF / Cathepsin F IHC Antibodies

A06600-1 has real IHC data from human paraffin sections of stomach and thyroid cancer tissue (catalog IHC captions); no IF/ICC data are listed (catalog applications and images).

Real IHC data

Anti-Cathepsin F/CTSF Antibody ®

Cat # A06600-1

View product →

A06600-1 is listed for human IHC (catalog applications and reactivity). Its IHC captions show staining in paraffin sections of human stomach and thyroid cancer tissue (catalog IHC captions).

Which to pick: Choose A06600-1 for human paraffin-section IHC; its listed IHC concentration is 2–5 μg/ml (datasheet), and its captions document staining at 2 μg/ml after EDTA pH 8.0 retrieval (catalog IHC captions). The captions do not report the fixative (catalog IHC captions). No listed SKU supports an IF/ICC or cross-species recommendation: A06600-1 lists human reactivity and IHC, but no IF/ICC application or image (catalog applications, reactivity and images).

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

References

UniProt Consortium. UniProt entry Q9UBX1 (CATF_HUMAN, Cathepsin F).
Human Protein Atlas. CTSF tissue IHC expression (reliability: Enhanced).
Human Protein Atlas. CTSF subcellular location (ICC-IF): Mainly localized to vesicles. In addition localized to the plasma membrane and endoplasmic reticulum..
Human Protein Atlas. CTSF antibody validation summary (2 antibodies).
Adipose-derived stem cells alleviate radiation-induced dermatitis by suppressing apoptosis and downregulating cathepsin F expression. Stem cell research & therapy 2021 — PMC8351374.
Overexpression of cathepsin F, matrix metalloproteinases 11 and 12 in cervical cancer. BMC cancer 2005 — PMC1175083.
Cathepsin F and Fibulin-1 as novel diagnostic biomarkers for brain metastasis of non-small cell lung cancer. British journal of cancer 2022 — PMC9174239.
Cathepsin F is a potential marker for senescent human skin fibroblasts and keratinocytes associated with skin aging. GeroScience 2023 — PMC9886782.
PubMed PMID:10318784 — UniProt-cited evidence.
PubMed PMID:10198209 — UniProt-cited evidence.
PubMed PMID:10661872 — UniProt-cited evidence.