Calcofluor White Staining 50 FAQs and 30 MCQs
Master fungal diagnostics with our ultimate Calcofluor White Staining resource! This comprehensive guide includes:
🔍 50 Essential FAQs Covering:
✓ Principle: How Calcofluor binds to fungal chitin/cellulose
✓ Protocol: Step-by-step staining for Candida, Aspergillus, Pneumocystis
✓ Microscopy: Optimal UV filter sets (DAPI vs. FITC)
✓ Troubleshooting: Background fluorescence, false positives
✓ Clinical Apps: Rapid fungal detection in KOH mounts
📝 30 Challenging MCQs On:
◼️ Stain preparation (KOH/CFW ratios)
◼️ Distinguishing hyphae vs. pseudohyphae
◼️ Artifact identification (cotton fibers, starch)
◼️ Comparison to PAS/GMS stains
🎯 Perfect For:
• Medical mycologists • Clinical lab technicians • USMLE/PLAB aspirants
Calcofluor White Staining 50 FAQs:
What is Calcofluor White Stain?
A fluorescent dye that binds to chitin and cellulose in fungal and parasitic cell walls.
What does Calcofluor White Stain detect?
Fungi, yeasts, and parasites like Pneumocystis, Acanthamoeba,
and Microsporidium.Is Calcofluor White specific to fungi?
No, it binds to any structure containing chitin or cellulose (e.g., plant cells, cotton fibers).
What is the principle of Calcofluor White Staining?
It binds to β-1,3 and β-1,4 polysaccharides (chitin/cellulose) and fluoresces under UV light.
What color does Calcofluor White Stain produce under UV light?
Fungal and parasitic elements fluoresce apple-green, while other materials appear reddish-orange.
How is Calcofluor White Stain prepared?
Dissolve 1g Calcofluor White powder in 100 mL distilled water (1% solution).
Why is Potassium Hydroxide (KOH) added?
KOH clears debris, improving fungal element visibility.
What is the role of Evans Blue in staining?
It reduces background fluorescence and enhances contrast.
How long does the staining process take?
Approximately 1–5 minutes.
What magnification is used for observation?
100X–400X under a fluorescent microscope.
Can Calcofluor White Stain be autoclaved?
Yes, it withstands autoclaving (121°C for 15 minutes).
How should the stain be stored?
At room temperature, protected from light.
What fungal infections can Calcofluor White detect?
Candida, Histoplasma, Pneumocystis, and dermatophytes.
Can it detect parasites?
Yes, including Acanthamoeba, Naegleria, and Microsporidium.
Is it useful for diagnosing Pneumocystis jirovecii?
Yes, it highlights cysts in bronchoalveolar lavage (BAL) samples.
Can it be used for Acanthamoeba keratitis diagnosis?
Yes, from corneal scrapings.
Does it work on yeast bud scars?
Yes, it stains chitin-rich bud scars more intensely.
Can it differentiate live vs. dead fungal cells?
No, it stains both live and dead cells with chitin.
Is it used in histopathology?
Yes, for rapid fungal detection in tissue samples.
Can it be used in textile industries?
Yes, as a fabric brightener.
What does apple-green fluorescence indicate?
Presence of fungi or parasites.
What does reddish-orange fluorescence mean?
Non-specific background or other biological material.
Why do cotton fibers fluoresce strongly?
They contain cellulose, which binds Calcofluor White.
Do Pneumocystis cysts show any unique features?
They appear as round, 5–8 µm cells with a bright cyst wall.
Can yeast cells be distinguished from Pneumocystis?
Yes, yeast shows budding and deeper internal staining.
Do amoebic trophozoites fluoresce?
No, only cysts fluoresce.
What causes yellowish-green background in tissues?
Non-specific binding; fungi appear brighter.
How can background fluorescence be reduced?
Using Evans Blue or adjusting microscope filters.
What are the advantages of Calcofluor White Staining?
Fast, sensitive, and works on non-culturable fungi.
What are its limitations?
Requires a fluorescent microscope (expensive).
Is it more sensitive than KOH wet mount?
Yes, but KOH is cheaper and faster for routine use.
Can it replace culture methods?
No, but it aids in rapid preliminary diagnosis.
Why is it not used in all labs?
Due to the cost of fluorescent microscopes.
Does it work on all fungi?
Mostly, but some may require additional tests.
What is the excitation wavelength for Calcofluor White?
347 nm (UV light), but violet/blue light also works.
What is the emission wavelength?
475 nm (apple-green fluorescence).
Why is UV light preferred?
It provides maximum fluorescence intensity.
Can brightfield microscopy be used?
No, only fluorescence microscopy works.
What if the fluorescence is too weak?
Check stain freshness, microscope filters, and light source.
Can old Calcofluor White Stain be used?
Yes, if stored properly (up to 1 year).
How does Calcofluor White compare to Gram stain?
Gram stain differentiates bacteria; Calcofluor targets fungi/parasites.
Is it better than PAS (Periodic Acid-Schiff) staining?
PAS is more specific for fungi in tissues, but Calcofluor is faster.
Can it be combined with other stains?
Yes, with KOH or Evans Blue for better clarity.
Why was it replaced by KOH in some labs?
KOH is cheaper and doesn’t need a fluorescent microscope.
Can it detect Cryptosporidium?
Yes, but modified acid-fast staining is more common.
Does it work on Dirofilaria larvae?
Yes, it can stain some nematode larvae.
Can it be used in plant pathology?
Yes, for detecting fungal infections in plants.
Is it used in environmental mycology?
Yes, for studying fungal growth in slide cultures.
Does it stain ascospores?
No, only vegetative fungal cells.
Are there any safety concerns with Calcofluor White?
It is generally safe but should be handled with standard lab precautions.
Calcofluor White Staining 30 MCQs:
- What does Calcofluor White primarily bind to?
a) Proteins
b) Lipids
c) Chitin and cellulose
d) DNA - Which type of microscope is required for Calcofluor White Staining?
a) Brightfield
b) Fluorescent
c) Phase-contrast
d) Electron - What color do fungi fluoresce under UV light with Calcofluor White?
a) Red
b) Blue
c) Apple-green
d) Yellow - Which of the following is NOT detected by Calcofluor White?
a) Candida albicans
b) Pneumocystis jirovecii
c) Acanthamoeba cysts
d) Bacterial cells - Why is Evans Blue used as a counterstain?
a) To kill microbes
b) To reduce background fluorescence
c) To enhance fungal growth
d) To dissolve chitin
- What is added to Calcofluor White for clearing debris in skin/nail samples?
a) HCl
b) 10% KOH
c) NaCl
d) Ethanol - How long should the stain be left before observation?
a) 10 seconds
b) 1 minute
c) 30 minutes
d) 1 hour - What is the optimal magnification for observing Calcofluor White-stained samples?
a) 40X
b) 100X–400X
c) 1000X
d) 2000X - Which wavelength is best for exciting Calcofluor White?
a) 200 nm
b) 347 nm (UV light)
c) 550 nm
d) 700 nm - How should Calcofluor White Stain be stored?
a) Frozen
b) At room temperature, protected from light
c) In direct sunlight
d) Under vacuum
- Which parasite’s cysts are detected by Calcofluor White?
a) Plasmodium
b) Acanthamoeba
c) Giardia
d) Entamoeba histolytica - What structure in yeast is intensely stained by Calcofluor White?
a) Nucleus
b) Mitochondria
c) Bud scars
d) Ribosomes - Why do cotton fibers interfere with interpretation?
a) They dissolve in KOH
b) They fluoresce strongly (cellulose content)
c) They inhibit fungal growth
d) They turn black - Which fungal infection is rapidly diagnosed using Calcofluor White?
a) Onychomycosis
b) Tuberculosis
c) Hepatitis B
d) Malaria - What is the size range of Pneumocystis cysts observed with Calcofluor White?
a) 1–2 µm
b) 5–8 µm
c) 10–15 µm
d) 20–30 µm
- What is the main advantage of Calcofluor White Staining?
a) Rapid detection of fungi/parasites
b) Cheaper than KOH
c) Works without a microscope
d) Stains only dead cells - What is a major limitation of this technique?
a) Requires a fluorescent microscope
b) Only works on bacteria
c) Takes 24 hours for results
d) Cannot detect yeast - Which of the following is a false-positive artifact in Calcofluor White Staining?
a) Cotton fibers
b) Fungal hyphae
c) Candida cells
d) Pneumocystis cysts - Why is KOH often preferred over Calcofluor White in some labs?
a) Lower cost and no need for a fluorescent microscope
b) Better for bacterial staining
c) Works faster (5 seconds)
d) Doesn’t require a coverslip - Which organism’s TROPHOZOITES do NOT fluoresce with Calcofluor White?
a) Candida
b) Acanthamoeba
c) Aspergillus
d) Cryptococcus
- Which component of fungal cell walls binds Calcofluor White?
a) Chitin
b) Ergosterol
c) Mannoproteins
d) Phospholipids - What is the emission peak wavelength of Calcofluor White?
a) 300 nm
b) 475 nm (apple-green)
c) 600 nm
d) 700 nm - Which of these is a non-fungal application of Calcofluor White?
a) Textile brightening agent
b) Gram staining
c) Acid-fast bacilli detection
d) Viral culture - Which counterstain reduces background fluorescence in Calcofluor White?
a) Safranin
b) Evans Blue
c) Methylene blue
d) Iodine - What is the shelf life of properly stored Calcofluor White Stain?
a) 1 week
b) 1 month
c) 1 year
d) 10 years - Which light source is LEAST effective for Calcofluor excitation?
a) Red light
b) UV light
c) Violet light
d) Blue-violet light - What does apple-green fluorescence indicate in a BAL sample?
a) Bacteria
b) Pneumocystis cysts
c) Red blood cells
d) Epithelial cells - Which fungal structure does NOT stain with Calcofluor White?
a) Hyphae
b) Yeast cells
c) Ascospores
d) Pseudohyphae - Why is Calcofluor White used in Papanicolaou (Pap) smears?
a) To detect HPV
b) To enhance yeast cell visibility
c) To stain nuclei
d) To fix cells - Which industry uses Calcofluor White as a fabric whitener?
a) Textile
b) Pharmaceutical
c) Food
d) Automotive
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