Which stain use in CSF examination
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stains used in CSF examination microbiology
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Here is a thorough, textbook-backed summary of all the stains used in CSF examination:
Stains Used in CSF (Cerebrospinal Fluid) Examination
1. Gram Stain - Most Important Routine Stain
The Gram stain is the cornerstone of CSF microbiology. All CSF specimens should be concentrated by centrifugation before staining. Sensitivity ranges from 60-90% depending on organism type and concentration:
- Gram-positive: Streptococcus pneumoniae, Listeria monocytogenes, Group B strep
- Gram-negative diplococci: Neisseria meningitidis
- Gram-negative bacilli: E. coli, H. influenzae
Note: Sensitivity for Listeria and gram-negative bacilli can be as low as 50% or less.
Fig. 30.10 - CSF Gram stain showing gram-negative diplococci characteristic of N. meningitidis (Henry's Clinical Diagnosis, p. 193)
2. India Ink (Negative Stain) - For Cryptococcal Meningitis
Used to detect Cryptococcus neoformans/C. gattii in CSF. India ink particles cannot penetrate the polysaccharide capsule, so the capsule appears as a clear halo around the yeast cell against a dark background. Sensitivity is less than 50% (especially in non-HIV patients), so the cryptococcal antigen (CrAg) test has largely replaced it - but India ink remains useful when antigen testing is unavailable.
Fig. 60.14 - India ink preparation of CSF with C. neoformans (Henry's Clinical Diagnosis, p. 2114)
3. Acridine Orange Stain - Fluorescent/Supplementary
A fluorescent stain used when Gram stain is negative but clinical suspicion remains high (e.g., many PMNs but no organisms seen). More sensitive than Gram stain for:
- Bacteria and fungi (appear bright orange/red under fluorescence)
- Gram-negative organisms
- Differentiates amoebas (brick red) from leukocytes (bright green) in primary amoebic meningoencephalitis (Naegleria fowleri)
4. Ziehl-Neelsen (ZN) / AFB Stain - For TB Meningitis
Used when tuberculous meningitis is suspected. Detects Mycobacterium tuberculosis as acid-fast bacilli (appear red on blue background). Sensitivity in CSF is low (10-40%), so culture and PCR are also essential.
5. Giemsa / Wright-Giemsa Stain
Used for:
- Parasites: Trypanosoma species (African sleeping sickness), Naegleria, Acanthamoeba
- Malignant cells in CSF cytology
- Also helps identify cell morphology in the CSF differential
6. KOH (Potassium Hydroxide) Preparation
Used for fungal elements - clears cellular debris and highlights fungal hyphae and yeast. Can be combined with Calcofluor white (fluorescent) for enhanced visibility.
Summary Table
| Stain | Target Organism/Purpose | Key Feature |
|---|---|---|
| Gram stain | Bacteria (meningitis) | 60-90% sensitivity; most routine |
| India ink | Cryptococcus (fungi) | Negative stain; halo capsule; <50% sensitivity |
| Acridine orange | Bacteria, fungi, amoeba | Fluorescent; more sensitive than Gram |
| ZN / AFB stain | Mycobacterium tuberculosis | Acid-fast; low CSF sensitivity |
| Giemsa/Wright | Parasites, malignant cells | Cell morphology, trypanosomes |
| KOH ± Calcofluor white | Fungal elements | Clears debris; shows hyphae/yeast |
- Henry's Clinical Diagnosis and Management by Laboratory Methods, pp. 193-195, 2114-2135
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