1. Lab Medicine Conference : Cerebrospinal Fluid Analysis

2. Cerebrospinal Fluid (CSF)
Adults produce 450 to 500 cc per day
150 cc in adult CNS at any one time
Neonates have 30 to 60 cc
Children have 100 cc
80 % produced by ventricular choroid plexuses
Reabsorbed by arachnoid villi
Drains into dural sinuses

3. Suspected Diagnoses for Which CSF Exam is Indicated
Meningitis
Encephalitis
Brain abscess
Neurosyphilis
Subarachnoid hemorrhage
Demyelinating conditions :
Multiple sclerosis
Guillian-Barre
CNS malignancies

4. Usual Recommended Tests to Run on Sequential Tubes of CSF from an LP
First and third tubes
Cell count & differential
Second tube
CSF total protein, glucose, +/- other chemistries
Fourth tube
Gram stain, other stains, cultures

5. Priority Ranking of Tests to Run If Only Small Amount of CSF Obtained
Gram stain / culture
Cell count / differential
Protein / glucose
Chemistries

6. Contraindications to Lumbar Puncture
Intracranial mass lesion with impending herniation
Cutaneous infection or suspected subcutaneous abscess at LP site
Systemic coagulopathy
Could result in cord compression from para-spinal hematoma
Unrestrainable patient

7. Potential Complications of Lumbar Puncture
Uncal or brainstem herniation
0.3 to 1.2 % mortality if papilledema present
less likely if smaller amounts of fluid removed
Arachnoiditis : can occur if needle carries in povidone-iodine
Epidermoid tumors (delayed) : from use of needle without stylet
Nerve root injury : less likely if needle bevel vertical
Induced meningitis ; paraspinal abscess
Mortality
from hyperflexion of head & tracheal obstruction
or from vagally induced asystole
Post-procedure headache : 12 to 39 %

8. CSF Exam First step is measure the opening pressure (OP) :
normal 80 to 180 mm H2O with pt. recumbent
can be "falsely" elevated by Valsalva, head-up position, or jugular compression
should vary 5 to 10 mm H2O with respiration
Queckenstedt & Tobey Ayer tests (involving jugular compression & seeing the effect on OP) are no longer recommended

9. Causes of Elevated CSF Opening Pressure
Meningitis
Intracranial mass lesions
SAH
CHF
SVC obstruction
Thrombosis of intracranial venous sinus
Acute elevation of serum osmolarity

10. Causes of Low CSF Opening Pressures
Severe dehydration
Circulatory collapse
Chronic serum hyperosmolality
Dural tears with CSF leak
Neurosurgical procedures
Subdural hematomas in elderly
Barbiturate intoxication
Complete spinal subarachnoid block

11. CSF Appearance Normal is clear & consistency similar to H2O
Causes of visual turbidity :
> 200 WBC's per mm3
> 400 RBC's per mm3
Bacteria
Aspirated epidural fat
Evil aliens (this was to see if you are paying attention)

12. Causes of CSF Clot Formation
Traumatic tap
Increased protein from :
subarachnoid block
neurosyphilis
tuberculosis
Metastatic mucinous adenocarcinoma of the meninges

13. Xanthochromia of the CSF
Is yellow - orange - brown coloration in supernatant of centrifuged CSF
Produced by lysis of red cells
Involves 3 pigments :
oxyhemoglobin (red) : occurs in CSF within 2 hours of a SAH
bilirubin (yellow) : converted from hemoglobin in 12 hours
methemoglobin (brown)

14. Causes of Xanthochromia Besides Red Cell Lysis
Direct serum bilirubin levels > 10 to 15 mg %
CSF protein levels > 150 mg %
Sample contamination with povidone iodine
Systemic hypercarotenemia
CSF melanin from meningeal melanosarcoma

15. CSF Glucose Normally 60 to 70 % of serum level
Is 100 % ratio in neonates (immature CSF / blood barrier)
In adults with serum glucose > 300 mg %, no further increase in CSF glucose occurs
CSF level takes 2 hours to equilibrate with change in serum glucose

16. Causes of Hypoglycorrhachia
(CSF to Serum glucose ratio < 0.6)
Systemic hypoglycemia
Impaired glucose transport
Increased CNS use of CSF
Increased use of CSF glucose by bacteria & leucocytes
Typical with bacterial, tuberculous, or fungal meningitis
Also sometimes with SAH, viral meningitidies, sarcoidosis, neoplasms

17. CSF Protein Normal adult range is 17 to 55 mg %
Normal neonate level is up to 150 mg %
Increased levels usually associated with CNS inflammatory processes, especially infections
Has relation ratio with serum protein levels, so elevations of serum protein may cause elevations in CSF protein

18. Noninfectious Causes of Elevated CSF Protein
Traumatic LP
1 mg % increase per 1000 RBC's per mm3
Interference with CSF / blood barrier
Cerebral hemorrhage
SAH
Cerebral thrombosis
Endocrine
Diabetes mellitus
Hyperthyroidism
Hypoparathyroidism
Hyperadrenalism

19. Other Noninfectious Causes of Elevated CSF Protein
Guillian-Barre Syndrome
Multiple sclerosis
Collagen vascular diseases
Subacute sclerosing panencephalitis
Mechanical obstruction of CSF circulation
tumors, abscesses, cord compression
Elevated serum protein levels (multiple myeloma, etc.)
Medications / toxins :
Phenytoin, ethanol, heavy metals

20. Causes of Low CSF Protein Levels
Chronic leakage from CSF otorrhea or rhinorrhea
Chronic increased ICP
Removal of CSF via neurosurgical procedures or repeated LP's

21. CSF Cell Counts Normal adult : 0 to 5 lymphs or monos
Even one poly is abnormal
Normal neonates have 0 to 30 cells & up to 60 % polys
Increased neutrophils usually indicate infectious process

22. Comparisons of Cell Counts in Viral Versus Bacterial Meningitis
Typically > 500 WBC's / mm3 & mainly polys
10 % of cases have < 50 % polys
Viral
Typically < 100 WBC's / mm3 & mainly monos
10 % of cases have > 50 % polys (especially if early)
90 % convert to mononuclear pleocytosis by 12 hours

23. Infectious Causes of Very Low CSF Cell Counts
Meningitis from :
Neisseria meningitidis
Hemophilus influenzae
Overwhelming Strep. pneumoniae infection

24. Causes of Increased Neutrophils in the CSF
Infectious
Bacterial meningitis
Early tuberculous meningitis
Early viral meningitis
Early mycotic meningitis
Noninfectious
3 to 4 days post - hemorrhagic infarct
SAH or intracerebral hematoma
Injection of antibiotics or antimetabolites
Injection of contrast media
Repeated LP's

25. Causes of Increased Lymphocytes in the CSF
Infectious
Tuberculous, fungal, or leptospiral meningitis
Partially treated bacterial meningitis
Viral or syphilitic meningoencephalitis
Subacute sclerosing panencephalitis
Measles
Noninfectious
Multiple sclerosis, Guillian-Barre Syndrome
Polyneuritis
Temporal arteritis or periarteritis
Chronic ethanol abuse
Intravenous drug abuse

26. Causes of Increased Eosinophils in the CSF
Infectious
Bacterial, fungal, or viral meningitis
Cysticercosis
Noninfectious
Allergic reaction to foods, meds, dyes, or envenomation
Intrathecal foreign substances or contrast dye
Synthetic intrathecal shunts
Periarteritis nodosa
Allergic bronchial asthma
Acute polyneuritis
Rabies vaccination

27. Causes of Increased Macrophages in the CSF
Infectious
Tuberculosis
Noninfectious
Presence of erythrocytes
Acute intracranial bleeding
Mycotic meningitis
Trauma to CNS
Contrast media

28. Age Related Causes of Bacterial Meningitis

29. Intersection with line B
Intersection with line B. Join the marks on lines A & B with the ruler, and read off the probability of acute bacterial versus acute viral meningitis where the ruler intersects the central probability scale.

30. CSF Gram Stain Should be done on uncentrifuged CSF if CSF cloudy
Should be done on centrifuged CSF if CSF clear
Identifies 80 % of bacterial CSF infections
False positive only if LP tray or stain itself is contaminated

31. CSF gram stain showing E. coli

32. CSF gram stain showing Listeria monocytogenes

33. CSF gram stain showing Neisseria meningitidis

34. CSF gram stain showing Streptococcus pneumoniae

35. CSF gram stain showing Staphylococcus aureus

36. CSF gram stain of Pneumococcal meningitis

37. Use of Acrinidine Orange Stain (AOS) for CSF
Is fluorochrome stain for bacterial nucleic acids
Bacteria stain bright orange
Background of cellular debris stains yellow - pale green
Takes 2.5 minutes to prepare (versus 3.5 minutes for gram stain)
Useful if bacteria not seen on gram stain (increases pickup rate > 75 %)

38. Other CSF Tests for Meningitis
Lactic acid
Levels > 35 mg % in 90 % of bacterial meningitis
Numerous false positives (neoplasm, injury, etc.)
LDH
Elevated (especially LDH-5) with bacterial meningitis, but is nonspecific
C-reactive protein
If elevated has high sensitivity & specificity for bacterial meningitis, but is a technically difficult assay
Quelling Reaction
Antisera cause swelling in pneumococci & Hemophilus influenzae

39. Other CSF Tests for Meningitis (cont.)
Limulus amebocyte lysate assay
Requires 60 minutes
Not 100 % sensitive
CSF amino acids
Elevated with bacterial meningitis
May be useful for dx if partial treatment
Countercurrentimmunoelectrophoresis CIE)
Takes 30 to 60 minutes
Precipitant line forms between bacterial antigens & serum with known antibodies
Can be useful in partially treated meningitis
False positives & cross-reactions occur

40. Causes of False Negative CIE
Amount of antigen too small (if < 10,000 bacteria per ml.)
If infection early, not enough time for antigen to dissolve off the bacteria
Poor antibody quality for some strians (as for group B meningococcus & pneumococci types 7 & 14)

41. Sensitivity of CIE in Meningitis
Meningococcal : 50 to 90 %
Strep. pneumoniae : 50 to 100 %
Hemophilus influenzae : 80 %
Group B strep : 60 to 90 %

42. Latex Agglutination Antigen Tests for Meningitis
More sensitive than CIE for pneumococci & meningococci
Only takes 15 minutes to perform
Not affected by antigen excess
Less false negatives than CIE

43. Other Tests to Consider for Suspected Non-Bacterial, Non-Viral Meningitis
Acid fast stain
Mycobacterial culture
India ink prep (for Cryptococcus)
Cryptococcal antigen
Fungal culture

44. Charges at H.M.C. for CSF Cultures & Microbial Stains
Gram stain & culture : $ 48
Sensitivity (antibiotic) : $ 45 to $ 105
Agar diffusion vs. dual vs. add anerobic
Fungal smear : $ 21
Fungal culture : $ 48
AFB smear & culture : $ 50
CIE : $ 37

45. Charges at H.M.C. for Other Standard Studies on CSF
Cell count & diff. : $ 67 (stat)
Glucose (stat) : $ 35
Protein (stat) : $ 35
Cryptococcal antigen : $ 35
Lactate : $ 26

46. Charges at H.M.C. for Miscellaneous Studies on CSF
Darkfield exam : $ 54
VDRL : $ 16
India ink prep : $ 22
IgG : $ 20
Immunochemistry eval. : $ 126
ph by electrode : $ 26
Sperm count (rule out sperm embolus) : $16

47. Total Charges at H.M.C. for Different Patterns of CSF Test Ordering
CBC/diff., gm. stain / culture, glucose, protein : $ 185
All standard, & culture / sensitivity studies : $ 322
All standard, & culture / sensitivity, & misc. studies : $ 462

48. Summary of Lab Studies on CSF for Meningitis
Measure opening pressure
Send four tubes
Check gram stain
If gram stain negative :
Consider AOS
Consider CIE +/- LA
If clinical suspicion for meningitis, start broad spectrum antibiotics prior to initial lab results