The cerebrospinal fluid (CSF) is formed by the choroid plexuses within the ventricles, passes into the subarachnoid space around the spinal cord and finally over the surface of the brain, to be absorbed in the arachnoid villi in the sagittal sinus. The composition of the CSF changes during circulation from its creation to its final absorption. For example, the CSF protein is lowest in the lateral ventricles, intermediate in the lumbar subarachnoid space, and highest in the subarachnoid spaces over the surface of the cerebral hemispheres (where it is occasionally sampled in mistake for subdural fluid). Except when there is obstruction to the flow of CSF from the ventricular system or potential downward herniation of the brain, CSF is obtained by the lumbar route.
While performing a lumbar puncture needles must contain stylets to avoid the possibility of inducing implantation dermoids and leading to paraparesis some years later.
- A cloudy appearance normally indicates infection.
- More important is the distinction between traumatic haemorrhage and subarachnoid bleeding. In subarachnoid haemorrhage the CSF, however red, runs like CSF and does not have the viscosity of blood.
- Xanthochromia of the supernatant may reflect bleeding several hours previously but a similar colour is seen when the protein content is very high from any cause.
- A shimmer' on flicking the tube suggests increased cells and/or protein >1.5g/L.
The opening pressure should always be measured using a manometer attached to the needle.
Pressure measurements obtained by the usual manometric method will be in centimetres of water (cmH2O), but those obtained with the aid of a transducer will be in milimetres of mercury (mmHg). The conversion factor is approximately 1.3, by which the pressure in mmHg must be multiplied to give the value in cmH2O.
The normal CSF pressure in a neonate is in the range 0-5.7mmHg, mean 2.8mmHg (0-7.6cmH2O), mean 3.8cmH2O). The upper limit of the CSF pressure in older children is said to be similar to the adult value of 14mmHg (19cmH2O). The upper limit in the infant must be lower but information is limited.
In the neonatal period and later, a white blood cell (WBC) count up to 5/mm3 is normal. The distribution is skewed to smaller numbers, such that the median neonatal WBC count is 1/mm1. It is wise to regard even 1 neutrophil/mm3 as abnormal. The recognition of CSF malignant cells requires specialist expertise, but eosinophils are detectable with standard staining.
In the evaluation of CSF cell count when there has been a traumatic lumbar puncture haemorrhage, one retrospective study found that the CSF WBC count was only 20% of that expected by the number of red cells. However, inferences based on such calculations must be made with caution.
The protein in the CSF consists mostly of albumin, synthesized entirely extrathecally, and immunoglobulins - IgG, etc. - synthesized both extrathecally and intrathecally.
Many laboratories even in children's hospitals give the upper limit of normal as the adult value of 0.45g/L (45mg/dL) total protein. In fact, the total protein begins at a high level in the neonatal period and falls to a plateau between 6 months and 10 years, followed by a slight increase up to the age of 16 years. For most of infancy and childhood the upper limit of normal is about 0.32g/L (32mg/dL).
Increase in CSF protein (albumin) occurs when there is disruption of the blood-brain barrier (and notably in polyneuropathy).
Traumatic blood contamination may be expected to increase the CSF protein by around 0.01lg/L for each 1000 red cells/uL but, as with similar calculations on cell counts, caution is indicated.
Because albumin comes entirely from outside the CSF, its concentration reflects the blood-CSF barrier. The CSF:serum albumin quotient (Qalb) = CSF albumin (mg/L)/serum albumin (g/L). A Qalb >9 indicates blood-CSF barrier impairment.
Immunoglobulin - quantitative indices
Although immunoglobulins may be increased because of disruption of the blood-brain barrier an important cause is intrathecal synthesis of immunoglobulin. Various formulae or indices are used to give a quantitative measure of intrathecal synthesis of IgG, the most important of the immunoglobulins.
The CSF IgG:albumin ratio = CSF IgG (mg/L)/CSF albumin (mg/L).
The upper limit of normal for children under 18 months is 0.17, and for children over 18 months 0.22. Values above these figures suggest intrathecal IgG synthesis.
The CSF:serum IgG quotient (QlgG) = CSF IgG (mg/L)/serum IgG (g/L).
The 'normal' upper limit for the CSF:serum IgG index is 1.02 under the age of 18 months and 0.78 over that age, a higher value indicating increased intrathecal IgG synthesis. This is an index of focal IgG production, adjusted both for leakage of IgG due to blood-brain barrier dysfunction and for serum albumin and IgG concentration. With these upper limits, false positives occur in ~2% of cases.
In the above calculations the degree of intrathecal IgG synthesis may be falsely elevated when measurements are made after a traumatic lumbar puncture.
Immunoglobulin - qualitative: oligoclonal bands
The presence of two or more bands of immunoglobulin on isoelectric focusing (IEF) of CSF is referred to as 'oligoclonal bands' (OCBs). This CSF oligoclonal banding is only indicative of local CNS IgG production when the banding pattern in the CSF is distinct from the pattern seen in the serum. For this reason a serum sample must be sent with a CSF sample, as is done of course for the CSF:serum IgG index.
While OCBs are well known to assist in the diagnosis of multiple sclerosis it is important to recognize that they are found in many other situations with infections (subacute sclerosing panencephalitis, rubella panencephalitis, neurosyphilis, neuroborreliosis) or autoimmunity (anti-NMDA receptor encephalitis).
Immunoglobulin - quantitative and qualitative
Although it has been suggested that in adults - where multiple sclerosis is a predominant condition - it is only necessary to request IEF for OCB, in children it is wise to request both quantitative (CSF:serum IgG index) and qualitative (isoelectric focusing for OCB) measures to determine whether there is local CSF IgG synthesis.
1 Also known as CSF lgG index or simply as IgG index.
The proportion of asialotransferrin (desialotransferrin, tau protein, t protein) to total transferrin may be measured simply, quickly and cheaply by 2D gel electrophoresis. Normally asialotransferrin forms more than 8% of CSF transferrin, but in elF2B-relaled disorders (vanishing white matter disease and several other phenotypes) it is below 8%. Although this test has not yet been validated in infancy, from toddlerhood onwards it has a high degree of sensitivity and specificity to predict mutations in the genes responsible for one of the five subunits of eII'2B. It may be a first-line investigation once a leukoencephalopathy has been detected on MRI/MRS.
Myelin basic protein
This component of myelin may be found in increased quantities at the time of acute demyelination (as in acute disseminating encephalomyelitis), but data on sensitivity and specificity are limited.
Although cytokines may be estimated fairly readily by means of commercially available kits, such estimations are not particularly helpful in clinical diagnosis.
The most important cytokine that is involved in certain virus infections and autoimmune disorders is interferon-alpha (IFN-α). The normal IFN-α value is less than 2 IU/mL A serum sample is helpful but not always necessary for interpretation of CSF IFN-α. CSF IFN-α is raised acutely in herpes encephalitis and in cerebral systemic lupus erythematosus. It is consistently elevated for the first 12 months of life in Aicardi-Goutieres syndrome.
The normal CSF glucose is two-thirds of the blood glucose level, which must always be estimated simultaneously, the blood sample taken immediately before the lumbar puncture.
Various degrees of low CSF glucose may be found in all inflammatory meningeal disorders whether infective or autoimmune, and in malignant meningitis. Low CSF glucose may also be seen after subarachnoid haemorrhage.
An important cause of low CSF glucose is GLUT1 deficiency. This is an important treatable cause of early-onset epileptic seizures, hypotonia, movement disorders, learning difficulties and microcephaly. A 4-6 hour fast is recommended with a blood glucose measurement immediately before the lumbar puncture for the CSF glucose estimation. In GLUT1 deficiency the CSF/blood glucose ratio has a mean of 0.35 with a range 0.19-0.49, and the lactate is always normal or low.
Paired samples of fasting serum and CSF are taken for lactate. The upper limit of normal is usually given as 2.4mmol/l. for serum and 2.2mmoI/L for CSF lactate. CSF lactate is often but by no means always increased in mitochondrial disorders, and in certain organic acidaemias. It is normal or reduced in GLUT] deficiency.
In certain circumstances the lactate/pyruvate (L/P) ratio is estimated. It is normally <20. The lactate/pyruvate (L/P) ratio is increased in mitochondrial disorders but not in pyruvate dehydrogenase or carboxylase deficiency.
In some disorders the CSF:plasma ratio is diagnostic while in others the level of a particular amino acid may be abnormal. For example, in glycine encephalopathy the CSF:plasma glycine ratio is elevated to 0.1 or more (normal <0.025).
In pyridoxal phosphate-responsive seizures (pyridox(am)ine phosphate oxidase deficiency) causing neonatal seizures, threonine may be elevated in CSF and glycine in plasma.
In serine biosynthesis disorders (in particular 3-phosphoglycerate dehydrogenase and phosphoserine amino transferase deficiency), CSF serine and glycine are reduced while fasting plasma serine may be reduced less obviously.
In creatine synthesis disorders (especially guanidinoacetate-methyl transferase deficiency), CSF creatine is reduced, but brain MRS to see the absent creatine peak is the best investigation.
Measurement of these metabolites may be of value in patients with
• hyperphenylalaninaemia and suspected disorders of biopterin metabolism
• movement disorders such as dopa-responsive-dystonia
• suspected pyridox(am)ine-5'-phosphate oxidase (PNPO) deficiency causing neonatal seizures responding to pyridoxal phosphate .
NB Pyridoxal-5 -phosphate itself may be conveniently measured in the CSF in the same specimen used for detection of biogenic amines.
Pterins should always be measured in the same specimen of CSF as biogenic monoamines, but their estimation is also important in suspected inflammatory disorders, whether infective or autoimmune.
Neopterin in particular is increased in virtually all inflammatory and immune-mediated disorders (including Aicardi-Goutieres syndrome). Neopterin, which has a short half-life, is a significantly more sensitive marker of CNS inflammation than CSF pleocytosis.
|CSF pterins and monoamines|
|Sepiapterin reductase||±1 ↓||↑||N||↑||↓||↓||N|
|Abbreviations: AADC = aromatic L-amino acid decarboxylase; AD = autosomal dominant; AR = autosomal recessive; BH2 = 7,8-dihydrobiopterin; BH4 = tetrahydrobiopterin; GTP-CH = guanosine triphosphate cyclohydrolase; 5HIAA = 5-hydroxyindole acetic acid; HVA = homovanillic acid; N = within reference range for age and laboratory; 30MD = 3-0-methyldopa|
Succinylpurines are increased in adenylosuccinate lyase deficiency in which there is early-life epileptic encephalopathy.
IFN-α has been mentioned under cytokines. IFN-α is increased in congenital rubella, in CNS human immunodeficiency virus (HIV) infection, in the acute phase of herpes simplex encephalitis (in the infectious but not in the parainfectious relapse of that condition) and in Aicardi-Goutieres syndrome in the first year of life. IFN-α is also increased in acute neurological systemic lupus erythematosus.
Blood sampling with all diagnostic lumbar punctures
A blood sample must always be taken at the same time as lumbar puncture (preferably immediately before) to allow proper interpretation of CSF findings.
Indications for CSF examination
Lumbar puncture is indicated in all cases of suspect meningitis or encephalitis, except when skin petechiae make meningococcal infection virtually certain, or if brain swelling, mass lesion, or obstructive hydrocephalus is thought likely (see below, Contraindications).
The remaining indications may be summarized as follows (with findings in parentheses).
• Acute polyneuropathy or Guillain-Barre syndrome (increased total protein without increased cells).
• Suspect poliomyelitis (increased cells - lymphocytes and/or polymorphs).
• Subacute polyneuropathy (increased total protein and increased immunoglobulin synthesis).
• Acute disseminated encephalomyelitis (increased total protein, increased immunoglobulin synthesis, mild or moderate increase in lymphocytes).
• Multiple sclerosis (OCBs).
• Benign intracranial hypertension, after MRI (increased pressure).
• Suspect regressive disorder:
- Subacute sclerosing panencephalitis and rubella panencephalitis (OCBs; paired blood and viral titres)
- Krabbe disease and metachromatic leukodystrophy (increased protein)
- Mitochondrial (increased lactate, protein)
- Aicardi-Goutieres syndrome (increased lymphocytes or neopterin and IFN-α).
• Early-onset epileptic seizures:
- Glycine encephalopathy (increased glycine and increased CSF/serum glycine ratio)
- GLUT1 deficiency (decreased glucose, decreased CSF/blood glucose ratio, normal lactate)
- Pyridox(am)ine oxidase deficiency (elevated threonine in blood and CSF, decreased CSF pyridoxal-5 -phosphate).
• Movement disorders - both hypo- and hyperkinetic forms have specific abnormalities, in particular:
- Dopa-responsive dystonia
- Tyrosine hydroxylase deficiency
- Aromatic acid decarboxylase deficiency.
Contraindications to lumbar punctures
The most important contraindication to lumbar puncture is the suspicion of an intracranial mass lesion, brain swelling, obstructive hydrocephalus, spinal cord mass or spinal cord swelling. Clinical judgement is necessary here because no single sign or constellation of signs is consistently present in any of these situations. In the acute situation, where the history is measured in hours or days and the symptoms suggest bacterial meningitis, a critical matter is to select those few children in whom immediate diagnostic lumbar puncture is not indicated. Failure to localize a painful stimulus applied to the head (using Glasgow Coma Scale methodology) is an absolute contraindication to lumbar puncture. Tonic non-epileptic seizures or deterioration in the Glasgow Coma Scale score are strong warning signs of impending brain herniation and indicate that lumbar puncture may be dangerous (or be falsely blamed for a catastrophic outcome). Treatment may be implemented, imaging performed and the need for a lumbar puncture reconsidered after several hours.
In the subacute situation, in which the history is of the order of 2-4 weeks, with features such as headache, vomiting, weight loss, behaviour change or intermittent fever, the diferential diagnosis includes posterior fossa tumour, missed tuberculous meningitis or brain abscess. Evidence suggesting a risk of cerebral herniation includes a larger than expected head size, a cracked-pot percussion note over the coronal sutures (McEwan's sign). papilloedema, loss of retinal venous pulsation, and an abnormal head posture, either tilted or held stiffly in an anxious-looking manner.
Biochemical investigations in CSF
|Albumin||Routine||↓ in blood-brain or blood-CSF barrier, disruption and inflammatory polyneuropathy. May not be ↑ in mitochondrial Guillain-Barre syndromelike neuropathy. May be ↑ in central demyelination|
|Amino acids||Neonatal hiccups in ill baby (hiccups common in normal newborn infants)||Obtain paired samples of plasma and CSF||↑ glycine and ↑ CSF/plasma glycine ratio in glycine encephalopathy|
|Neonatal seizures/ encephalopathy/stiff baby/ microcephaly||Obtain paired samples of plasma and CSF||↓ serine and glycine in serine synthesis disorders|
|Suspect mitochondrial disorder||Obtain paired samples of plasma and CSF||Alanine and proline ± ↓ in mitochondrial disorders|
|Headache, vomiting||Obtain paired samples of plasma and CSF||Glutamine ↑ in OTC deficiency|
|Asialotransferrin (% of total transferrin) (= π or tau protein)||Pleomorphic central white matter disorder with onset from neonatal to late adolescence. Ataxia (often progressive spastic ataxia) ± dementia ± optic atrophy ± peripheral neuropathy ± macrocephaly ± coma. Deterioration with febrile illness or after head bump or fright||Test not yet validated in neonates and first year of life. Blood transferrin should be normal||↓ (<8%) in elF2B-related disorder (includes vanishing white matter but not exclusively so)|
|Biogenic amines (monoamine metabolites) (always analyse both amines and pterins together)||Infantile hypertonia with oculogyric crises and autonomic features. Infantile immobility and bradykinesia. 'Cerebral palsy'/delay ± oculogyric crises (NB easily misdiagnosed as absence epileptic seizures) and normal brain MRI. Acquired movement disorder not typical of known conditions. Acquired gait disturbance when features atypical for Segawa disease (cognitive delay, parkinsonism, oculogyric crises) argue against immediate levodopa trial||Organize in advance with supraregional laboratory. Freeze samples immediately at bedside on dry ice antioxidant mixture. If blood admixed, spin at once and transfer clear C5F to new tubes before freezing. Do not give levodopa within 1 week before||Patterns of dopamine, 5-HIAA and HVA vary in different monoamine neurotransmitter disorders - see also Pterins below|
|Creatine||Global delay, expressive language impairment, early-onset 'generalized' epilepsy, movement disorder (mixed, may be dystonic/ballistic)||Needs to be evaluated by in vitro H-MRS for lack of creatine peak (frozen immediately -discuss with lab in advance)||↓ in GAMT and AGAT deficiencies and in creatine transporter defect [SLC6A8]|
|GABA||In very exceptional cases of hyper-or hypotonia||Organize with supra regional laboratory. Not likely to be a helpful investigation||↑ in GABA-transaminase deficiency and ↑ in succinic semialdehyde dehydrogenase deficiency. ↓ in some cases of hyperekplexia but not helpful in diagnosis|
|Glucose||Epileptic seizures, developmental delay, ataxia, movement disorder, evolving microcephaly||Always take blood for plasma glucose immediately before a lumbar puncture||Numerous inflammatory and other cellular causes of ↓ glucose. Hypoglycaemia. ↓ absolute level of CSF glucose and CSF/blood glucose ratio indicates GLUT1 deficiency|
|Guanidinoacetate||Severe learning disability, epilepsy, movement disorder (often ballistic)||Urine usually||↑ in creatine synthesis disorder, will need proton brain MRS|
|Lactate||Suspect mitochondrial disorder||Blood for lactate before lumbar puncture||↑ value suggests a mitochondrial disorder|
|5-MTHF||Decreasing head growth with agitation, delay, sleep disturbance in infancy followed by developmental arrest or regression with later dyskinesia, ataxia, pyramidal signs, epileptic seizures, and on brain MRI white matter signal change||May not be a primary disorder. Homocysteine will be increased||↓ in cerebral folate deficiency. ↓ in 5-MTHF reductase deficiency (a form of homocystinuria)|
|Myelin basic protein||Suspect acute demyelination||Sensitivity and specificity not established||↑ in acute demyelination in multiple sclerosis, ADEM, neuromyelitis/ neuromyelitis optica|
|Neopterin||Suspect 6TP-CH deficiency||Measure with other pterins and biogenic amines||↓ in GTP-CH deficiency|
|Suspect inflammatory CNS disorder, infective or autoimmune, incl. Aicardi-Goutieres syndrome||More sensitive than CSF pleocytosis; short half-life||↑ in inflammatory CNS disorders (including Aicardi-Goutieres syndrome)|
|Poliols||Unexplained leukodystrophy||Can also be done in urine||↑ in ribose-5-phosphate isomerase deficiency|
|Pterins (always analyse both amines and pterins together; see also Neopterin)||As for biogenic amines (always analyse both amines and pterins together)||As for biogenic amines: discuss with supraregional laboratory. Sepiapterin may need separate analysis (see below)||As for biogenic amines. ↓ neopterin (and others) in various monoamine disorders|
|Congenital infectionlike picture||↑ neopterin in inflammation when T-lymphocytes or macrophages involved; autoimmunity; Aicardi-Goutieres syndrome|
|Pyridoxal 5'-phosphate (PLP)||Intractable neonatal epileptic seizures. Possibly other neurological phenotypes||Do pyridoxal phosphate trial first (Chapter 2.16)||↓ in PNPO deficiency|
|Sepiapterin||Developmental delay with atypical features, especially bulbar symptoms, oculogyric crises, diurnal variation||May be missed on standard pterin study||Sepiapterin reductase deficiency|
|Succinylpurines||Learning disability, autism. As for urine succinylpurines||Freeze specimen for Bratton-Marshall test||↓ ADSL deficiency ↑ AlCA-ribosiduria|