CSF PDF

CEREBROSPINAL FLUID CSF is produced in the choroid plexuses of the two lumbar ventricles and the third and fourth ventricles. Blood–brain barrier is the tight-fitting structure of the endothelial cells in the choroid plexuses that prevent the passage of many molecules. Meninges- it lines the br...

CEREBROSPINAL FLUID CSF is produced in the choroid plexuses of the two lumbar ventricles and the third and fourth ventricles. Blood–brain barrier is the tight-fitting structure of the endothelial cells in the choroid plexuses that prevent the passage of many molecules. Meninges- it lines the brain and spinal cord. Composed of three layers: A. Duramater (outer layer) – lines the skull and vertebral canal B. Arachnoid mater (Spiderweb-like) – filamentous inner membrane Subarachnoid space – where CSF flows C. Pia mater (innermost layer) – lines the surface of the brain and spinal cord  First recognized by Cotugno in 1764 3rd major body fluid Approximately 20 mL is produced every hour in the choroid plexuses and reabsorbed by the arachnoid villi Total volume: Adults: 90 to 150 mL Neonate: 10 to 60 mL Functions: Supply the nutrients to the nervous system Remove metabolic waste Produce a mechanical barrier to cushion the brain and spinal cord against trauma  SPECIMEN COLLECTION Method of collection: Lumbar puncture between third, fourth, or fifth lumbar vertebra CSF ORDER OF DRAW PURPOSE PRESERVATION TUBE 1 Chemical and Serologic tests Frozen TUBE 2 Microbiology Room Temperature TUBE 3 Hematology Refrigerated TUBE 4 Microbiology/ Serology If only 1 CSF tube is available: Microbiology → HEMATOLOGY → CHEMICAL AND SEROLOGIC TESTS A 4th tube may be drawn for microbiology to better exclude skin contamination or additional serologic tests GROSS EXAMINATION APPEARANCE SIGNIFICANCE Crystal clear NORMAL Hazy, Turbid, WBC counts over 200/uL Cloudy, Milky RBC counts over 400/uL ↑ Microorganisms ↑ Proteins Oily Radiographic contrast media Clotted, Protein, clotting factors Pellicle  APPEARANCE SIGNIFICANCE Xanthochromic Supernatant is pink, orange or yellow ↑ Serum bilirubin ↑ Protein concentration Presence of pigments: - Carotene - Melanoma Presence of RBC degradation products: - Pink: Very slight amount of oxyhgb - Yellow: Conversion of oxyhgb to unconjugated bilirubin - Orange: Heavy hemolysis Grossly bloody - RBC counts >6, 000/uL  Tubes of CSF. Appearance left to right is normal, xanthochromic, hemolyzed, and cloudy.  TRAUMATIC TAP VS. INTRACRANIAL HEMORRHAGE Distribution of Uneven Even (1=2=3) Blood (1>2>3) Clot Formation + - Supernatant Clear Xanthocromic Erythrophages - + D-dimer - + INTRACRANIAL HEMORRHAGE TRAUMATIC TAP MICROSCOPIC PARAMETERS  CSF CELL COUNT 1. Total Cell Count 2. WBC Count 3. RBC Count 4. DIFFERENTIAL COUNT Any cell count should be performed immediately because WBCs (particularly granulocytes) and RBCs begin to lyse within 1 hour, and 40% of the leukocytes disintegrate after 2 hrs Specimens that cannot be analyzed immediately should be refrigerated  1. TOTAL CELL COUNT Dilution: NSS (0.85% NaCl) An improved Neubauer counting chamber is routinely used for performing CSF cell counts. Fuchs-Rosenthal- type chamber may be used Cells are counted in the four corner squares and the center square on both sides of the hemocytometer Counting and calculation of the number of uL are done using the same procedure as WBC count Neubauer Counting Chamber  2. WBC COUNT Procedure: 1. Dilute (3% Acetic acid, NSS) 2. Charge Both side of Hemocytometer 3. Count in 5 large squares per side 4. Calculation Normal Values: Adult: 0 to5 WBCs/uL Neonates: as many as 30 mononuclear cells/uL  CSF DILUTION CLARITY DILUTION Clear May be counted undiluted Slightly Hazy 1:10 Hazy 1:20 Slightly cloudy 1:100 Cloudy 1:200 Slightly bloody Bloody, Turbid 1: 10,000  3. RBC COUNT Done in cases of Traumatic Tap Calculation: RBC count = Total cell count – WBC count Corrections for contamination: 1 WBC for every 700 RBCs 8 mg/dL protein for every 10, 000 RBCs/uL  4. DIFFERENTIAL COUNT Performed on a stained smear Specimen should be concentrated prior to preparation of smear Concentration techniques: Centrifugation for 5 to 10 mins Cytocentrifugation Cytocentrifugation- adds albumin to increase the cell yield and decrease cellular distortion Increased in number of normal cells in CSF: Pleocytosis (Abnormal condition) Normal cells found in CSF: Lymphocytes and monocytes Adult: Predominance of lymphocytes (70:30 ratio) Children: Predominance of monocytes (30:70 ratio) CHEMISTRY TESTS  1. TOTAL PROTEIN Normal values: 15 to 45 mg/dL Major CSF protein: Albumin 2nd most abundant: Prealbumin (Transthyretin) Alpha globulins: Haptoglobin, ceruloplasmin Beta-globulins: Beta2-transferrin (TAU) protein Gamma globulins: IgG and some IgA  Most frequently performed chemical test CSF can be identified based on the appearance of the extra isoform of tau transferrin that is found only in CSF. NOT found in normal csf: 1. IgM 2. Fibrinogen 3. Beta-lipoprotein  CSF PROTEIN DETERMINATION Turbidimetric 1. 3% Trichloroacetic acid (TCA) ✔ Preferred method; precipitates both albumins and globulins 2. SSA (3%) Total ✔ Precipitates albumin only; to precipitate globulins, add Protein sodium sulfate (Na2SO4) Dye-Binding Coomasie Brilliant Blue ✔ Color change from red to blue occurs when protein binds to dye ✔ ↑Protein = ↑ Blue color CSF/ Serum Abnormal: >9 9-14 = Slight impairment Albumin Index 15-30 = moderate impairment Protein >30= Severe impairment Fractions 100= complete damage to BBB IgG index NV =0.77  CSF ELECTROPHORESIS Done in conjunction with serum electrophoresis For the detection of oligoclonal bands in the gamma region The presence of 2 or more oligoclonal bands in CSF but not in serum is valuable diagnosis of Multiple sclerosis Other conditions with oligoclonal banding in CSF but not in serum are: Neurosyphilis, encephalitis, neoplastic disorders, Guillain barre syndrome  MYELIN BASIC PROTEIN Protein component of the lipid-protein complex that insulate the nerve fibers Presence of MBP in CSF indicate destruction of myelin sheath Used to monitor the course of MS  2. GLUCOSE Normal values: 60-70% of the plasma glucose. - If the plasma glucose is 100 mg/dL, then a reference CSF glucose would be approximately 65 mg/dL ↑ in Cases of: Result of plasma elevations ↓ in cases of: Bacterial, tubercular and fungal meningitis Normal in cases of: Viral meningitis  3. LACTATE Valuable aid in diagnosing and managing meningitis cases Normal values: 25 mg/dL Other tests: + Gram stain Agents: Agent: M. Agent: C. + Culture Enteroviruses tuberculosis neoformans + Limulus ✔ Poliovirus + AFB stain + Gram stain = lysate ✔ Echovirus + Pellicle classic ✔ Coxsackievir web-like clot starburst us formation after pattern 12-24 hours + India ink = refrigeration capsule (unstained); background (black)  Microbiology Tests This can take anywhere from 24 hours in cases of bacterial meningitis to 6 weeks for tubercular meningitis. Gram stain -The CSF should be centrifuged at 1500 g for 15 minutes, and slides and cultures should be prepared from the sediment. Organisms encountered most frequently include Streptococcus pneumoniae, Haemophilus influenzae , Escherichia coli , and Neisseria meningitidis. Streptococcus agalactiae and the gram-positive rods Listeria monocytogenes may be encountered in newborns. Fungal meningitis - Cryptococcus neoformans detect using India ink preparation. - Gram stain for the classic starburst pattern produced by Cryptococcus  LIMULUS LYSATE Detects Gram negative bacterial endotoxin in body fluids and surgical instruments Regent: Blood of Horseshoe crab Principle: in the presence of endotoxin, the amoebocytes (WBCs) will release lysate (protein), + clumping/ Clot formation  SEROLOGY ELISA Latex agglutination - detect the presence of C. neoformans antigen in serum and CSF provide a more sensitive method than the India ink preparation VDRL (Venereal disease research laboratory test) - recommended by the CDC to diagnose neurosyphilis FTA- ABS (fluorescent treponemal antibody absorption) - remains positive in the serum of treated cases of syphilis. Rheumatoid Factor - most common cause of false-positive reactions