CHAPTER 14 Cell counts on CSF and other body fluids - PowerPoint Presentation

1. CHAPTER 14Cell counts on CSF and other body fluidsBy Bamlaku Enawgaw

2. ObjectivesAt the end of this chapter, students will be able to:Define terms related to body fluidsIdentify different types of body fluidsDefine CSFDefine Serous fluids: pleural, pericardial, peritoneal (Ascitic), and Synovial fluidExplain the analysis of CSFPerform cell counts on different body fluids Undertake QC measures in body fluid examination

3. What are body fluids? What is there difference from blood?Are extra vascular fluids (body cavity fluids) fluids contained with in the closed cavities of the body serve as lubricants as membranes move against each other314.1. Introduction to Body Fluids

4. Commonly analyzed Body fluids include:Cerebrospinal fluid (CSF) ………Brain & spinal cordSerous fluidPeritoneal/Ascetic fluid……Abdominal & pelvic cavitiesPericardial fluid…………….Around heartPleural fluid…………………Around lungsSynovial fluid …………….….fluid from the jointsHydrocele fluid (semen) ………………Tests cavities14.1. Introduction to Body Fluids

5. Examination of body fluidsGross examination - Turbidity - Color - Clot formationWet mount – Fungus, parasitesCell count…… total and diff WBC, RBC Culture , gram stain and acid fast stainProtein and glucose determination 5

6. 14.2. Cerebrospinal fluid (CSF)Clear sterile extravascular fluid circulates in the brain, sub-arachnoid space & spinal cordCSF is not an ultra filtrate of plasmaSimilar to plasmaless protein, less glucose and more chloride ionVolume: Adult 90 -150 ml Newborn 10 – 60ml450-750 mL per day or 20 ml per hour CSF is formed

7. Function of CSFCerebrospinal fluid serves to protect the underlying tissue of the central nervous system.prevent traumaRegulate volume of intracranial pressurecirculate nutrientsremove metabolic waste products from the CNS Act as lubricant

8. Location of CSFFig. Collecting a CSF specimenTube 1 = Chemical and immunologic testsTube 2 = Microbiology (culture and gram stain)Tube 3 = Hematology (gross examn, total WBC & Diff)1-2ml of CSF is collected by lumbar puncture/spinal tapBetween 3rd and 4th or 4th and 5th lumbar vertebraeCollected with three sterile tubesCSF collection

9. CSF Cont’dii. Clinical SignificanceDiagnosis of meningitis ….bacteria, viral, fungal, parasitesHemorrhage….sub-arachnoids or intracerebral Malignancy: leukemia…lymphomaiii. Principle of CSF analysisVisually and microscopicallycounted within 1 hour of collection. If delay is unavoidable store at 2-8oC.1v. Equipment and Reagents: same as for WBC counting on whole blood

10. Examination methods Gross appearanceIs visual assessment of CSF for turbidity, color and viscosityNormal CSF = crystal clear in appearance with viscosity comparable to waterAbnormal CSF may appear cloudy, smoky, hazy, opalescent, turbid or grossly bloody

11. Turbidity may be graded from 0 to 4+ as follows: 0 = crystal clear fluid1+ = faintly cloudy, smoky or hazy2+ = turbidity present but news print easily read3+ = news print not easily read through tube4+ = grossly turbid, news print cannot be seen

12. Cont’dBloody specimensTraumatic tap = 1st tube is bloody and last tube is clearHemorrhage = all tubes are same bloody color

13. Gross appearance cont’d2. ColorNormal CSF = crystal clearXanthochromia: is yellow coloration of CSFResult of release of hemoglobin from lysed red blood cells increase in bile pigments3. ViscosityNormal CSF = waterClotting may occur from increased fibrinogen resulting from a traumatic tap meningitis or subarachnoid block

14. Microscopic cellular enumerationCell count = RBC and WBCNR: 0-5 WBC/µL or 0-5 x 106/LNeonates have higher value of 0-30 cells/µLLow WBC with turbidity = high concentration of bacteriaWBC between 100-10,000 x 106/L = bacterial meningitisWBC >50,000 x 106/L = intraventricular rupture of brain abscess

15. Total Leucocyte Count on CSFIf CSF is clearMix well the undilutedcount the cells in 9mm2 area Multiply the number by 10/9If CSF is slightly turbidprepare a 1:10 dilutioncount the cells in 9mm2Multiply counted cells by 100/9 If CSF is purulent, similar to bloodHow the volume correction factor is obtained ?

16. Technique for Counting Mixture of WBC and RBCTo find the true WBC count when the CSF is bloodyPerform the WBC and RBC counts on the patient’s blood and CSF.RBCCSF x WBCblood= WBCADDED RBCBLOOD  Corrected WBCCSF = WBCCSF – WBCADDED

17. Excercise  Example:RBCBLOOD = 5 x 106/mm3RBCCSF = 2,500/mm3WBCBLOOD = 12 x 103/mm3WBCCSF = 70/mm3 WBCADDED = 2,500 x 12,000 5 x 106 = 6/mm3 Corrected WBCCSF = 70 – 6 = 64/mm3

18. Differential Leukocyte Count on CSFCentrifuge the CSF at 2500 rpm for 10 min (if it is clear)Remove supernatant Re-suspend the sediment and prepare a smearStain using Wright stain Wash off stain with water and air-dryIdentify the types of leucocytes (PMNs or mononuclear cells)Count at least 100 cells using the oil immersion objective

19. Differential Leukocyte Count on CSFCells in the spinal fluid may include: granulocytes mature or reactive lymphocytes, mononuclear phagocytes, plasma cells, blast cells and malignant cellsOthers like nucleated red cells, and intracellular bacteria.Lupus erythematosus (LE) cells are rarely foundOther rare cells unique to spinal fluid (ependymal cells, choroidal cells) may be found

20. Vii. Interpretation of CSF diff countNormal range <5 cells/mm3Increased PMN indicate bacterial infectionIncreased Eosinophils indicate: systemic parasitic and fungal infection, drug reactionIncreased basophils indicatechronic leukemia, Chronic granulocytic leukemiaParasitic infections

21. Interpretation of CSF diff cont’dIncreased lymphocytes indicate: viral infections, Fungal meningitisSyphilitic meningeal encephalitisIncreased monocytes (>2%) indicate:Tuberculosis meningitisSyphilis and viral encephalitisSubarachnoid hemorrhageLeukemiaIncreased macrophagesInfectious diseasesCNS leukemia, Lymphoma, Malignant myeloma,

22. Interpretation of CSF diff cont’dPlasma cells increased in:Viral disorders such as herpes simplex infectionMeningeal encephalitisSyphilitic involvement of the CNSHodgkins diseasePost subarachnoid hemorrhageErythrocytes increased in:Traumatic tap specimensPatients with a bleeding subarachnoid hemorrhage or intracerebral hemorrhageChronic myelogenous leukemia or erythroleukoblastic conditions

23. Associated findingsDecreased glucose in CSF with normal blood glucose Bacterial infection Elevated protein = inflammatory reaction or bacterial infectionViral infections have no dramatic effect on CSF glucose or protein levels

24. Count both sides of counting chamber (18 mm2 area)Increasing the number of cells to be counted for differential count (instead of 100 cells count 200 cells)Check staining quality (e.g. staining time, pH, filtering)Proper centrifugation (Speed and time) Properly follow SOP viii. Quality control

25. ix. Sources of errordilution, charging, counting , calculatingDelay in analysisCentrifugation time and speed preparation for Diff countStaining timeImproper handling of sample

26. Pleural fluid, pericardial fluid, peritoneal/ascitic fluid, synovial fluid, semenSame protocol followed as with CSFBut for effusions using anti-coagulant (9ml: 1ml)A high count of lymphocytes in pleural and peritoneal fluids is suggestive of tuberculosis 2614.3. Other body fluids

27. Definition of terms Effusion: an increase in volume of any serous fluidTransudates: effusion as a result of systemic diseases like liver cirrhosis, congestive heart diseaseExudates: are effusions resulting from inflammatory responses that directly affect the serous cavity (includes infections and malignancies).

28. Characteristics of Serous Effusions: Transudate versus ExudateObservation / TestTransudateExudateAppearanceWatery, clear, pale yellow, does not clotCloudy, turbid, purulent, or bloody; may clot (fibrinogen present)WBC countLow, < 1,000/µL with > 50% mononuclear cells500-1,000 cells/µL or more, with increased PMNs, increased lymphocytes with TB or rheumatoid arthritisRed cell countLow, unless from a traumatic tap> 100,000/µL, especially with a malignancyTotal proteinLow>3g/dl (or > than half the serum level)Lactate dehydrogenaseVaries with serum levelIncreased (>60% of serum level because of cellular debris)GlucoseNot applicableLower than serum level with some infections and high cell counts

29. Review Questions What is the function of body fluids in general?What is the role of CSF in our body?Mention at least three different types of body fluidsWhat is the difference between transudates and exudatesDefine serous fluidsDefine CSFHow do you perform the total leukocyte count on a: a) clear CSF b) slightly turbid CSF c) purulent CSF How do you correct the total leukocyte count to a true value when the count is performed on a sample of CSF that is slightly turbid due to traumatic tap? How is the differential leukocyte count performed on a sample of CSF?

30. Thanks