Body fluids

Body Fluids

Overview of Body Fluids

  • Types of body fluids:

    • Cerebrospinal Fluid (CSF)

    • Synovial Fluid

    • Serous Fluids

Cell Counts on Body Fluids: Overview

  • Examination Methodology

    • Involves assessing gross appearance (color, turbidity, volume).

    • Requires red and white blood cell (RBC, WBC) counts, and WBC differential on cytocentrifuge slides.

    • Slides should be prepared as soon as possible (preferably within 1 hour) to prevent deterioration of WBCs; deterioration can start within 30 minutes.

    • Samples must be gently mixed before each testing step.

  • Methods of Counting

    • Counts are typically performed using a hemocytometer.

    • Automated analysis is available with specific approved instruments for body fluid analysis.

    • Reports of counts are valid if within linearity for the analyzer, without cellular interference flags.

  • Sample Preparation and Dilutions

    • Cell counts are done on undiluted fluids when clear; dilutions may be needed if the sample is hazy, cloudy, turbidity, or bloody.

    • Hematological values:

    • No specific value for RBC count in serous fluids; clinical context comes from the sample's appearance (e.g., slightly bloody, bloody, grossly bloody).

    • Dilution Protocols

    • RBC dilutions use isotonic saline.

    • WBC dilutions utilize glacial acetic acid or a Türk solution (mix of glacial acetic acid and methylene blue).

    • Total dilutions should be minimized.

    • Acetic acid should never be used for synovial fluids due to hyaluronic acid coagulation.

  • Handling Synovial Fluids

    • Synovial fluids are typically viscous; hyaluronidase powder is added before counting to liquefy the samples.

    • Amount of dilution depends on turbidity and cell counts from undiluted samples.

  • Counting Procedure

    • WBC > 200/μL or RBC counts > 400/μL indicates slight hazy appearance.

    • If slightly bloody, perform undiluted RBC count, but for WBC, a 1:2 dilution is preferred (using Turk solution).

    • If the sample is bloody, then conduct a 1:200 RBC and either a 1:2 or 1:10 dilution for WBC.

    • Proper calibration of pipettes is crucial.

  • Hemocytometer Counting

    • Typically, all nine squares on both sides of the hemocytometer should be counted for body fluids; if counts are substantially high, fewer squares may be counted, and volume calculations adjusted accordingly.

Standard Operating Procedures (SOP) for CSF Cell Counting

  • Refer to specific SOP documentation for detailed protocols with hemocytometer usage.

Cytocentrifuge Slide Preparation

  • Cytocentrifuge is essential for slide preparation following cell count completions.

  • A relevant video tutorial can be viewed for instructional guidance.

Cerebrospinal Fluid (CSF) Examination

Anatomy and Collection

  • CNS Structure: Surrounded by the meningeal membrane which comprises three layers:

    • Dura Mater: Thick outer layer providing protection.

    • Arachnoid Mater: Middle protective layer.

    • Pia Mater: Innermost layer closely attached to the CNS tissues.

  • Subarachnoid Space: Between the arachnoid and pia mater, this space contains CSF—protecting and supporting the CNS, maintaining ionic environment by circulating nutrients and removing waste.

Blood-Brain Barrier

  • Composed of astrocytes that regulate substance transfer:

    • Permitted: Water, oxygen, glucose, essential amino acids.

    • Prevented: Larger molecules like proteins and drugs.

  • CSF Circulation: Flows through the ventricular system (brain chambers).

CSF Production

  • Specialized capillaries in pia mater (Choroid plexus) are the main source of CSF production, producing about 20 mL/hour.

  • Normal volume ranges include:

    • Adults: 90-150 mL

    • Children: 60-100 mL

    • Neonates: 10-60 mL

Sample Collection Guidelines

  • CSF must be tested within 1 hour of collection to avoid degradation causing falsely low WBC/RBC counts. Cellular membranes become destabilized faster at lower protein levels in CSF.

  • Samples should be collected in sterile, non-anticoagulated tubes, typically involving three or four tubes containing 2-4 mL each, collected in sequence.

Indications for CSF Collection

  • Suspected meningeal infection.

  • Subarachnoid hemorrhage.

  • Central nervous system malignancy.

  • Demyelinating diseases.

  • Collected between L3 and L4 spinal vertebrae under sterile conditions.

CSF Collection Tubes

  • Tube #1: For chemistry; likely to be contaminated by blood and debris—requires centrifugation for protein, glucose, lactate testing.

  • Tube #2: For microbiology; includes gram stains and cultures.

  • Tube #3: For hematology; assessing cell counts, differentials, and xanthochromia.

  • Tube #4: For additional chemistry, hematology, cytology, serology, flow cytometry, or molecular biology tests.

Sample Analysis and Handling

  • Post-analysis, samples should be refrigerated unless sent to microbiology (where pathogens may be impacted by low temperatures). For microbiology, samples are better incubated at 37°C until testing.

Blood Sample Coordination

  • Blood draws should accompany CSF analyses for complete blood counts (CBC) and chemistry testing.

Gross Examination of CSF

  • Normal CSF Characteristics: Non-viscous, clear, colorless.

  • Cloudiness: Indicates presence of microorganisms, increased WBC/RBC counts, and proteins.

  • Blood presence:

    • Traumatic tap: Blood in the first tube, progressively clearer in following tubes.

    • Pathologic hemorrhage: All tubes equally bloody, often due to subarachnoid hemorrhage.

CSF Characteristics

Traumatic Tap

Pathologic Hemorrhage

Supernatant Color

Clear

Colored or hemolyzed

Tube Appearance

Clears progressively

Same appearance in all tubes

Other Features

Bone marrow contamination; presence of cartilage cells

Erythrophages; may have siderophages and bilirubin crystals

Centrifuge Testing of Bloody CSF

  • Centrifuge and assess supernatant for color changes:

    • Clear/cytologically normal potentially indicates a traumatic tap.

    • Yellow/pinkish color suggests subarachnoid hemorrhage alongside potential yellow (xanthochromia).

Normal Cell Counts for CSF

  • Per established guidelines:

    • Adults: 0-5 WBC/µL; 0 RBCs.

    • Neonates: 0-30 WBC/µL; 0 RBCs.

  • Reports often focus on WBC only.

Quantification of Cell Counts

  • Counts are reported in $x imes 10^6/L$.

  • Analyzer may be needed if RBC counts are excessively high.

  • WBC count distinctions:

    • In the $1000s$: indicative of bacterial meningitis.

    • In the $100s$: indicative of viral meningitis.

    • Differential analysis of predominant cells suggests type of meningitis.

WBC Differential in CSF

  • Normal Constituents: Lymphocytes and monocytes present in small numbers; adults predominantly lymphocytes, while neonates show monocytes.

  • Increased Neutrophils: Search rigorously for bacteria, known early indicators in meningitis.

Other Cells in WBC Differential

  • Ependymal cells: Large cells with abundant cytoplasm appearing in clumps; not definitive for diagnosis and often confused with malignant cells.

  • Cartilage Cells: Large cells with red cytoplasm/signifies vertebral penetration.

  • Siderophages: Macrophages characterized by ingestion of RBCs, showing hemosiderin as dark granules, indication of hemorrhage occurring 48 hours prior.

  • Lymphoblasts: Can be observed in various counts, suggestive of pathological states or infections.

Serous Body Fluids

Overview of Serous Fluids

  • Types: Include but not limited to pleural, pericardial, and peritoneal fluids.

  • Functions: Lubrication between organs to reduce friction, protective cushioning.

Effusion Types

  • Definitions: Effusions indicate fluid accumulation in serous cavities, categorized as transudates or exudates.

    • Transudates: Result from systemic processes, generally due to increased hydrostatic pressure causing fluid leakage—linked with conditions like congestive heart failure, kidney, liver failures.

    • Characteristic Metrics:

      • Specific Gravity: < 1.016

      • Protein: < 3 g/dL

      • Lactate Dehydrogenase: < 200 IU

      • WBC Count: < 1000/µL (predominantly mononuclear cells)

    • Exudates: Result from inflammatory processes where both fluid and proteins are involved—associated with infections or malignancies.

    • Characteristic Metrics:

      • Specific Gravity: > 1.016

      • Protein: > 3 g/dL

      • Lactate Dehydrogenase: > 200 IU

      • WBC Count: > 1000/µL

Serous Fluid Appearance

  • Normal serous fluid should be pale yellow and transparent.

    • Transudates: Generally straw-colored and clear.

    • Exudates: Typically cloudy or hazy due to infectious processes.

    • Bloody: May signal trauma or malignancy.

    • Milky: Indicative of chyle presence, especially in pleural fluids.

Series of Differential Cell Types Found in Serous Fluids

  • Lymphocytes, Histiocytes (macrophages), Mesothelial cells: Normal cellular constituents, changes indicate disease.

  • Neutrophils: Presence suggests abnormal processes; hyper-segmented appearances may warrant bacterial scans.

  • Mesothelial Cells: Show varied morphology, notable in effusions, may appear frayed or in giant cell formations, common in sterile inflammatory contexts and less in infections.

  • Erythrophages and Siderophages: Indicate prior bleeding; erythrophages denote monocyte ingestion of RBCs, while siderophages show granules related to iron metabolism.

  • Neutrophils: Higher incidence demands searches for bacteria and responses to gram stains.

  • Lupus Erythematosus Cells: Can be indicative of Lupus, characterized by intact neutrophils that have engulfed a mass of nuclear material, necessitating reporting upon identification.

Synovial Fluid

  • Normal Characteristics: Straw-colored, clear, viscous fluid found in joint cavities, aiding lubrication.

  • Differential Cell Types: Includes lymphocytes, monocytes, histocytes, whereas LE cells can sporadically appear in specific disease states.

  • Presence of Crystals:

    • Cholesterol Crystals: Appearing large and flat, indicate chronic effusions like Rheumatoid arthritis.

    • Calcium Pyrophosphate Crystals: Small rhomboid, found in pseudo-gout; weakly birefringent.

    • Monosodium Urate Crystals: Large needle-like appearances associated with gout; they show strong birefringence dependent on the crystal’s orientation relative to polarized light.

Bronchoalveolar Lavage Specimen

  • Procedure Description: Introduces warm saline into the lungs and withdraws it, enabling identification of cellular organisms and contents inaccessible otherwise, especially during lung dysfunction.

  • Sample Handling: Samples ought to be mixed cautiously and examined within biosafety cabinets, as there are aerosolization risks.

    • Expected Cell Types: Includes neutrophils, macrophages, lymphocytes, but not mesothelial cells; ciliated epithelial cells must be reported as they suggest upper respiratory tract sample collection, rather than deep lung samples.

  • Carbonaceous Histocytes: Possible findings indicative of tobacco usage.

  • Monitoring for Infection: Pneumocystis jiroveci may indicate infection in HIV-positive patients.