Microscopic Examination of Urine

Microscopic Examination of Urinary Sediment

Introduction

  • The microscopic examination of urinary sediment is the third component of routine urinalysis following physical and chemical evaluations.
  • Purpose: Detect and identify insoluble materials in urine.
  • Contributors to urine constituents:
    • Blood, Kidney, Lower Genitourinary Tract, External Contamination
  • Components of urine sediment:
    • Red Blood Cells (RBCs)
    • White Blood Cells (WBCs)
    • Epithelial Cells
    • Casts
    • Bacteria
    • Yeast
    • Parasites
    • Mucus
    • Spermatozoa
    • Crystals
    • Artifacts
  • Clinical significance:
    • Some components may be normal unless in increased amounts, necessitating both identification and quantitation during examination.

Microscopic Analysis Procedural Variations

  • Variations affecting microscopic analysis include:
    • Methods for sediment preparation
    • Volume of sediment examined
    • Methods and equipment for visualization
    • Results reporting methods

Standardization and Cost-effectiveness

  • Development of protocols to enhance standardization and cost-effectiveness in microscopic urinalysis.

Macroscopic Screening

  • Many laboratories perform microscopic examination of urine sediment based only on specific criteria to enhance cost-effectiveness.
  • Key parameters for consideration typically include:
    • Color
    • Clarity
    • Presence of Blood, Protein, Nitrite, Leukocyte Esterase, Glucose
  • Laboratory-designated criteria can be integrated into automated instruments.
  • Variability in detection rates:
    • Studies indicate a significant difference in percentages of abnormal specimens undetected using standard parameters.
  • Patient Population Considerations:
    • Special populations include pregnant, pediatric, geriatric, diabetic, immunocompromised, and renal patients.
  • The Clinical and Laboratory Standards Institute (CLSI) advises performing microscopic examinations when:
    • Requested by a physician
    • Testing a laboratory-specified patient population
    • Abnormal physical or chemical results are obtained.

Specimen Preparation

  • Freshness/Preservation: Specimens should be examined fresh or adequately preserved, as formed elements can disintegrate, especially in dilute alkaline urine.
  • Refrigeration can cause precipitation of non-pathological crystals, obscuring other sediment elements.
  • To mitigate precipitation, warm specimens to 37°C prior to centrifugation.
  • Clean-Catch Midstream Specimen: Minimizes external contamination, similar to physical and chemical analyses, yet dilute random specimens may yield false negatives.
  • Mix Thoroughly: Ensure proper mixing before decantation into a centrifuge tube.

Technical Tips

  • Tip 7-1: Warm refrigerated specimens to 37°C before centrifuging to dissolve amorphous urate crystals.

Specimen Volume

  • Recommended centrifuge volume: 10 to 15 mL (often 12 mL as multiparameter reagent strips can easily be immersed).
  • If specimen volume is less, the actual volume used must be noted for result correction (e.g., the results of a 6 mL sample are multiplied by 2 to account for volume).

Centrifugation

  • Consistency in Method: Centrify at 5 minutes with a Relative Centrifugal Force (RCF) of 400 to achieve optimal sediment with minimal element damage.
  • Conversion of RPM to RCF is done using the formula:
    RCF=1.118imes10−5imesextradiusincmimesextRPM2RCF = 1.118 imes 10^{-5} imes ext{radius in cm} imes ext{RPM}^2
  • Routine calibration of centrifugation is essential and should avoid the use of braking mechanisms that disrupt the sediment.
  • Ensure all centrifugation is performed in capped tubes to prevent biohazardous aerosols.

Sediment Preparation

  • A uniform volume of urine and sediment should remain following decantation.
  • Common volumes after centrifugation range from 0.5 to 1.0 mL; volume centrifuged divided by sediment volume equals the concentration factor (24 and 12 represented in examples).
  • Tip 7-2: Use commercial systems providing constant volumes for suspension and sediment resuspension.
  • Resuspension Technique: Use gentle agitation methods to ensure equal distribution during microscopic examination without damaging cellular elements.
  • Volume Examined: Recommended slide volume is 20 μL (0.02 mL) covered with a 22 x 22 mm cover slip.
    • Use of commercial systems controls the sediment volume for consistency.

Commercial Systems

  • Systems have improved the conventional method of glass slide preparation.
  • Some current systems include:
    • KOVA
    • Urisystem
    • Count-10 System
    • Quick-Prep Urinalysis System
    • CenSlide 2000 Urinalysis System
    • RS Urine Sediment Workstation
  • Features include:
    • Capped, calibrated centrifuge tubes
    • Decanting pipettes for precise volume control
    • Slides ensuring a monolayer of sediment, promoting consistent quantitation
    • Closed systems like Cen-Slide and RS minimizing specimen exposure

Examining the Sediment

  • Examining Methodology: Perform consistent examinations across a minimum of 10 fields under both low (10x) and high (40x) power magnifications.
  • The initial examination under low power is for casting detection and general composition assessment.
  • Use high power for definitive identification of important elements.
  • Tip 7-3: Use reduced light for visual accuracy during examinations with bright-field microscopy.

Reporting the Microscopic Examination

  • Reporting practices vary among labs but should maintain internal consistency.
  • Common reporting methods are:
    • Casts: average number per low-power field (lpf)
    • RBCs and WBCs: average number per high-power field (hpf)
    • Semiquantitative reporting of elements as “rare,” “few,” “moderate,” “many,” or graded as 1+, 2+, 3+, and 4+.
  • Laboratories must define reference values based on sediment concentration factor in use, which may vary (e.g., Urisystem concentration factor of 30 gives reference of 0 to 8 WBCs/hpf).