MICROSCOPIC

EXAMINATION OF
URINE

By:
PROFESSOR JOMEL B. VASQUEZ
● Identification of insoluble substances
(formed elements)
● Red blood cells (RBCs)
● White blood cells (WBCs)
● Epithelial cells
● Casts
● Bacteria
● Yeast parasites
● Mucus
● Spermatozoa
● Crystals
● Artifacts
● Preparation of sediment
● Volume of sediment examined
● Methods of visualization
● Reporting of results
● Commercial systems: KOVA
● Calibrated centrifuge tubes,
special slides to control
volume, decanting pipettes,
grids for better quantitation
Macroscopic Screening
Correlations
Screening Test Significance
● Color Blood
● Clarity Hematuria versus
hemoglobinuria/myoglobinuria
● Confirm pathologic or
nonpathologic cause of turbidity
● Blood RBCs/RBC casts
● Protein Casts/cells
● Nitrite Bacteria/WBCs
● Leukocyte esterase WBCs/WBC casts/bacteria
● Glucose Yeast
Specimen Preparation
➢ Examine when fresh or preserved
➢ RBCs, WBCs, casts lysed in dilute,
alkaline urine
➢ Refrigeration precipitates crystals
➢ Less contamination (epithelial cells)
from a midstream clean-catch specimen
➢ Mix specimen before decanting to the
centrifuge tube
Specimen Volume
● Centrifuge 10–15 mL urine (reagent
strips fit into 12 mL)
● Always cap tubes
● Too little volume = fewer formed
elements
● Some laboratories correct for
volume
Centrifugation
● Standardize speed and time of
centrifugation
● 5 min. at relative centrifugal force
(RCF) of 400 is ideal
● RCF corrects for variations in the
diameter of centrifuge heads;
revolutions per minute does not
● Do not brake the centrifuge
Postcentrifuge Sediment
➢ 0.5–1.0 mL after decantation
➢ Concentration factor: volume of
urine centrifuged/sediment volume
➢ Probability of detecting low
quantities of formed elements
➢ Aspirate rather than pour off urine
(pipettes available for this)
➢ Mix sediment gently, not vigorously
Volume of Sediment Examined

➢ Commercial systems control this

➢ Glass slide method:
➢ 20 μL or 0.02 mL
➢ 22 x 22 glass cover slip
➢ Do not overflow cover slip
➢ Heavier elements (casts) flow
outside
Examination of
Sediment
➢ Minimum 10 lpfs and 10 hpfs
➢ Low power: casts, general composition
➢ Scan edges for casts with glass slide method
➢ High power: identification
➢ Initial focusing: low power, reduced light
➢ Focus on epithelial cell, not artifacts that
are in a different plane
➢ Use fine adjustment continuously for best
view
Microscopic Reporting
➢ Consistent within laboratory
➢ Rare, few, moderate, many, or full
field or 1+, etc. – semiquantitative
➢ Casts: average per lpf
➢ RBCs, WBCs: average per hpf
➢ Epithelial cells, crystals, etc., in
semiquantitative terms
Sediment Stains
➢ Low refractive index elements are often
difficult to see under bright-field microscopy
➢ Supravital stain: crystal violet and Safranin O
➢ Most common
➢ Increases refractive index
➢ Stains nuclei, cytoplasm, inclusions
➢ Sedi-Stain, KOVA stain, etc.
➢ Acetic acid will enhance WBC nuclei
 Sediment Stains
➢ Lipid stains
➢ Oil Red O and Sudan III for triglycerides and
neutral fats; cholesterol polarizes
➢ Gram stain
➢ Identification of bacterial casts
➢ Hansel stain
➢ Urinary eosinophils
➢ Methylene blue and eosin Y: better than Wright
stain
➢ Prussian blue stain
➢ Hemosiderin granules seen with hemoglobinuria
Microscopy
➢ Bright field most common in urinalysis
➢ Reduced light is essential
➢ Magnification is 10x and 40x
➢ Par focal means minimal adjustment
when changing objectives (use fine
adjustment)
➢ Lower light using the rheostat
➢ Condenser can be raised up and down
➢ Do not use the aperture diaphragm
Care of the Microscope
1. Carry microscope with two hands, supporting
the base with one hand.
2. Always hold the microscope in a vertical
position.
3. Only clean optical surfaces with a good quality
lens tissue and commercial lens cleaner.
4. Do not use the 10x and 40xobjectives with oil.
5. Clean the oil immersion lens after use.
6. Always remove slides with the low-power
objective raised.
7. Store the microscope with the low-power
objective in position and the stage centered.
Microscopy
● Phase-contrast microscopy
● Increases refractive index

● Polarizing microscopy
● Crystals and lipids
● Ability to split light into two beams
● Crystals are multicolored
● Cholesterol produces Maltese cross formations

● Interference-contrast microscopy
● Three-dimensional images
SEDIMENTS CONSTITUENTS
● RBC
Yeast cells
Oil droplets Air bubbles
Dysmorphic RBC
Glomerular bleeding
Strenous Exercise
Clinical Significance
●NV = 0-3 or 5/hpf
●Glomerular damage
●Vascular injury
WHITE BLOOD CELLS
EOSINOPHILS
3 TYPES OF EPITHELIAL CELLS
●Squamous epithelial cells
●Transitional epithelial
cells
●Renal tubular epithelial
cells
EPITHELIAL CELLS
SQUAMOUS EPITHELIAL CELLS
SPHERICAL TRANSITIONAL
EPITHELIAL CELLS
Caudate transitional epithelial
cells
Columnar PCT cells with granules
and fat globules
Oval DCT cells
Collecting duct RTE cells
Clinical Significance
● RTE cells
● Necrosis of renal tubules
exposure to heavy metals
drug induced toxicity
hemoglobin & myoglobin toxicity
viral infections
pyelonephritis, allergic reactions
malignancy, salicylate poisoning
acute allogenic transplant
rejection
Prussian blue stained hemosiderin
granules
OVAL FAT BODIES
Oval Fat Bodies
LIPIDURIA
●Severe tubular
necrosis
●Diabetes mellitus
●Trauma
BACTERIA
YEAST CELLS
PARASITES
SPERMATOZOA
MUCUS CELLS
CASTS
●Found in the kidney
●Formed within the
lumens of the DCT and
collecting ducts
●Lpf is used
Cast Composition and
Formation
●Tamm Horsfall Protein
● Increased – stress and
exercise
● Hyaline casts
● Formation:
● Aggregated TH fibrils attached to
RTEs
● Interweaving of protein fibrils which
form a loose network
● More interweaving to form solid
matrix
● Attachment of elements to matrix
● Detachment of fibrils from RTEs
● Excretion of cast
●Cylindroids
tapered end structures which
are formed at the junction of
ascending loop of henle and DCT
HYALINE CASTS
Hyaline cast under phase contrast
microscopy
RBC CASTS
Granular dirty cast
WBC CASTS
Disintegrating WBC cast
BACTERIAL CAST
EPITHELIAL CELL CAST
●Advanced tubular
destruction
●Urinary stasis
●Viral infections & renal
transplant rejection
●Tamm-Horsfall protein
RTE cells
RTE cast with bilirubin stained
cells
FATTY CASTS
Clinical significance
● Lipiduria
● Nephrotic syndrome
● Toxic tubular necrosis
● Diabetes mellitus
● Crush injuries
GRANULAR
CASTS
WAXY CASTS
BROAD CASTS
URINARY CRYSTALS
●Liver diseases
●Inborn error of
metabolism
●Renal damage
Crystal Formation
●Formed by the
precipitation of urine
solutes
●temperature
●pH
Normal crystals seen in acidic
urine

●Amorphous urates
●Uric acid
●Acid urates
●Sodium urates
AMORPHOUS URATES
URIC ACID CRYSTALS
ACID URATES AND SODIUM
URATES
● Acid urates are similar to
ammonium biurates
● Sodium urates are needle shaped
crystals
CALCIUM OXALATE
CRYSTALS
Normal crystals in alkaline
urine
●Amorphous phosphates
●Triple phosphates
●Calcium phosphates
● Calcium carbonate
● Ammonium biurates
AMORPHOUS
PHOSPHATES
TRIPLE PHOSPHATES
CALCIUM CARBONATE
AMMONIUM BIURATES
ABNORMAL URINE CRYSTALS
● Found in acidic urine
● Cystine crystals
● Cholesterol crystals
● Radiographic dye crystals
● Crystal associated with liver
diseases
● Sulfonamide crystals
● Ampicillin crystals
CYSTINE CRYSTALS
CHOLESTEROL CRYSTALS
RADIOGRAPHIC DYE CRYSTALS
● Similar to cholesterol
crystals
● Specific gravity is markedly
elevated
LIVER DISEASE CRYSTALS
●Tyrosine crystals
●Leucine crystals
●Bilirubin crystals
TYROSINE CRYSTALS
LEUCINE CRYSTALS
BILIRUBIN CRYSTALS
SULFONAMIDE CRYSTALS
AMPICILLIN CRYSTALS
URINARY SEDIMENT ARTIFACTS
● Starch
● Oil droplets
● Air bubbles
● Pollen grains
● Fibers
● Fecal contamination
Fiber
●THANK
YOU.