Specimen (Urine) Volume: 10-15 ml, average of 12ml

INTRODUCTION • For the multi-parameter reagent strip testing

• The third part of urinalysis is the MICROSCOPIC
EXAMINATION of the urine sediment. Ideal container of urine prior to centrifugation –
• Purpose: detect and identify insoluble materials conical centrifuge tube
present in urine.
• The blood, the kidney, the lower urinary tract, and
external contamination all contribute formed
elements to the urine.

Microscopic analysis is subject to several procedural

variations including:
• methods by which the sediment is prepared
• volume of sediment actually examined
• methods and equipment used to obtain visualization
• manner in which the results are reported

MACROSCOPIC SCREENING
• Physical and chemical examination of the urine is
done first to assess for abnormalities.
• Plays primary role in the decision to perform
microscopic analysis
• Parameters considered significant vary among
laboratories but usually include color, clarity, blood,
protein, nitrite, leukocyte esterase, and possibly,
glucose.

SPECIMEN COLLECTION
• The urine must be collected in a clean, sterile
container free from fibers, talc and other contaminants.
• All urine collections must avoid contamination with
menstrual blood, vaginal secretions and feces.
• The patient will be asked to collect another sample if COMMERCIAL SYSTEMS
any contamination is found. • Currently available systems:
▪ KOVA (Hycor Biomedical, Inc., Garden Grove, CA)
SPECIMEN PREPARATION ▪ Urisystem (ThermoFisher Scientific, Waltham, MA)
• Fresh or adequately-preserved specimen ▪ Count-10 (V-Tech, Inc., Pomona, CA)
• Formed elements (RBCs, WBCs, hyaline casts) ▪ Quick-Prep Urinalysis System (Globe Scientific,
disintegrate rapidly in dilute, alkaline urine Paramus, NJ)
• Refrigeration may cause precipitation of amorphous ▪ CenSlide 2000 Urinalysis System (International
materials Remote Imaging Systems, Norwood, MA)
• Warming at 37 C prior to centrifuging may dissolve ▪ R/S Workstations 1000, 2000, 2003 (DioSys,
some crystals Waterbury, CA)
• MSCC minimizes external contamination
• Dilute, random specimens may cause FALSE- • Cen-Slide and R/S Workstations do not require
NEGATIVE readings. manual loading of the centrifuged specimen onto a
• Care must be taken to thoroughly mix the specimen slide; considered closed systems that minimize
prior to decanting a portion into a centrifuge tube. exposure to the specimen.

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• Cen-Slide provides a specially designed tube that
permits direct reading of the urine sediment.
• R/S Workstations consist of a glass flow cell into
which urine sediment is pumped, microscopically
examined, and then flushed from the system.

REPORTING THE MICROSCOPIC EXAMINATION

• The terminology and methods of reporting may differ
slightly among laboratories.
• Routinely, casts are reported as the average number
per low-power field (lpf)
• RBCs and WBCs as the average number per high-
power fields (hpf’s).
• Epithelial cells, crystals, and other elements are
frequently reported in semi-quantitative terms such
as rare, few, moderate, and many, or as 1+, 2+, 3+,
and 4+.

• Hyaline casts 0-2/lpf

• Coarse granular casts 1-2/lpf
• RBC 0-2/hpf
• Epithelial cells few
• Bacteria many

EXAMINATION OF THE SEDIMENT

• The sediment must first be observed using the low
power objective to observe for the presence of casts,
amorphous materials and squamous epithelial cells.
• After (LPO): shift to HPO then observe and quantitate
cells and crystals.
• If one needs to confirm the identity of the cast,
HPO is commonly used.
• Minimum of 10 fields per objective

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 SEDIMENT EXAMINATION TECHNIQUES

SEDIMENT STAINS
• STAINING
- increases overall visibility of sediment elements being
examined using bright-field microscopy by changing
the refractive index
- imparts identifying characteristics to cellular
structures, such as nuclei, cytoplasm, and inclusions

Sternheimer-Malbin stain
• consists of Crystal Violet and Safranin O
• absorbed well by WBCs, epithelial cells and casts,
providing clearer delineation of structure and
contrasting colors of the nucleus and cytoplasm

0.5% Toluidine blue

• A metachromatic stain
• Provides enhancement of nuclear detail
• Can be useful in the differentiation between WBCs &
renal tubular epithelial cells
• Used in the examination of cells from other body fluids

2% Acetic Acid
• Enhances nuclear detail
• Cannot be used for initial sediment analysis because
RBCs are lysed by the acetic acid

Lipid Stains
• Oil Red O and Sudan III
• Used to demonstrate the presence of free fat droplets,
lipid-containing cells and casts in the urinary sediment.
• Polarizing microscopy can also be used
• Triglycerides & neutral fat stain orange-red
• Cholesterol do not stain but is capable of
polarization INTERFERENCE-CONTRAST MICROSCOPY
• Two types:
Gram Stain 1. Modulation contrast (Hoffman)
• Used primarily in the microbiology section 2. Differential-interference contrast (Nomarski)
• For the differentiation between gram-positive (blue)
and gram-negative (red) bacteria
• Role in routine urinalysis: identification of bacterial
casts, which can easily be confused with granular
casts.
• A dried, heat-fixed preparation of the urine sediment
must be used.

Hansel Stain
Preferred stain for urinary eosinophils
• Clinical Significance: drug-induced allergic reaction
• Consists of methylene blue & eosin Y
• Wright’s stain can also be used.
• Staining is performed on a dried smear of the
centrifuged specimen or a cytocentrifuged
preparation of the sediment.

Prussian Blue Stain

Yellow-brown granules may be seen in renal tubular
epithelial cells and casts or free-floating in the urine
sediment after episodes of hemoglobinuria
• To confirm that these granules are hemosiderin, the
Prussian blue stain for iron is used and stains the
hemosiderin granules a blue color.
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 CELLS IN THE URINE SEDIMENT RBC’s are frequently confused with:
Monohydrate Calcium oxalate crystals, Air bubbles,
Red blood cells (RBC’s) Yeast cells, Oil droplets
▪ To resolve: Add acetic acid to a portion of the
• Appearance: Smooth, non-nucleated, biconcave disc
sediment, if the suspected cells are RBCs they will
approximately 7mm in diameter
be lysed and therefore will disappear from view
• In concentrated urine specimen (hypersthenuria),
leaving the yeast, air bubbles, CaOx and oil droplets
the cells shrink due to loss of water therefore the
intact.
appearance of RBCs are CRENATED.
▪ You can also use supravital stains.
• In dilute specimens (hyposthenuria), the cells
absorb water, swell, and then lyse rapidly releasing
their hemoglobin leaving only their cell membrane;
thus, the empty cells are termed as GHOST CELLS.

• Reporting of results: average number of RBC’s per

10 high power fields (e.g. 6-8/hpf)
• Normal: 0-2/ or 0-3/hpf
• Clinical significance: RENAL OR GENITOURINARY
ORGAN BLEEDING (HEMATURIA) primarily due to
glomerular membrane or vascular injury
- The number of red blood cells present is
indicative of the extent of the damage or injury
▪ DYSMORPHIC RBCs – RBC’s with irregular
cellular protrusions, with blebs and punch-out
centers are indicative of GLOMERULAR
BLEEDING
• Clinical Correlations:
- Urine Color
- Strip Reaction (Blood)

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 EOSINOPHILS
• Clinical Significance: Drug-Induced Interstitial
Nephritis, UTI and renal transplant rejection
WHITE BLOOD CELLS (WBC’s) • Hansel stain: preferred stain used for eosinophils;
Wright's can also be used
• Appearance: Larger than RBCs, 12mm, with visible • Percentage of eosinophils in 100 to 500 cells is
cytoplasmic granules determined
• Most predominant WBC is NEUTROPHIL containing • NOT normally seen in urine
granules and nuclei. • Finding of more than 1% eosinophils is considered
• When exposed to hypotonic urine, the cell absorbs significant.
water and swell & when subjected to Brownian motion,
they sparkle and their granules become more
prominent and they are termed as GLITTER CELLS.

MONONUCLEAR CELLS
▪ Lymphocytes, monocytes, macrophages, histiocytes
▪ Lymphocytes are the smallest WBCs; they may
resemble RBCs. They are seen in early cases of renal
transplant rejections.
▪ •Monocytes, macrophages, and histiocytes are large
cells and may appear vacuolated or contain inclusions.

• Sources of Error: Differentiate WBC from RTE

• Clinical significance: PYURIA (increased urinary
WBCs) indicating INFECTION OR INFLAMMATION
OF THE GENITOURINARY SYSTEM.
• Clinical correlations: LE, Nitrite
• Reporting of results: average number of WBCs per
10 high power fields (e.g., 10-15hpf)
• Normal: 0-5/hpf

• If you still want to confirm if the cell is White Blood Cell,

the med tech can add acetic acid to the sediment:
• Acetic Acid will make the nuclei of the WBCs more
visible.

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 1. SQUAMOUS EPITHELIAL CELL
• Appearance: LARGEST CELL in urine sediment
usually flattened with abundant, irregular cytoplasm
and prominent nuclei
• Reporting: rare, few moderate, many / low power field
(e.g. many/lpf)
• Originate from the lining of the vagina and female
urethra and the lower portion of the male urethra.

• The TOLUIDINE BLUE stain can also enhance the

nuclei of WBC to differentiate them from RTE.
• RTE cells are larger than WBCs and have
eccentrically (off-center) located nucleus.

EPITHELIAL CELLS
✓ derived from the linings of the genitourinary system
✓ represent normal sloughing of old cells
• Clinical significance: Represent normal sloughing and
• Squamous pose NO CLINICAL SIGNIFICANCE
• Transitional (Urothelial) • Most commonly seen in FEMALE: a frequent
• Renal Tubular Epithelial (RTE) Cell contamination in female specimen
• Remedy: Midstream Clean Catch collection

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 Variation of SEC • Increased numbers are seen singly, in pairs or in
• Significance: Squamous epithelial cell studded with groups (syncytia) and are indicative of invasive
coccobacillus (the bacteria almost cover the surface urologic procedures such as CATHETERIZATION but
of the cell and extend beyond the edges) of no clinical significance.
• CLUE CELLS for Gardnerella vaginalis; the
causative agent of Bacterial vaginosis

2. TRANSITIONAL/UROTHELIAL
• Spherical, polyhedral, or caudate with distinct,
CENTRALLY LOCATED NUCLEUS
• May resemble RTE (Eccentric)
• Originate from the renal pelvis, calyces, ureters
and bladder and upper portion of the urethra.

• Reporting: rare, few moderate, many /high power

field (e.g. many/hpf)
• NORMALLY PRESENT IN URINE BUT IN SMALL
AMOUNTS, ALSO REPRESENTS NORMAL
SLOUGHING.

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• Variation:
• Abnormal vacuoles and irregular nuclei are
indicative of malignancy and viral infection and must
be referred for cytological exam.

3. RENAL TUBULAR EPITHELIAL CELLS

• Cells of the renal tubules that appear Rectangular,
polyhedral, cuboidal, or columnar with an
ECCENTRIC (OFF-CENTER) NUCLEUS
• Coarsely granular cytoplasm
• Often resembles casts
• THE MOST CLINICALLY SIGNIFICANT OF ALL
EPITHELIAL CELLS

• RTE cells from PCT are fine cell that has absorbed fat
globules
• Cells from DCT are smaller than those from the PCT
and are round or oval
• Collecting duct RTE cells are cuboidal and are never
round
• Cells from CD that appear in groups of 3 or more are
called renal fragments. They are frequently seen as
large sheets of cells. PCT and DCT cells are not
seen in large sheets of cells

• Reporting: rare, few, moderate, or many, or as the

actual number per high-power field
• The presence of more than 2 RTE/hpf indicates
TUBULAR INJURY and should be referred for
cytologic exams
• Increased amount is indicative of TUBULAR
necrosis caused by exposure to heavy metals, drug-
induced toxicity etc.

OVAL FAT BODIES

• Highly Refractile RTE Cells that absorb lipids present
in the glomerular filtrate
• Confirmed by staining the sediment using Sudan III or
Oil Red O fat stains.
• Composition of droplets (triglycerides, neutral fats and
cholesterol)
• Under polarized light, they exhibit Maltese Cross
Formation

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(Continuation---Oval Fat Bodies) BUBBLE CELLS
• Clinical significance: Lipiduria associated with • RTE cells containing large, nonlipid-filled vacuoles
nephrotic syndrome. • Seen in acute tubular necrosis and may be seen along
• Are also significant in cases of severe tubular with RTE and Oval Fat Bodies.
necrosis, diabetes mellitus, and in trauma associated • They represent injured cells in which the endoplasmic
with: reticulum has dilated prior to cell death.
- Release of fat from the bone marrow
• Manner of reporting: average number/hpf
BACTERIA
• Not normally present in urine especially if it is collected
under sterile conditions and are analyzed within two
hours after receipt of specimen.
• SPECIMEN WITH A PH OF 8.5 AND 9.0 IS
REJECTED BECAUSE OBVIOUSLY URINE IS
CONTAMINATED WITH BACTERIAL
CONTAMINANTS

• Appearance: Cocci (spherical) or Bacilli (rods) forms

• Reported as few, moderate, many per high power
objective.
• To be considered UTI, bacteria should be
accompanied with predominant WBCs

• Clinical Significance:
- UTI (should be correlated with nitrite and LE test
accompanied by WBC’s in the urine sediment.)
- The presence of motile organisms in a drop of fresh
urine collected under sterile conditions correlates well
with a positive urine culture.

• Observing bacteria for motility also is useful in

differentiating them from amorphous phosphates and
urates. Phase microscopy aids in the visualization of
bacteria.
• The presence of bacteria can be indicative of either
lower or upper UTI. Specimens containing increased
bacteria and leukocytes are routinely followed up
with a specimen for quantitative urine culture.

• Most frequently associated bacteria include the family

Enterobacteriaceae, Staph and Enterococcus
species
• Of course, you cannot identify the bacteria through
microscopic exam, urine culture must still be done.

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 PARASITES

Trichomonas vaginalis
- most frequent parasite encounted in urine
- pear-shaped flagellate with undulating membrane
- a sexually transmitted pathogen causing vaginal
inflammation
- Reported as rare, few, moderate, or many per hpf

YEAST
Schistosoma haematobium
• Appearance: REFRACTILE OVAL STRUCTURES
- ova with terminal spine lacerates the bladder causing
that may contain a BUD.
bloody urine
• In severe infections, they may appear branched, with
- agent of schistosomiasis predominantly in Samar and
mycelial forms and hyphae may be visible.
Leyte.
• Reported as rare, few, moderate or many per hpf.

• Clinical significance: Primary yeast cells in urine are

from a fungus known as Candida albicans - the
causative agent of vaginal moniliasis.
• Are also significant in cases of immunocompromised
patients and those with DIABETES MELLITUS

• Could also be a lab contaminant if there is a delay in

analysis.
• TRUE yeast infection should be accompanied with the Could also be found in cases of fecal contamination
presence of WBCs. usually Enterobius vermicularis ova.

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 SPERMATOZOA/SPERMS
IDENTIFY!
• Oval, slightly tapered heads with long flagella-like tails
• Usually seen in urine after sexual intercourse,
masturbation and nocturnal emission (wet dreams)
• Not reported unless the test is for medico-legal
cases as in rape cases.

▪ CLINICAL SIGNIFICANCE:
 RARE
 considered clinically significant in cases of MALE
INFERTILITY OR RETROGRADE EJACULATION
 This happens when sperms are expelled into the
bladder instead of the urethra.
 Increased amounts of semen results to increased
PROTEIN in urine.

MUCUS THREADS
• These are composed of protein materials produced by
the glands and epithelial cells from the lower
genitourinary tract and the RTE cells.
• Major protein constituent is TAMM-HORSFALL
PROTEIN now known as UROMODULIN.

• Appear as thread-like structures with low refractive

index
• Clumps of mucus threads are usually mistaken for
HYALINE CASTS
• Reported as rare, few, moderate and many/low power
field (lpf)
• Frequent in FEMALE but of no clinical significance
when seen whether on males or females

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