MICROSCOPIC EXAMINATION OF URINE

Urine sediment preparation

1st step

Transfer 10-15 mL of urine in a test tube

Urine sediment preparation

Recommended urine volume

12 mL

Urine sediment preparation

2nd step

Centrifuge tube at 400 RCF for 5 mins

Urine sediment preparation

3rd step

Decant urine

Urine sediment preparation

After decanting, how many mL of urine remains in the tube?

0.5 or 1.0 mL

Urine sediment preparation

4th step

Transfer 20 uL (0.02 mL) urine sediment to glass slide with 22×22 mm coverslip

Urine sediment preparation

5th step

Examine microscopically (10 LPF, 10 HPF, under reduced light)

If < 12 mL urine is available for microscopy, we must?

Centrifuge 3 mL

No centrifugation needed if urine volume is?

< 3 mL

RCF formula

1.118 × 10² x radius in centimeters x RPM²

Quantitative measure of formed elements of urine using hemacytometer

Addis count

Addis count recommended specimen

12-hour urine

Addis count recommended preservative

Formalin

Addis count normal values: RBC

0-500,000 / 12-hr

Addis count normal values: WBCs and ECs

0-1,800,000 / 12hrs

Addis count normal values: Hyaline casts

0-5000 / 12hrs

Where is the first lens system located?

Objective

Second lens system location

Eyepiece (ocular lens)

Ability to distinguish 2 small objects that are a specific distance apart

Resolution

Microscopes requiring minimum adjustment when switching objectives

Parfocal

Used to remove dust on the optical surface of the microscope

Camel-hair brush

Used to clean the optical surfaces of the microscope

Lens paper

Used to clean any contaminated lens

Commercial lens cleaner

How to remove oil on lens?

Dry lens paper, then lens paper moistened w/ lens cleaner

Xylene is not recommended because of its?

Toxic fumes

Performs initial focusing (LPO and SO)

Moves stage noticeably up and down

Corase adjustment know

Focuses and controls the light on the specimen

Condenser

Controls the amount and angle of light

Aperture diaphragm

Further magnifies image 10x (secondary magnification)

Oculars

Controls the diameter of the light beam reaching the slide

Field diaphragm

Sharpens the image (HPO and OIO)

Fine adjustment knob

Adjusts distance between oculars horizontally

Interpupillary distance control

Lenses which form primary (initial) image of specimen

Objectives

Regulate intensity of the light

Rheostat

Microscope used for routine UA

Bright-field microscopy

Used for translucent elements with low refractive index

Phase-contrast microscope

Phase-contrast microscopy produces what around an object?

Halo of light

How to convert BF to PC microscope?

Replace objective lens and condenser with PC objective lens and condenser

For identification of cholesterol in oval fat bodies, fatty casts, and crystals

Polarizing microscope

Detects presence or absence of birefringence

Polarizing microscopy

How to convert BF into polarizing?

Add 2 filters (1 below the condenser and 1 between objective and oculars)

For identification of T. pallidum

Dark-field microscope

How to convert BF to DF?

Replace condenser with a DF condenser that contains an opaque disk

Visualization of fluorescent substances and microorganisms

Fluorescence microscope

3D microscopy image and layer-by-layer imaging of a specimen

Interference contrast microscope

What can be adapted for interference contrast microscopy?

Bright-field

Two types of interference contrast mircosope

Nomarski

Hoffman

Nomarski aka?

Differential interference contrast

Hoggman aka?

Modulation contrast

Sediment stains

Delineates structure and contrasting colors of the nucleus and cytoplasm

Sternheimer-Malbin

Sediment stains

Identifies WBCs, epithelial cells, and casts

Sternheimer-Malbin

Sediment stains

Most commonly used supravital stain

Sternheimer-Malbin

Sediment stains

Sternheimer-Malbin composition

CV + safranin O

Sediment stains

Enhances nuclear detail and differentiates WBCs and RTE cells

Toluidine blue

Sediment stains

Lyses RBCs, enhances nuclei of WBCs, distinguishes RBCs from WBCs, yeast, oil droplets, and crystals

2% acetic acid

Sediment stains

Stains triglycerids and neutral fats but not cholesterol

Oil Red O

Sudan III

Sediment stains

Lipid stains color

Orange-red

Sediment stains

Identifies free fat droplets and lipid-containing cells and casts

Oil Red O

Sudan III

Sediment stains

Differentiates Gram-pos and Gram-neg bacteria

Identifies bacterial casts

Gram stain

Sediment stains

Stains eosinophilic granules

Identifies urinary eosinophils

Hansel stain

Sediment stains

Hansel stain composition

Eosin Y + Metyhlene blue

Sediment stains

Stains structures containing iron

Identifies hemosiderin granules

Prussian blue

Sediment stains

Prussian blue aka?

Rous test

Sediment stains

Stains DNA

Phenathridine

Sediment stains

Stains nuclear membranes, mitochondria, and cell membranes

Carbocyanine

Normal value for RBCs in urine

0-2 or 0-3 / HPF

RBC characteristic in hypertonic urine

Crenate / shrink

RBC characteristic in hypotonic urine

Swell / Lyse

RBC in hypotonic urine known as?

Ghost cell

RBC characteristic in glomerular membrane damage

Dysmorphic, with projections, fragmented (acanthocytes)

RBCs in urine sources of false positive

Yeasts, oil droplets, air bubbles, monohydrate calcium oxalate crystals

Remedy for false positive in RBCs in urine

2% acetic acid

Normal value of WBCs in urine

0-5 or 0-8 / HPF

Most predominant WBC in urine

Neutrophils

In hypotonic urine, neutrophils will _______ and granules undergo ____________

Swell ; Brownian movement

Neutrophils in hypotonic urine known as?

Glitter cells

Neutrophils, when dying, form?

Blebs and finger-like projections (myelin forms)

Normal value of eosinophils in urine

1%

Significant eosinophil value

> 1%

Eosinophil > 1% in urine is seen in?

Acute interstitial nephritis

Increased lymphocytes in urine seen in?

Renal transplant rejection

Increased monocytes and histiocytes in urine seen in?

Chronic inflammation and radiation therapy

When using Sternheimer-Malbin stain, glitter cells appear?

Pale blue

When using Sternheimer-Malbin stain, leukocytes appear?

Pale pink

Considered as the point of reference in microscopic examination of urine

Squamous epithelial cell

Largest cell with abundant, irregular cytoplasm and prominent nucleus

SECs

SEC cell size

30-50 um (5-7x more than an RBC)

SEC covered with G. vaginalis

Clue cell

Clue cell characteristic

Shaggy cytoplasm

Transitional epithelial cell aka?

Urothelial / Bladder cell

TEC cell size

20-30 um (4-6x size of RBC)

Spherical, polyhedral or caudate with centrally located nucleus

TEC

TEC derived from?

Renal pelvis, calyces, ureter, urinary bladder, and upper male urethra

What cell is increased following catheterization?

TEC

Most clinically significant epithelial cell

Renal tubular epithelial cell

Origin of RTE

Nephron

RTE nucleus?

Eccentric`

If RTE is from PCT:

Large, oblong, cigar-shaped with grainy cytoplasm

If RTE is from DCT:

Smaller, round to oval, may be mistaken for WBCs

If RTE is from CD:

Columnar, polygonal, or cuboidal with flat edge