URINALYSIS

Outline
I. Urine examination
overview
A. Specimen collection and
evaluation
B. Gross examination of urine

II. Chemical testing of Urine

III. Post-lecture quiz

 References
• Henry's
Clinical Diagnosis and Management by
Laboratory Methods
• Part 3 - chapter 29

• Urinalysis and Body Fluids by Susan Strasinger

• Part 2 - chapters 4, 5, and 6
Types of urine specimens
Specimen Purpose
Random Routine screening
First morning Routine screening
Pregnancy test
Orthostatic protein
24-hour/Timed Quantitative chemical test
Catheterized Bacterial Culture
Midstream clean- Routine screening
catch Bacterial culture
Suprapubic Bacterial culture
aspirate Cytology
Three glass Prostatic infection
collection
Basic/Routine Urinalysis
(Overview)
• Specimen Evaluation

• Gross or Physical Examination

• Chemical Testing

• Sediment Examination
Specimen Collection
• Observe standard precautions. Always wear gloves!

• Containers:
• Should be disposable.
• Should be made of clear, transparent material with a leak-proof cover.
• Should have a wide mouth and flat bottom.
• Recommended capacity is 50 ml.
• Some require sterile containers.

• Observe proper LABELLING!

Urinalysis Container
 What are grounds for
rejecting a specimen?
• WALANG LABEL!!!

• Wrong label, wrong request form, or wrong na lahat!

• Contaminated specimens (presence of stool or toilet

paper)

• Insufficient volume

• Improperly transported specimens

Specimen Integrity and
Preservation
• Should be promptly delivered and tested within 2 hours.

• Should be refrigerated or have a preservative added if

testing cannot be done immediately.

• Refrigerate at 2 to 8 deg C for up to 24 hours.

• Bring to room temperature prior to testing.

 Physical Examination of
Urine
COLOR CAUSE
Colorless Recent fluid intake
Pale yellow Polyuria, Diabetes Insipidus, DM
Dilute specimen
Dark Yellow Concentrated, Dehydrated
B complex intake
Antibiotics
Yellow-Green Bilirubin/Biliverdin
Green Pseudomonas infection
Pink Presence of RBCs
Red Presence of Hemoglobin or Myoglobin
Beetroot intake
Menstrual contamination
Physical Examination of Urine
COLOR CAUSE
Red-Brown RBCs oxidized to methemoglobin
Myoglobin
Brown Alkaptonuria (Homogentisic acid)
Black Melanin/Melanogens, Malignant
Melanoma
Intake of Methyldopa,
Metronidazole
 Physical examination of
Urine
• Non-pathologic causes of turbid urine
• presence of squamous epithelial cells, semen, vaginal
cream or mucus
• fecal contamination (reject the specimen)
• talc powder
• amorphous phosphates, carbonates and urates,
radiographic contrast media

• Pathologic causes of turbid urine

• presence of bacteria
• chyluria (due to lymph obstruction, ex filariasis)
• fat globules (usually triglycerides) or increased proteins
(nephrotic syndrome)
 Physical Examination of
Urine
AMINO ACID DISORDER ODOR
Cystinuria Rotten eggs
Hawkinsinuria Swimming pool
Ketoacidosis Fruity, Sweet
Isovaleric acidemia and Sweaty feet
Glutaric acidemia
Maple Syrup Urine Disease Maple syrup
Methionine malabsorption Cabbage
Phenylketonuria Mousy, Musty
Trimethylaminuria Rotting fish
Tyrosinemia Rancid
Specific Gravity
• Determined by amount of Urea, NaCl, Sulfates,
Phosphates.
• Normal range is 1.016 to 1.022 in a 24 hour period.

• Hyposthenuria
• SG is less than 1.007
• Seen in pyelonephritis, glomerulonephritis

• Isothenuria
• No variability in SG, fixed at 1.010
• Indicative of severe renal damage

• Methodsto determine: Reagent strip,

Refractometer, Urinometer, Falling drop method
Add/Subtract 0.001 for every 3 degrees above/below calibration
temperature.
Subtract 0.003 for every 1 g/dL of protein
CHEMICAL
ANALYSIS
OF URINE
Chemical Examination of urine
•A strip containing multiple chemical reactions per pad.

• Can
be read manually, but now there are automated
methods available.

• Tests commonly included in the strip are:

• pH, protein, glucose, ketones, blood, bilirubin/urobilinogen, nitrites,
leukocyte esterase, specific gravity
SYSMEX MEDITAPE UC
• Urinalysis
test strip with reagent pads for
determination of diagnostic parameters in
human urine
COBAS
COMBUR STRIP
•A chemical reagent strip
containing the ff tests (in
order):
• Specific Gravity
• pH

• Leukocytes

• Nitrite

• Protein

• Glucose

• Ketone

• Urobilinogen

• Bilirubin

• RBCs/Blood

• Control/Comparison
 Urine pH
• Range is 4.5 to 8.0
• Reagents: Methyl red and Bromthymol blue (pH
indicator)

• First morning specimen is usually more acidic.

• Specimens taken after meals are more alkaline
(“alkaline tide”).

• Clinical significance:
• Respiratory or Metabolic acidosis or alkalosis
• Renal tubular acidosis
• Renal calculi formation
• Treatment of UTI
• Identification/Precipitation of crystals
• Determine unsatisfactory specimens
 Urine protein
• Average protein concentration is 2 to 10 mg/dL.
• One third of protein excreted is Albumin.
• Others are: globulin light chains, B2-microglobulin,
lysozyme, Tamm-Horsfall (uromucoid)

• Important indicator of renal disease.

• Functional proteinuria: Exercise, dehydration, CHF, cold
exposure, fever
• Transient proteinuria: Pregnancy, Postural

• 24hour urine collection is useful for kidney

disease.
• If not available, a UPCR is done instead.
 Urine protein
• Reagent strip method: protein error of pH
• Proteins have a net negative charge. Their presence in
urine changes the pH. (Yellow to Green or Blue)
• More sensitive to Albumin
• Detects 5 to 20mg/dL of Albumin

• Sulfosalicylic acid method (Qualitative)

• Formation of precipitate indicates presence of protein.
• Detects 5 to 10mg/dL of Albumin, globulins,
glycoproteins, bence jones proteins
• Affected by contrast media for up to 3 days.
Micral test (Roche) and
Immunodip test (Dx Chem Ltd)
• Semiquantitative
immunologic test

• Enzyme linked immunoassay

• Uses a gold-labeled antihuman
antibody, which binds to
albumin.
• Color changes from white to red.
• Color intensity reflects amount
of albumin present in the
sample.
Urine glucose
• Glycosuriaoccurs when blood glucose level is
greater than 180 mg/dL.
• For
diabetes monitoring, specimen is tested 2
hours after a meal.

• Othersugars present in urine (usually

disaccharides):
• Fructose – inherited enzyme deficiency
• Galactose – deficient galactokinase
• Lactose – appearance is normal in late
pregnancy
• Pentose
• Sucrose – sucrase deficiency (Sprue)
Urine glucose
• Reagent strip method – glucose oxidase and
peroxidase double sequential enzyme
reaction.
• Specific only for glucose!

• Copper reduction test

• Testsfor presence of all reducing sugars:
fructose, lactose, galactose, maltose, and
pentoses.
Urine Ketones
• Acetoacetic
acid (20%), acetone (2%), 3-
hydroxybutyrate (78%)
• Up to 50mg/dL of acetoacetic acid can be present prior to
onset of symptoms.
• Lacticacidosis can present with high butyrate and low
acetoacetate. This may not be detected in the reagent
strip.

• Ketonuria can be present in:

• Diabetes
• Hyperemesis of pregnancy
• Cechexia
• After administration of anesthesia
• Exposure to cold
• Heavy exercise
• Low carb diet
Urine Ketones
• Commonly used methods: Nitroprusside reagent strip
or Tablet tests

• Reagent strip method: Nitroprusside reaction (Na

nitroferricyanide and glycine)
• Reacts with acetoacetic acid and acetone in an alkaline
medium.
• Read the result at 60 seconds/ 1 minute.
• Can detect 10 mg/dL acetoacetic acid, and 70 mg/dL of
acetone.

• Tablettest: Na nitroprusside, glycine, and alkaline

buffer (same lang)
• Detects 5 to 10 mg/dL acetoacetic acid, and 20 to 25 mg/dL of
acetone.
• Used when urine has an interfering color. Can also be used
with blood and plasma.
• Sensitive to humidity. Store properly!
Blood in the Urine
• Hematuria – rbcs in the urine

• Hemoglobinuria – free hemoglobin in the urine.

• Indicates intravascular hemolysis (with associated
decreased haptoglobin, increased unconj bilirubin,
urobilinogen, and hemosiderin).
• Hemosiderin will be present in the urine after 2 to 3 days.

• Myoglobinuria
– usually in strenuous exercise and
rhabdomyolysis.

• Reagentstrip method: peroxidase-like activity of

heme in hemoglobin.
• In all cases the urine appears red to red-brown. The reagent
strip is also positive in all cases.
• Serum is pink in hemoglobinemia
• Serum is normal in myoglobinemia
Bilirubin/Urobilinogen
• Normal adult urine contains about 0.02 mg of bilirubin.
• Normal output of urobilinogen is 0.5 to 2.5 mg/24 hours.
• Bilirubin
– indicator of liver disease (intra or
extrahepatic).
• Urobilinogen – increased amounts appear during
hepatocellular damage (hepatitis, drugs, toxins,
cirrhosis), and hemolysis.

• Reagent strip method: coupling reaction of bilirubin with

a diazonium salt in acid medium or Ehrlich aldehyde
reaction.
• Specimen should be fresh because bilirubin can quickly hydrolyze.
• Specimens left standing will oxidize, causing a false negative
result.
Bilirubin/Urobilinogen
• Ictotest (tablet test for bilirubin, brand name)
• Tablets contain nitrobenzene diazonium toluene,
sulfosalicylic acid, and sodium bicarbonate.
• Add water and sample then observe color change.
• Tablets are hygroscopic. Protect from moisture and
humidity.
• Exposure to light can cause decomposition of the
tablet.
Testing for UTI
• Patients
can be asymptomatic. Commonly in the following
populations:
• Pregnant
• Elderly
• Diabetic patients
• Previous history of UTI

• Indirect
assessment of UTI includes: Nitrites and Leukocyte
Esterase activity in the urinalysis reagent strip.
• The gold standard is still a bacterial culture.
Nitrites
• Bacteria
present in significant numbers ( > 105 to 106/mL)
can reduce nitrate to nitrite.
• This is true for GRAM-NEGATIVE bacteria which are the common agents
of UTI.
• Other bacteria or yeasts who lack the reductase enzyme may produce a
negative result!

• Reagent strip method: Diazonium salt formation (Greiss

reaction).
• Arsanilic acid reacts with nitrite in the urine to form a salt.
Leukocyte Esterase
• This
enzyme is present mostly in neutrophil primary
granules.
•A positive result indicates significant amounts of wbcs in the
urine, and may suggest an infection.

• Reagent strip method: naphthol chloroacetate reaction.

• Neutrophil leukocyte esterase catalyzes the hydrolysis of esters to
produce alcohols and acids.
• The alcohols react with a diazonium salt to form a colored compound.
There are more tests not present in
the reagent strip, for your own
reading…
• SeeHenry’s chapter 29, basic examination of
urine
• Other tests include:
• Ascorbic acid test
• Serotonin test (5 hydroxyindoleacetic acid
test)
• Testing for melanin/melanosomes
• Test
for porphyrins (test for urobilinogen vs
porphobilinogen)
• Watson-Schwartz test
• Inverse Ehrlich aldehyde reaction/ Hoesch test
 Principles of Examination
Test Principle Reference
Range
UROBILINOGE Azo coupling method. 0.03 to 0.97
N Color changes from light pink to mg/dL
red.
BLOOD Peroxidase-like reaction of Less than
hemoglobin. 5 cells/HPF
Color changes from white to blue.
PROTEIN Protein error of pH indicator. Less than
Color changes from yellow to 30 mg/dL
blue-green.
GLUCOSE Enzymatic method (glucose 2 to 20 mg/dL
oxidase peroxidase).
Color changes from yellow to
blue-green.
KETONE Nitroprusside method. Less than
Color changes from buff pink to 2 mg/dL
 Principles of Examination
Test Principle Reference
Range
BILIRUBIN Azo coupling method. Less than 0.05
Color changes from light tan to mg/dL
light pink.
NITRITES Griess reaction. (Diazo reaction) Positive or
Color changes from light green to Negative only
purple.
LEUKOCYTES Measurement of leukocyte Less than 12
esterase activity. cells/uL
Color changes to violet.
PH pH indicator method. 4.5 to 7.5
Color changes from orange to
green
(indicates pH of 5 to 9)
ALBUMIN Protein error of pH indicator. Less than 23.8
Color changes from yellow to blue- mg/L
green.
AUTOMATED URINALYSIS
QUESTION TIME!!!
5 questions lang pramis.

-Oh no di ako nakinig kanina-

What is read in A and B?

(A) (B)
What is the correction
factor for temperature?
This is a urinometer.

0.001 is added for every 3

degrees above calibration
temperature.
0.001 is subtracted for
every 3 degrees below
calibration temperature.
What does this test measure?

Benedicts test ito.

This tests for the presence of reducing

sugars.
Negative yung BLUE.
What is the principle of this test?

This is the acetest (ketone tablet test).

Nitroprusside reaction is the principle.

Positive in moderate amounts (middle).

Positive in large amounts (two tablets).
Is this a direct or indirect
method to measure
Specific Gravity?

This is a refractometer.
It is an indirect method to measure specific gravity.
It measures the refractive index of the solution (urine).

The only direct method is URINOMETRY!

 THE END!

(OF PART 1)
-cry ka nalang muna kasi mahaba pa
ang laban-