1. Urinalysis
Course: IDPT 5005
School of Medicine, UCDHSC
Francisco G. La Rosa, MD
Francisco.LaRosa@uchsc.edu
Assistant Professor, Department of Pathology
University of Colorado at Denver Health Science Center, Denver, Colorado
2. Specimen
Collection
– First morning voiding (most concentrated)
– Record collection time
– Type of specimen (e.g. “clean catch”)
– Analyzed within 2 hours of collection
– Free of debris or vaginal secretions
5. Types of Analysis
− Macroscopic Examination
− Chemical Analysis (Urine
Dipstick)
− Microscopic Examination
− Culture (not covered in this lecture)
− Cytological Examination
6. Macroscopic Examination
Odor:
− Ammonia-like: (Urea-splitting bacteria)
− Foul, offensive: Old specimen, pus or inflammation
− Sweet: Glucose
− Fruity: Ketones
− Maple syrup-like: Maple Syrup Urine Disease
Color:
− Colorless Diluted urine
− Deep Yellow Concentrated Urine, Riboflavin
− Yellow-Green Bilirubin / Biliverdin
− Red Blood / Hemoglobin
− Brownish-red Acidified Blood (Actute GN)
− Brownish-black Homogentisic acid (Melanin)
7. Macroscopic Examination
Turbidity:
− Typically cells or crystals.
− Cellular elements and bacteria will clear by
centrifugation.
− Crystals dissolved by a variety of methods (acid or
base).
− Microscopic examination will determine which is
present.
11. Uses and Limitations of Urine Glucose
Detection
Significance
– Diabetes mellitus.
– Renal glycosuria.
Limitations
– Interference: reducing agents, ketones.
– Only measures glucose and not other sugars.
– Renal threshold must be passed in order for glucose to spill
into the urine.
Other Tests
– CuSO4 test for reducing sugars.
12. Detection of Reducing Sugars* by
CuSO 4
Sugar Disease(s)
- Galactose Galactosemias
- Fructose Fructosuria, Fructose
Intolerance, etc.
- Lactose Lactase Deficiency
- Pentoses Essential Pentosuria
- Maltose Non-pathogenic
* NOT Sucrose because it is not a reducing sugar
13. Urine versus Blood Glucose
Urinalysis Glucose Result
++
+
trace
Negative
200 400 600 800 1000
Blood Glucose (mg/dL)
14. The Urine
Dipstick: Bilirrubin
Negative Chemical Principle
+ (weak)
Bilirubin + Diazo salt Acidic Azobilirubin
--------->
++ (moderate)
Read at 30 seconds
+++ (strong) RR: Negative
15. Uses and Limitations of Urine Bilirrubin
Detection
Significance
- Increased direct bilirubin (correlates with urobilinogen and serum
bilirubin)
Limitations
- Interference: prolonged exposure of sample to light
- Only measures direct bilirubin--will not pick up indirect bilirubin
Other Tests
- Ictotest (more sensitive tablet version of same assay)
- Serum test for total and direct bilirubin is more informative
17. Uses and Limitations of Urine Ketone Detection
Significance
- Diabetic ketoacidosis
- Prolonged fasting
Limitations
- Interference: expired reagents (degradation with exposure to
moisture in air)
- Only measures acetoacetate not other ketone bodies (such as in
rebound ketosis).
Other Tests
- Ketostix (more sensitive tablet version of same assay)
- Serum glucose measurement to confirm DKA
18. The Urine
Dipstick: Specific
Gravity
1.000
Chemical Principle
1.005
X+ + Polymethyl vinyl ether / maleic anhydride
1.010 --------------->
X+-Polymethyl vinyl ether / maleic anhydride + H+
1.015
H+ interacts with a Bromthymol Blue indicator to
1.020 form a colored complex.
1.025 Read up to 2 minutes
RR: 1.003-1.035
1.030
19. Uses and Limitations of Urine Specific Gravity
Significance
- Diabetes insipidus
Limitations
- Interference: alkaline urine
- Does not measure non-ionized solutes (e.g. glucose)
Other Tests
- Refractometry
- Hydrometer
- Osmolality measurement (typically used with water deprivation test)
20. The Urine
Dipstick: Blood
Negative
Chemical Principle
Trace (non-hemolyzed)
Lysing agent to lyse red blood cells
Moderate (non-hemolyzed)
Diisopropylbenzene dihydroperoxide +
Tetramethylbenzidine
Trace (hemolyzed)
Heme
------------> Colored Complex
+ (weak)
++ (moderate) Read at 60 seconds
RR: Negative
Analytic Sensitivity: 10 RBCs
+++ (strong)
21. Uses and Limitations of Urine Blood Detection
Significance
- Hematuria (nephritis, trauma, etc)
- Hemoglobinuria (hemolysis, etc)
- Myoglobinuria (rhabdomyolysis, etc)
Limitations
- Interference: reducing agents, microbial peroxidases
- Cannot distinguish between the above disease processes
Other Tests
- Urine microscopic examination
- Urine cytology
22. The Urine
Dipstick: pH
5.0
6.0 Chemical Principle
H+ interacts with:
6.5
Methyl Red (at high concentration; low pH) and
Bromthymol Blue (at low concentration; high
7.0
pH), to form a colored complexes
(dual indicator system)
7.5
8.0 Read up to 2 minutes
R.R.: 4.5-8.0
8.5
23. Uses and Limitations of Urine pH Detection
Significance
- Acidic (less than 4.5): metabolic acidosis, high-protein diet
- Alkaline (greater than 8.0): renal tubular acidosis (>5.5)
Limitations
- Interference: bacterial overgrowth (alkaline or acidic),
“run over effect” effect of protein pad on pH indicator pad
Other Tests
- Titrable acidity
- Blood gases to determine acid-base status
24. pH Run Over Effect
Glucose
Bilirubin
Ketones
Specific Gravity Buffers from the protein area of
the strip (pH 3.0) spill over to the
Blood pH area of the strip and make the
pH pH of the sample appear more
acidic than it really is.
Protein
Urobilinogen
Nitrite
Leukocyte Esterase
25. The Urine
Dipstick: Protein
Chemical Principle
“Protein Error of Indicators Method”
Negative Pr
H Pr
H Pr
Trace H
Pr Pr
+ (30 mg/dL) H H
H Pr
++ (100 mg/dL) Tetrabromphenol Blue
H+ H H
+ +
(buffered to pH 3.0)
H+ H H
+ +
+++ (300 mg/dL) Pr Pr
Pr
Pr Pr
++++ (2000 mg/dL) Pr
Read at 60 seconds
RR: Negative
26. Causes of
Proteinuria
Functional Renal
- Severe muscular exertion - Glomerulonephritis
- Pregnancy - Nephrotic syndrome
- Orthostatic proteinuria - Renal tumor or infection
Pre-Renal Post-Renal
- Fever - Cystitis
- Renal hypoxia - Urethritis or prostatitis
- Hypertension - Contamination with vaginal
secretions
28. Uses and Limitations of Urine Protein
Detection
Significance
- Proteinuria and the nephrotic syndrome.
Limitations
- Interference: highly alkaline urine.
- Much more sensitive to albumin than other proteins
(e.g., immunoglobulin light chains).
Other Tests
- Sulfosalicylic acid (SSA) turbidity test.
- Urine protein electrophoresis (UPEP)
- Bence Jones protein
29. Proteins in “Normal” Urine
Protein % of Total
Daily Maximum
Albumin 40%
60 mg
Tamm-Horsfall 40%
60 mg
Immunoglobulins 12%
24 mg
Secretory IgA 3%
6 mg
Other 5%
31. Uses and Limitations of Urobilinogen
Detection
Significance
- High: increased hepatic processing of bilirubin
- Low: bile obstruction
Limitations
- Interference: prolonged exposure of specimen to oxygen
(urobilinogen ---> urobilin)
- Cannot detect low levels of urobilinogen
Other Tests
- Serum total and direct bilirubin
32. The Urine
Dipstick: Nitrite
Chemical Principle
Acidic
Negative Nitrite + p-arsenilic acid -------> Diazo compound
Diazo compound + Tetrahydrobenzoquinolinol
Positive
----------> Colored Complex
Read at 60 seconds
RR: Negative
33. Uses and Limitations of Nitrite Detection
Significance
- Gram negative bacteriuria
Limitations
- Interference: bacterial overgrowth
- Only able to detect bacteria that reduce nitrate to nitrite
Other Tests
- Correlate with leukocyte esterase and
- Urine microscopic examination (bacteria)
- Urine culture
35. Uses and Limitations of Leukocyte Esterase Detection
Significance
- Pyuria
- Acute inflammation
- Renal calculus
Limitations
- Interference: oxidizing agents, menstrual contamination
Other Tests
- Urine microscopic examination (WBCs and bacteria)
- Urine culture
36. Microscopic Examination
General Aspects
Preservation
- Cells and casts begin to disintegrate in 1 - 3 hrs. at room temp.
- Refrigeration for up to 48 hours (little loss of cells).
Specimen concentration
- Ten to twenty-fold concentration by centrifugation.
Types of microscopy
- Phase contrast microscopy
- Polarized microscopy
- Bright field microscopy with special staining
(e.g., Sternheimer-Malbin stain)
37. Microscopic Examination
Abnormal Findings
Per High Power Field (HPF) (400x)
– > 3 erythrocytes
– > 5 leukocytes
– > 2 renal tubular cells
– > 10 bacteria
Per Low Power Field (LPF) (200x)
– > 3 hyaline casts or > 1 granular cast
– > 10 squamous cells (indicative of contaminated specimen)
– Any other cast (RBCs, WBCs)
Presence of:
– Fungal hyphae or yeast, parasite, viral inclusions
– Pathological crystals (cystine, leucine, tyrosine)
– Large number of uric acid or calcium oxalate crystals
38. Microscopic Examination
Cells
Erythrocytes
- “Dysmorphic” vs. “normal” (> 10 per HPF)
Leukocytes
- Neutrophils (glitter cells) More than 1 per 3 HPF
- Eosinophils Hansel test (special stain)
Epithelial Cells
- Squamous cells Indicate level of contamination
- Renal tubular epithelial cells Few are normal
- Transitional epithelial cells Few are normal
- Oval fat bodies Abnormal, indicate Nephrosis
48. Microscopic Examination
Bacteria & Yeasts
Bacteria
- Bacteriuria More than 10 per HPF
Yeasts
- Candidiasis Most likely a contaminant
but should correlate with
clinical picture.
Viruses
- CMV inclusions Probable viral cystitis.
92. Case Diluted urine, request a voided urine in the morning
If persisting low SG, possible diabetes insipida
1 A microscopic may give negative results
Glucose Negative
Bilirubin Negative
A 35-year old man undergoing routine pre
employment drug screening.
Ketones Negative
Physical characteristics: Clear.
S.G. 1.001 Microscopic: Not performed.
Drugs Identified: None.
Blood Negative
pH 5.5
Questions:
Protein Negative - What is your differential diagnosis?
- What would you do next to confirm your
Urobilinogen 0.2 mg/dL suspicion?
- Would you order a microscopic analysis
Nitrite Negative on this sample?
L.E. Negative
93. Case Possible gallbladder or hepatic disease.
No hemolytic anemia. Perform bilirubins in serum
2 Microscopic unlikely to provide additional info
Glucose Negative
Bilirubin +++
A 42-year old woman presents with “dark urine”
Ketones Negative
Physical characteristics: Red-brown.
S.G. 1.020 Microscopic: Not performed.
Blood Negative
Questions:
pH 5.5
- What is your differential diagnosis?
Protein Negative - Could this be a case of hemolytic anemia?
- How would you rule it out?
Urobilinogen 0.2 mg/dL - What tests would you order next? Why?
Nitrite - Would you order a microscopic analysis?
Negative
L.E. Negative
94. Case Possible UTI, request culture and antibiotic sensitivity
Negative Nitrite test: Gram positive bacteria
3 Lower SG may show less number of cells and bacteria
Un-common diagnosis in this type of patient
Glucose Negative
A 42-year old man presents painful urination
Bilirubin Negative
Ketones
Physical characteristics: dark red, turbid
Negative
Microscopic: leukocytes = 30 per HPF
S.G. 1.030 RBCs = >100 per HPF
Bacteria = >100 per HPF
Blood +++
Questions:
pH 6.5 - What is your suspected diagnosis?
- What would you do next?
Protein Trace
- What do you make of the nitrite test?
Urobilinogen 1.0 mg/dL - How would the microscopic exam differ if
the S.G. were 1.003?
Nitrite Negative - Is this a common diagnosis for this type of
patient?
L.E. +++
95. Case Diabetes
May be decompensated and with ketoacidosis
4 Ketones should become negative after treatment
Glucose ++
Bilirubin Negative
Ketones Trace A 27-year old woman presents with severe
abdominal pain.
S.G. 1.015
Physical characteristics: clear-yellow.
Blood Negative Microscopic: Not performed.
pH 6.0
Questions:
Protein Negative - What is the most likely diagnosis?
- What do you make of the ketone result?
Urobilinogen 1.0 mg/dL - What do you expect to happen to the ketone
measurement when treatment begins?
Nitrite Negative
L.E. Negative
96. Case Glomerulonephritis
RBC casts reveals renal cortex involvement
5 RBC cast are not always present in GN
Glucose Negative
8-year old boy presents with discolored urine
Bilirubin Negative
Ketones Physical characteristics: Red, turbid.
Negative
Microscopic: erythrocytes = >100 per HPF
S.G. 1.015 (almost all dysmorphic)
Red cell casts present.
Blood +++
Questions:
pH 6.5 - What is the most likely diagnosis in this
Protein case?
+
- Does the presence of red cell casts help you
Urobilinogen 1.0 mg/dL in any way?
- If the erythrocytes were not dysmorphic
Nitrite Negative would that change your diagnosis?
L.E. Negative
97. Case “Functional” proteinuria?
Microscopic may reveal a few leukocytes
6 Request protein concentration in 24 h urine
Glucose Negative
22-year old man presenting for a routine
Bilirubin Negative
physical required for admission to medical
Ketones Negative school
S.G. 1.010 Physical characteristics: Yellow
Microscopic: Not performed
Blood Negative
Questions:
pH 5.0
- What is your differential diagnosis?
Protein + - Would you order a microscopic analysis on
this sample?
Urobilinogen 0.2 mg/dL - What would you do next to confirm the
diagnosis?
Nitrite Negative
L.E. Negative