CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 1

Routine Urinalysis- Physical and Chemical Examination

f U i

University of Nebraska Medical Center

of Urine

CLS 500: Application and Interpretation of Clinical Laboratory Data

Objectives: On completion of this unit, participants will be able to:

• Describe considerations on collection, handling, and processing of urine specimens

• Identify screening tests for physical andIdentify screening tests for physical and chemical examination of urine

• Describe major sources of error in routine urinalysis procedures

• Define common terms applied to urinalysis and renal disease 2

Objectives (continued)• Correlate physical & chemical tests of

urine in:– Cystitis

– Pyelonephritis

– Renal glycosuria

– Diabetes mellitus

– Diabetes insipidus

– Hepatic, obstructive or hemolytic jaundice

– Acute and chronic glomerulonephritis

– Nephrotic syndrome3

Objectives (continued)

• Explain the significance of the following:– Pos Clinitest and neg dipstick for glucose– Pos dipstick for glucose and neg Clinitest– False positive dipstick for proteinp p p– False negative for ketones– False negative for bilirubin– Neg bilirubin dipstick and pos Ictotest– Pos bilirubin and neg urobilinogen– Neg bilirubin and pos urobilinogen

4

Objectives (continued)

• Predict the potential changes that may take place in a urine specimen that remains at room temperature for longer than 2 hours

5

What is Urine?• A fluid which is continuously formed

in and excreted from the body

• Composed of water and

metabolic waste productsmetabolic waste products

• An actual fluid biopsy of the kidney– kidney is the only organ with such a noninvasive means

by which to evaluate its status

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CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 2

Functions of the Kidney

• Produces urine

• Maintains electrolyte balance

• Maintains blood pH

• Produces hormones

• Excretes waste

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The Purpose of Urinalysis?

• To aid in the diagnosis of disease

• To monitor wellness (screening for asymptomatic, congenital, or hereditary disease))

• To monitor the progress of disease

• To monitor therapy (effectiveness or complications)

A Complete Urinalysis Providesa Fountain of Information

pancreas muscle

intoxication

acid base equilibrium inborn errors of metabolismwater status

cardiovascular system

blood carbohydrate metabolism

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bonekidney

drug abuse

pregnancy

liver

hormones

infection

respiratory system

fat metabolism

protein metabolism

urinary tract

gastrointestinal tract

nutritionelectrolytes

central nervous system

Types of Urine Specimens• Random• First morning• Midstream clean catch• Fasting• Catheterized• Suprapubic aspiration• Pediatric specimen• Timed collections (for quantitative testing)

– 2-hour postprandial– 24-hour, 12-hour, 6-hour 10

General Considerations

• Use clean, dry, disposable, sterile container

• Label properly

• ANALYZE WITHIN ONE HOUR

• Preserve urine constituents– Refrigeration (2-8C)

– Advantages vs disadvantages11

Changes at Room Temperature

• Increased pH• Decreased glucose• Decreased ketones• Decreased bilirubin• Decreased urobilinogen• Increased bacteria• Increased turbidity• Disintegration of red blood cells and casts• Changes color

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CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 3

Collection Errors

• Labeling (label the container, not lid)– Patient name

– Patient identification number (MRN, DOB)

– Date of collectionDate of collection

– Time of collection

– Test ordered

– Name of ordering physician/clinician

• Delay in testing– Testing after one hour

– Lack of refrigeration 13

Historical Perspective: Urinalysis• Physical examination

of urine– Odor

– Taste

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– Color

– Clarity

– Volume

Historical Perspective

• Chemical examination of urine– Limited reactions/large volumes required

– Time consuming/cumbersome

– Clinical usefulness was not realized

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Clinical usefulness was not realized

– Not routinely ordered

• Microscopic exam of urine– Clinical usefulness not

realized until invention of

the microscope

Reagent Strip Testing

• Technology and necessity

• Chemical reactions ‘miniaturized’

• Required less urine

U i t ll t

16

• Urine easy to collect

• Test results within minutes

• Easy to perform

• Increased test utilization

Brunzel, 2nd Ed, page 124

Reagent Strip Testing

• Ideal qualitative screening tool– Sensitive: Low concentration of substances

Negative result = normal

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– Specific: Reacts with only one substanceFalse negative and false positive

– Cost effective: Relatively inexpensive tool that provides information about the health status of the patient

Complete Urinalysis

• Physical Examination– Color, Clarity, Concentration

– Odor, Volume

C• Chemical Examination– 10 chemical reactions

• Microscopic Examination

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CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 4

Physical Examination of Urine

Color, Clarity, Foam,

Concentration, Odor, Volume

Physical Properties

• Volume– Average: 1200 to 1500 ml/24 hours

– ‘Extremes’: 600-2,000 ml/24 hrs

• TermsTerms– Anuria -no urine output

– Polyuria -increased urine output

– Oliguria -decreased urine output

– Nocturia -excessive output at night

– Dysuria -painful urination20

Color

BrownHemoglobin

BluePseudomonas infection

Red Green

ColorlessDilute Urine

YellowNormal

OrangePyridium

AmberBilirubin

Blood

Bright YellowVitamins

Medication

Dark GreenBiliverdin

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Clarity / Appearance

ClearLike water; easily read newsprint

CloudyLines are barely visible; cannot read newsprint

HazyBlurry but

lines are still visible

TurbidCannot see through it; often have particulates

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What Can Cause Urine to be Hazy, Cloudy, or Turbid?

Epithelial

Cells

White Blood

Red

Blood

Amorphous

MaterialCells Blood Cells

Blood Cells

Material

Sperm Fat Mucus Yeast

Bacteria Powder Casts Crystals

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Foam

• White foam– protein is

presentpresent

• Yellow foam – bilirubin is

present

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 5

Concentration

• Concentration is determined by the specific gravity test

• 2 ways specific gravity can be determinedR f t t– Refractometer

– Reagent Strip• Most labs use this method

• Increased glucose and protein levels do not interfere

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Odor• Faintly aromatic

– Normal

• Ammonia – Old urine

• Pungent– UTIU

• Fruity, Sweet– Ketones

• Mousy, Barny– PKU disease

• Maple syrup– Maple syrup urine

disease 26

Taste – thank goodness we don’t do this anymore!!

This doesn’t taste like lemonade!!!

Chemical Examination of Urine

Reagent Strip Testing

Reagent Strip Testing• Chemically impregnated absorbent

pads attached to an inert plastic strip

• Each pad is a specific chemical reaction that

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takes place upon contact with urine

• Chemical reaction causes the color of the pad to

change: reaction is timed

• Color compared to a color chart for interpretation

Reagent Strip Proper Storage

• Tightly closed container

• Cool dry place

• Avoid volatile fumes

• Expiration date

• Do not use if pads are discolored

• Do not touch pads

• Run positive and negative controls once/day30

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 6

Quality Control• Ensures reliability of results

• Evaluates reagent strip

• Run 2 controls once a day

• Positive control

N ti t l

31

• Negative control

• BOTH controls MUST be ‘ok’ else patient testing cannot be performed

The Dipstick Procedure

• Wear gown & gloves

• Mix the urine

• Insert reagent stripg p

• Remove excess urine

• Time the reactions

• Compare test areas to color chart

• Record results

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The Urine Dipstick10 Reactions on 1 Plastic Strip

• Glucose• Bilirubin• Ketones• Specific gravity

• Purpose of the test

• What is normal

• What is abnormal• Specific gravity• Blood• pH• Protein• Urobilinogen• Nitrites• Leukocytes

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• Causes of abnormal results

• Causes of false pos/neg results

Glucose

• All glucose is normally reabsorbed in the tubules unless the blood level is higher than the renalunless the blood level is higher than the renal threshold (160 to 180 mg/dl)

• Normal = Negative

• Abnormal = Diabetes mellitusImpaired tubular reabsorptionInborn errors of metabolism

34

Glucose

• Glucosuria

Glycosuria

• Caused by renal and non renal disease

Terms used interchangeably

35

• Caused by renal and non-renal disease– Pre-renal glycosuria: plasma glucose level

exceeds renal threshold (diabetes mellitus)

– Renal glycosuria: plasma glucose level below renal threshold, but tubules cannot reabsorb glucose back into bloodstream

Glucose• Sensitivity: ~ 100 mg/dl

• Specificity: – Reacts only with glucose

– False positive:

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False positive: • Strong oxidizing agents (bleach)

• Peroxides

– False negative: • Ascorbic acid (reducing agent)

• High ketone levels

• Improperly stored urine: glycolysis

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 7

Clinitest (tablet test)

• Is a copper reduction test (cupric to cuprous)

• Detects all reducing sugars

• Reacts with glucose, galactose, lactose, fructose, ascorbic acid, homogentisic acid; , , g ;not sucrose

• All children <2 years: screened for metabolic disorder (galactosemia)

37

• Clinitest is non-specific– Reacts with all reducing substances

• Clinitest not as sensitiveWill d t t l t 250 /dl

Glucose Dipstick vs Clinitest

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– Will detect glucose at 250 mg/dl

• Dipstick is specific for glucose (enzyme rxn)

• Dipstick more sensitive– Will detect glucose at 100 mg/dl

Bilirubin

• Bilirubin is formed from the breakdown of hemoglobin in the reticuloendothelial (RE) g ( )system

• Only conjugated bilirubin is found in urine

• Normal = Negative

• Abnormal = Liver disease-hepatic jaundiceObstruction-obstructive jaundice39 40

Bilirubin• False Negative

– Ascorbic acid inhibits

– High urine nitrites inhibit

– Low bilirubin concentration

I i h dli t t f li ht– Improper specimen handling: protect from light

• False Positive– Urine color interference

– Drug induced color changes: phenazyridine, indican-indoxyl sulfate

– Perform Ictotest to confirm presence of bilirubin41

Bilirubin Dipstick vs Ictotest

• Specificity is the same: both react with conjugated bilirubin

S iti it diff

42

• Sensitivity differs

Reagent strip: ~0.5 mg/dl

Ictotest: 0.05 – 0.1 mg/dl

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 8

Ketones

• Ketones are products of incomplete fat metabolism Normally fats are completely metabolized to CO2

d tand water

• Normal = Negative

• Abnormal = Inability to utilize carbohydratesDiabetes mellitus (DKA)Inadequate intake of carbohydratesExcessive loss of carbohydrates 43

Ketones

• Fruity odor = acetone

44

Ketones

• False Negatives– Improper storage

• Volatilization

• Bacterial breakdown

• False Positives– Compounds containing free-sulfhydryl groups

– Highly pigmented urines (color interference)

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Specific Gravity

• Specific gravity measures the concentrating and diluting abilities of the kidney; results are dependent on hydration statuson hydration status

• Normal = 1.002 to 1.035

Majority of urines are 1.010-1.025

• Abnormal = Increased SG means the urine is concentrated

= Decreased SG means the urine is dilute46

Specific Gravity

• Physiologically impossible

>1.040 (suspect interfering substance)

1.000 (suspect water)

• Sensitivity: 1.000

• Specificity: detects only ionic substances– Radiographic dye

– Mannitol

– Glucose47

Does not interfere with this method

Specific Gravity: Terms• Isosthenuria

– Fixed at 1.010

– Renal tubules lost absorption and secreting capability

• Hypersthenuria

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– Increased specific gravity

– Concentrated urine

– Hypertonic

• Hyposthenuria– Decreased specific gravity

– Dilute urine

– Hypotonic

Sensitivity issues:

Pregnancy testing

Urinary tract infection

Protein

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 9

Blood

• The presence of blood in the urine may indicate damage to the kidney or urinary tractg y y

• Normal = Negative

• Abnormal = Kidney stones (renal calculi)

Glomerulonephritis

Strenuous exercise

Hemolytic anemia

Transfusion reactions 49

Blood

• Blood in urine indicates pathology

• Two forms found in urine

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– Intact RBC

– Hemolyzed RBC

Blood

• Positive with hemoglobin or myoglobin

• False Positive– Menstrual contamination

– Microbial peroxidesMicrobial peroxides

– Oxidizing agents (bleach)

• False Negative– Ascorbic acid

– High levels of protein

– High nitrite reduces strip reactivity51

Blood: Terms

• Hematuria

(intact RBC)

H l bi iAll will give a

iti bl d

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• Hemoglobinuria

(hemolyzed RBC)

• Myoglobinuria

(muscle protein)

positive blood reaction

pH

• Kidneys help regulate the acid-base balance of the body Detects systemic acid base disordersbody. Detects systemic acid-base disorders

• Normal = 4.5 to 8.0

• Acidic = acidosis, high protein diet, starvation, dehydration, or diarrhea

• Alkaline = alkalosis, UTI, vegetarian diet, vomiting or chronic renal failure

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pH

• Normal: ranges from 4.5 – 8.0

• First morning void: acidic

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• Physiologically impossible: <4.5

>8.0

1. Urine not handled properly

2. Old urine

3. Treatment induced

CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 10

pH

• Invalid test results due to:– Improper handling of urine sample

Contamination of urine vessel prior to

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– Contamination of urine vessel prior to collection

– ‘Run-over’ phenomenon (dipstick technique)

Protein

• Normal kidneys excrete little protein (<10 mg/dl) Proteinuria associated with early renal disease

• Normal = Negative (Albumin reacts)

• Transient: occurs with fever, exposure to heat or cold, emotional stress or pregnancy, exercise

• Pathological: membrane damage, disorders affecting tubular reabsorption

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Protein

• The protein that is found in urine comes from– Bloodstream– Urinary tract

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• Proteinuria is an indicator of early renal disease

• Proteinuria also caused by non-renal disease– Multiple myeloma

Renal Causes of Proteinuria

• Glomerular damage:– Most serious cause of proteinuria– Most common cause of proteinuria– Glomerulonephritis

N h ti S d (hi h t l l f t i )

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– Nephrotic Syndrome (highest levels of protein)

• Tubular dysfunction:– Reabsorption capability decreased– Toxin exposure, inherited disorder– Fanconi’s syndrome: heavy metal poisoning

Protein• Sensitivity: ~ 10-25 mg/dl

• Specificity: reacts primarily with albumin

• False Positive• False Positive– Highly buffered or alkaline urine >8.0– Alkaline drugs– Improper storage and handling– Contamination of detergents

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Protein• False Negative

– Dilute urine

– Presence of other proteins• Uromodulin (Tamm-Horsfall protein matrix in casts)

• Globulins

M l bi• Myoglobin

• Free light chains (Bence-Jones protein)

• Hemoglobin

– Exercise/transitory conditions

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CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 11

Urobilinogen

• Urobilinogen is formed in the intestine from bili bi b b t i ( t i t d i th f )bilirubin by bacteria (most is excreted in the feces) but some is reabsorbed back into the bloodstream where small amounts are excreted in the urine

• Normal = 0.2 to 1.0 mg/dl

• Abnormal = Hemolytic disease

Liver disease

Cannot determine absence of UBG 61 62

Nitrites

• Some gram negative bacteria reduce dietary nitrates to nitrites. The bacteria that cause urinary tract infections (UTI) are often nitrite producersinfections (UTI) are often nitrite producers

• Normal = Negative

• Abnormal = UTI

Cystitis (bladder infection)

Pyelonephritis (kidney infection)

Rapid screening test for UTI 63

Nitrite• False Positive

– Substances that mask reaction color

– Foods (beets); Drugs

– Improper specimen storage/handling

F l N ti• False Negative– Ascorbic acid– Bacteria cannot reduce nitrates– Bladder time not sufficient: need 4 hours– Low nitrate levels (lacks dietary nitrates)– Antibiotic inhibition of bacteria– Further reduction of nitrites to nitrogen

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Leukocytes

• The presence of leukocytes in the urine indicate a possible urinary tract infection Can detect intactpossible urinary tract infection. Can detect intact WBC and lysed WBC (granulocytes)

• Normal = Negative

• Abnormal = Urinary tract infection (UTI): cystitis, pyelonephritis, urethritis

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Leukocytes• False Positive

– Substances that induce color mask

– Vaginal contamination

• False Negative• False Negative– Not waiting the two minutes

– Lymphocytes present; are not detected

– Increased glucose & protein

– Strong oxidizing agents

– Drugs

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CLS 500 Application and Interpretation of Clinical Laboratory DataRoutine Urinalysis- Physical and Chemical Examination of Urine 12

Leukocytes

• Abnormal:– Bacterial infection:

Cystitis (bladder infection)

Pyelonephritis (kidney infection)

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Pyelonephritis (kidney infection)

Urethritis (infection/inflammation of urethra)

– Non-bacterial infection:

Yeast

Trichomonas

Leukocytes

• Sensitivity: 5-15 WBC/hpf

• False positive

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– Vaginal contamination

– Color masking

• False negative– Oxidizing agents (bleach)

– Lymphocytes (no granules)

Ascorbic Acid (Vitamin C)• Interferes with Reagent Strip reaction

• Causes false negative reactions for

Blood Nitrite

Bilirubin Glucose

BBNG: “Bad Boys No Good”

BGNB: “Bad Girls No Better”

University of Nebraska Medical Center

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