Urine Analysics Full Report -8371-M.Saihetharan.pdf

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KIU
Faculty Name: Health science
Department Name: Biomedical science
Degree Name: BSC (hons) in Biomedical Science
Individual Assignment
Assignment Title (Urine analysis full report)
Subject Name: Clinical Biochemistry 1
Subject Code: BMS 1407
Examination ID Number: KIU/ X/ 8371
Name of Student: M. SAI HETHARAN
Lecturer’s Name: Jayani Dilhani Kumara
Program Code: 0406
Deadline of the Assignment: 25 / 07/ 2022
Submission Date of the Assignment: 22 / 07/ 2022

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DECLARATION
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Acknowledgment
I had to seek out the assistance and direction of a few reputable people in order to complete my work, and I
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gratitude to KIU University for providing me with an excellent assignment guideline during my multiple
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Many people, notably my classmates, provided insightful comments and ideas on my work, which motivated
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Contents
1 Introduction
1.1 Urine
1.2 Urine Composition
1.3 Urine Volume
1.4 Oliguria
1.5 Polyuria
1.6 Anuria
1.7 Urine Collection
1.8 Specimen Collection
1.9 Type of Specimen
1.10 Urinalysis
1.11 Urine Preservation
2 Physical Examination
2.1 Color
2.2 Odor
2.3 Clarity
2.4 Specific Gravity
2.5 Urinometer
2.6 Refractometer
2.7 Harmonic Oscillation Densitometry
3 Chemical Examination
3.1 Determination of Reducing Substance
3.1.1 Benedict Test
3.2 Determination of Ketone Bodies in Urine
3.2.1 Rothera’s Test
3.2.2 Gerhardt’s Test
3.3 Determination of protein in Urine
3.3.1 Sulfosalicylic Acid Test
3.3.2 Acid Heat Coagulation
3.4 Determination of Bilirubin
3.4.1 Fochets Test
3.5 Determination of Urobilinogen
3.5.1 Ehrich’s Test
3.6 Reagent Strip Method

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4 Microscopic Examination
4.1 Microscope
4.2 Centrifugation
4.3 Microscopic Contents
5 Abnormal Report Analysis
6 References
List of Figure
Figure 1……………………………………. Oliguria
Figure 2……………………………………. Polyuria
Figure 3……………………………………. Anuria
Figure 4……………………………………. Specimen Sample
Figure 5……………………………………. Urine Color Chart
Figure 6……………………………………. Pink Urine
Figure 7……………………………………. Dark Orange
Figure 8……………………………………. Dark Urine
Figure 9………………………………….... Variety of Urine Colors
Figure 10…………………………………… Cloudy Urine
Figure 11…………………………………… Urinometer
Figure 12…………………………………… Refractometer
Figure 13…………………………………… Benedict Equation
Figure 14…………………………………… Benedict Results
Figure 15…………………………………… Rothera’s Test Results
Figure 16…………………………………. Refractometer
Figure 17…………………………………. Gerhardt’s Test
Figure 18…………………………………. Sulphosalicyclic Acid Test Results
Figure 19…………………………………. Fochets Reagent
Figure 20…………………………………. Fochets Results

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Figure 21…………………………………. Ehrich’s Results
Figure 22…………………………………. Reagent Strip
Figure 23…………………………………. Centrifuged Strip
Figure 24…………………………………. Microscope
Figure 25…………………………………. Centrifuged Conical Tube
Figure 26…………………………………. Centrifugation
Figure 27…………………………………. Sediment
Figure 28…………………………………. Uri Stain Sediment Test
Table of Content
Table No 1: Urine Color and Causes
Table No 2: Urine odor and Causes
Table No 3: Specific Gravity and Causes
Table No 4: Color of Precipitable

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1. Introduction
1.1 Urine
A fluid that includes several metabolic waste products, particularly urea and other nitrogenous chemicals,
which the kidneys filter from the blood. The urinary bladder stores urine, which is then expelled from the body
through the urethra.
1.2 Urine Composition
Organic components: Urea, Creatinine, Uric acid, Hippuric acid, other substances
Inorganic components: Sodium Chloride, Potassium, Sulfate, Ammonium, Magnesium, Calcium
1.3 Urine Volume
• Normal Volume is 750-2000ml/24h
• Under 750ml (oliguria)-dehydration, infection, obstruction, renal stones, kidney, failure.
• Over 2000ml (polyuria)-diabetes insipidus, hypertension, nephrotic syndrome, ingestion of alcohol or
Drugs, endocrinal disorders
• Absent of urine (anuria)-Obstruction, Kidney failure, stenosis
Factors that influence urine volume
• Fluid intake
• Fluid loss from non-renal sources
• Variations in the secretion of antidiuretic hormone
• Need to excrete increased amounts of dissolved Solids
• Pathological conditions
1.4 Oliguria
❖ Oliguria refers to infrequent urination. That translates to fewer than 400 milliliters of pee every day
for adults.
❖ The specific amounts for infants and children are based on their weight (less than 1 milliliter per
kilogram per hour for infants, and less than 0.5 milliliters per kilogram per hour for children).
Dehydration: This is the most common cause of Oliguria.it most happens When you are Vomiting or
hard diarrhea
Figure 1: Oliguria

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1.5 Polyuria
Its potential severity, numerous origins, and intriguing pathophysiology, polyuria is a significant symptom or
indicator. While polyuria brought on by water diuresis is well understood and simple for doctors to identify,
that brought on by solute diuresis is more likely to be perplexing.
It examines the etiology and classification of polyuria, concentrates on solute diuresis as a source of polyuria,
and describes clinical and laboratory tests helpful for assessing polyureic patients. In order to determine
whether a patient has a water diuresis, a solute diuresis, or both (concurrently), as well as to determine whether
a solute diuresis is caused by electrolytes (such as sodium chloride or sodium bicarbonate), by nonelectrolytes
(such as glucose or urea), or by both, a step-by-step, logical approach is provided
Figure 2: Polyuria
1.6 Anuria
• The cause of anuria is divides Causes in to prerenal, renal, and post renal
• Post renal implies cessation of urinary Secretion due to a fall in blood pressure below the level which
may be due to diminution of the circulatory blood volume or slowing of the circulation rate.
• The renal causes meant stoppage of secretion in the presence of adequate filtration pressure and
unobstructed ureters.
• In these causes tubular damage or blockage occurs. It’s probable that the glomerular filtrate is almost
all reabsorbed direct into the blood stream owing to the distraction of the cells lining the tubules.
• It is probable that the glomerular filtrate is almost all reabsorbed direct into the blood stream owing
to the destruction of the cells lining the tubules.

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Figure 3: anuria
1.7 Urine Collection
• Urine must be collected in sterile bottle
• The routine urine examination must be conducted within 30 minutes
• In case of delay, urine must be refrigerated
• Delay in sample examination will result in:
✓ Decreased pH by the utilization of glucose by bacteria
✓ Increased pH by the conversion of urea to ammonia by bacteria
✓ Increased pH the tendency of phosphate to precipitate will increase
✓ Oxidation of urobilinogen to urobilin. It may give false negative
Figure 4: Specimen Sample

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1.8 Specimen Collection procedure
Patient preparation
Female patient
1. First thoroughly wash your hands
2. In the perineal region, use beta line swabs or hibiclens
3. Unfold the labia and wipe one side with betadine swabs or Hibiclens from front to back.
4. Hibiclene or the use of a second betaline swab
5. From front to back, wipe down the middle and toss.
6. Wipe the middle of the object from front to back with a third betaline swab or Hibiclens wipe, then
throw it away.
7. Use clean, dry gauze to dry the periurethral area and remove any extra beta line while maintaining the
labia apart.
For male patient
1. Hibiclene or the use of a second betaline swab,
2. From front to back, wipe down the middle and toss.
3. Wipe the middle of the object from front to back with a third betaline swab or Hibiclens wipe, then
throw it away.
4. Use clean, dry gauze to dry the periurethral area and remove any extra beta line while maintaining the
labia apart.
A. Urination should begin passing the first portion into the bedpan urinal or toilet
B. After the flow of urine has started the urine specimen container should be placed under
the patient collecting the midportion without contamination the container.
C. Any excess urine can pass into the bed pan urinal or toilet
D. Cover the urine container immediately with the lid being careful not to touch the inside
of the container or the inside of lid
E. Transfer the to specimen tube or container and place specimen in the transport of urine
container
F. Attach label to tube or container and place specimen in the transport bag.
G. Remove gloves and wash hands
H. Record date the time of collection and initials of the person collecting specimen on the
specimen container transport to the lab as soon as possible hours of collection or
refrigerate and transport lab to soon as possible

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1.9 Types of urine specimen
1. Random specimen
Chemical and microscopic examination, a voided specimen is usually more suitable. A randomly collected
specimen may be collected at unspecified times and is often more convenient for the patient. A random
specimen is suitable for more screening purposes
2. First morning specimen or 8-hour specimen
The patient should be instructed to collect the specimen immediately upon rising from night sleep. Other 8-
hours periods may be used accommodate insomniacs, night-shift workers, and in certain pediatric situations.
The bladder is emptied before lying down and the specimen is collected on arising so that the collected only
reflects the recumbent position. Any urine voided during the night should be collected and pooled with first
morning voided specimen.
3. Fasting specimen
This differs from a first morning specimen by being the second voided specimen after a period of fasting.
4. 2-hour post prandial specimen
The patient should be instructed to void shortly before consuming a routine meal and to collect a specimen 2
hours after eating.
5. 24-hour or timed specimen
To obtain an accurately timed specimen.it is necessary to begin and end the collection period with an empty
bladder. The following instructions for collecting a 24-hours specimen can be applied to any timed collection.
Day 1-7 AM patient voids and discards specimen. Patient collects all urine for the next 24 hours.
Day 2-7 AM patient voids and adds this urine to the previously collected urine.
6. Catheterized specimen
This specimen is collected under the sterile conditions by passing a hollow tube through the urethra into
bladder.
7. Mid-stream “clean catch” specimen
This specimen provides a safer less traumatic method for obtaining urine for bacterial culture. It also offers a
more representative and less contaminated specimen for microscopic analysis than the random specimen.
Adequate cleansing materials and a sterile container must be provided for the patient. The procedure for the
collection of a clean catch urine is described below in section VI of the policy.

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8. Suprapubic aspiration
Urine may be collected by external introduction of a needle into the bladder. It is free of extraneous
contamination and may be used cytologic examination.
9. Pediatric specimens
This may be a sterile specimen obtained by catheterization or by suprapubic aspiration. The random specimen
may be collected by attaching a soft. Clear plastic bag with adhesive to the general area of both boys and girls.
1.10 Urinalysis
It is a routine urine examination.it is most useful for clinicians as an indicator or health or disease.it particularly
used in renal metabolic disorders. Often it done for patients admitted to the hospital. The diagnostic
examination for all urinary and many metabolic illnesses must include a urinalysis. Its assessment includes
evaluation of physical characteristics (color, clarity, and volume). The parameters are influenced by collection
method and therefore with accuracy of some test results can decrease improper interpretation. When all these
parameters are evaluated in combination with clinical signs. Physical examination through history and other
laboratory tests, a diagnosis will often be attained.
1.11 Urine preservation
The routinely used method preservation 2o
C to 8o
c of urine sample and refrigeration should be increased
Specific gravity and precipitation of amorphous phosphates and urates.
Preservative Methods
1. Hydrochloric Acid: It used for detect adrenaline or nor-adrenaline, Vanilylamandelic acid (VMA),and
Steroids
2. Toluene: It forms a thin layer and hence physical Barrier against Bacteria and air.
3. Boric acid: Specimen sample should be Keep for 24 hours without refrigeration.
4. Thymol: It can inhibit bacteria and fungi.
5. Formalin: Excellent for preservation for formed elements.
It is not recommended for routine urine analysis because they interfere with reagent strip techniques and
chemical test for protein.
Physical Examination
This includes color, odor, Appearance, Specific gravity.
Chemical Examination
The routine urine analysis includes Ph, Protein, glucose, ketones, occult blood, bilirubin, urobilinogen, nitrite,
leukocyte esterase and strip test method for specific gravity.
Microscopic Examination
This includes RBC, WBC, epithelial cells, Casts, bacteria, Yeast, Parastesmatozoa, crystals, artifacts.

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2. Physical Examination
It includes color, odor, clarity, appearance, specific gravity.
2.1 Color
The normal color in fresh state is pale yellow or amber and is due to the presence of various pigments
collectively called urochrome.
The color of urine varies from almost colorless to black.
The most changes in urine color are harmless and temporary and may be due to certain foods, Dyes in foods,
supplements and prescription drugs. Unusual urine color can indicate an infection or serious illness.
The amount of water you consume affects the color of your normal urine. The yellow pigments in urine are
diluted by fluids, thus the more you drink, the clearer your pee appears; the less you drink, the more
concentrated the color is. Urine that is amber in hue can result from severe dehydration. However, urine can
change colors that are far from normal, such as red, blue, dark brown, and hazy white.
Urine Color Chart
Figure 5: Urine color Chart
Urine color and conditions
Colors Conditions
Colorless Dilute urine (diabetes mellitus, diabetes insipidus,
over hydration)
Red Hematuria,
Hemoglobinuria
, Porphyria,

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Myoglobinuria
Dark brown or black Alkaptonuria, Melanoma
Brown Hemoglobinuria
Yellow Concentrated Urine
Yellow green or green Biliverdin
Deep yellow with yellow foam Bilirubin
Orange or Orange brown Urobilinogen
Porphobilinogen
Milky-White Chyluria
Red or Orange fluorescence with Uv light Porphyria
Table No :1 Urine Color and Conditions
Urine color Specimen Examples
Figure No 8: Dark urine
Figure:7 Dark Orange
Figure:6 Pink Urine
Figure 9:Variety of urine Colors
2.2 Odor
Urine typically does not have strong odor, but odor is infected by urine concentration, diet medications the
presence of urine concentration, diet, medications, the presence of bacteria, inflammation and certain medical
conditions (E.g.; in diabetes mellitus a sweet or fruity urine odor is sometimes detected).
Improperly stored or aged samples may have strong odor. Some benign, non-harm full circulenes such
consuming specific foods or using specific medications may cause an odd or strong urine odor.
Dark
Orange
Dark
urine
pink
urine

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In some causes an unusual or storing urine odor may be due to benign conditions that are not harmful, such as
eating certain foot or taking certain medications.
Urine Odor Causes
Aromatic Normal
Foul, ammonia-like Bacterial decomposition, urinary tract infection
Fruity , sweet Ketones (diabetes mellitus ,starvation, vomiting)
Maple syrup Maple syrup urine disease
Mousy Phenylketonuria
Rancid Tyrosinemia
Sweaty feet Isovaleric acidemia
Cabbage Methionine malabsorption
Bleach Contamination
Table No 2 Urine Odor and Causes
2.3 Clarity
The physical property of turbidity refers to the degree of the clarity of the urine generally and is reported in
range from clear to flocent.
Normal urine described usually as clear to very mildly cloudy. Turbidly most offer co relates to the amount
of particulate matter present in the urine in the microorganisms and mucus.
Occasionally semen and feces can also contribute to turbidity the causes turbidity is usually determined by
microscopic examination.
Figure 10: Cloudy Urine
2.4 Specific gravity
Specific gravity is defined as the density of a solution compared with the density of similar volume distilled
water at the similar temperature. Urine specific gravity measured with the refractometer is an indirect
measurement of urine osmolality. The interpretation if USA is dependent on several factors such as hydration
status, electrolyte concentration tractions, serum creatinine and urea nitrogen. Concentrations, the
administration of certain medications and the administration of fluid therapy.
Cloudy urine

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Specific gravity Causes
USG less than 1.007 Hyposthenurias-this dilute urine indicating specific
gravity lower than that the plasma and glomerular
filtrate
USG between 1.008 and 1.013 Isothenuria- It equal to that of plasma and glomerular
filtrate
USG less than 1.025 Ploydipsia – Water intake greater than 50ml/Kg/day
Table No :3
2.5 Urinometer
A weighted float is connected to a scale that has been calibrated in terms of urine specific gravity. The
urinometer is composed of three elements. They are as follows: I. The float is the air-containing portion II.
Weight is the bottom end of the urinometer. Stem- Equipped with calibrations and numbers to indicate the
specific gravity.
Figure 11: Urinometer
Principle of urinometer: It is based on the principle of BUOYANCY. Because of increased density of urine
compared to that of water, the urinometer will float higher in urine than in water. The weighted float will
displace the volume of the urine which is equal to its weight. The weight of the weighted float is designed in
such a way that the urinometer sinks to the level of 1.000 in the distilled water.
Procedure of using urinometer: 1. An adequate amount of urine is poured into a proper-size container.
2. Urinometer is added with a spinning motion.
3. The scale reading is then taken at the bottom of the urine meniscus
Temperature adjustment made while calculating specific gravity: Due to the fact that urinometers are
calibrated at 15 degrees Celsius, a temperature correction must be applied in order to measure specific gravity
correctly. To do this, multiply the temperature by 0.001 for every 3 degrees above or below 15 degrees Celsius,
respectively

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Normal value of specific gravity:1.003-1.030
Disadvantages of Urinometer: At least 15ml of urine required and if the urine very turbid then measurement
will be very difficult.
2.6 Refractometer
Simply evaluating the refractive index, one could determine the sample's concentration of dissolved particles.
Benefits include the need for no temperature correction and the need for only one or two drops of urine.
Figure 12: Refractometer
2.7 Harmonic oscillation densitometry
The principle is – principle that the frequency of a sound wave entering a solution change in proportion to the
density of the solution. Advantages are that temperature corrections are not necessary.

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3. Chemical Examination
TheroutineurinalysicsincludeschemicaltestingforPh,Protein,glucose,ketones,occultblood,bilirubin,urobilinog
en nitrite, leukocyte esterase and strip test method for specific gravity. The urine analysis offered by
laboratories depends on type of dipstick that is used. The urinalysis offered by medical laboratories depends
on the type of dipstick that is used. These procedures are either qualitative (positive or negative) or semi-
quantitative measurements. Since the introduction of single and multiple-test reagent strips, test tapes, and
tablets, the chemical screening of the urine have become a sensitive and rapid procedure.
Abnormal constituents in urine includes:
1. Reducing substances (glucose, fructose, galactose, vitamin C)
2. Proteins (albumin, globulin, hemoglobin, Bence jones protein)
3. Ketone bodies (Acetone, acetoacetic acid)
4. Bile salts and bilirubin (bile pigments)
5. Urobilinogen (increased amount)
3.1 Determination of Reducing Substance
3.1.1 Benedict Test
Introduction
A chemical test called Benedict's test can be used to determine whether an analyte contains reducing sugars.
This test can therefore be used to identify simple carbohydrates that include a free ketone or aldehyde
functional group. The test is based on a compound mixture of sodium citrate, sodium carbonate, and the
pentahydrate of copper (II) sulfate known as Benedict's reagent, also referred to as Benedict's solution. A
brick-red precipitate that forms as a result of Benedict's reagent's interactions with reducing sugars is a sign
that the test is successful. The illustration below shows how exposure to reducing sugars causes Benedict's
reagent to change from clear blue to brick-red in color.
It should be mentioned that the Benedict's test can be utilized to determine whether there is glucose present in
a urine sample. Since glucose is an aldose whose open-chain forms an aldehyde group and this test detects
any aldehydes and -hydroxy ketones, the presence of glucose in the analyte results in a positive test result.
However, the presence of homogentisic acid, ascorbic acid, and other reducing agents in urine can also result
in a favorable response. As a result, a positive Benedict's test result does not automatically indicate that the
test participant has diabetes.
List some substances give positive results of Benedict test:
✓ Glucose
✓ Fructose
✓ Ribose

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Principle of Benedict Test
Figure 12: Benedic test
A reducing sugar has become an enediol when heated while still in the presence of an alkali (which is a
relatively powerful reducing agent). Benedict's reagent's cupric ions (Cu2+) are converted to cuprous ions
(Cu+) when reducing sugars are present in the analyte. These cuprous ions combine with the reaction mixture
to produce copper(I) oxide, that precipitates as a brick-red substance.
Procedure of Benedict Test:
1. 1. Pipette 5 ml of Benedict’s reagent into a test tube
2. Add 8 drops of urine to the Benedict’s reagent
3. Keep for 5 minutes in a boiling water bath.
4. Cool under running tap water
5. Observe the color change with precipitation
Figure No :14
Color of precipitate Reducing sugar
Green 0.5%
Yellow 1%
Orange 1.5%
Red 2%
Table No 4
Limitations of Benedict test:
1. False-positive reactions in the test can also be obtained if there are certain drugs present for example,
salicylates, isoniazid, streptomycin, penicillin, and p-amino salicylic acid.
2. The chemicals present in the concentrated urine may reduce Benedict’s reaction which includes urate,
creatinine, and ascorbic acid (the reduction is slight).

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3.2 Determination of Ketone bodies in urine
3.2.1 Rothera’s Test
Introduction:
Acetone (2%), acetoacetic acid (20%), and beta-hydroxybutyrate are the three intermediate products of fat
metabolism that are referred to as ketone bodies (78 percent). Ketone bodies are minutely eliminated in urine
(1 mg/24 hours) because digested lipids are entirely converted to water and carbon dioxide under normal
conditions.
Procedure:
1. Take 5 ml of urine in a test tube and saturate it with ammonium sulphate.
2. Add a small crystal of sodium nitroprusside.
3. Mix well.
4. Slowly run along the side of the test tube liquor ammonia to form a layer.
5. Observe the pink-purple ring at the interface
Figure No :15 Rothera’s test Results
Results and Observation:
• Immediate formation of purple permanganate colored ring at the interface
• Ketone bodies present (Positive)
• No formation of purple permanganate colored ring at the interface:
• ketone bodies absent (Negative)

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F i g u r e N o : 1 6 Example of portable ketonometer/ glucometer which can be used for evaluation of
diabetic patients.
3.1.2 Gerhardt’s test
Introduction:
Gerhardt’s test is based on the reaction of ferric chloride with acetoacetic acid to form a port wine or Bordeaux
red color. This test detects acetoacetic acid only, Acetone and beta-hydroxybutyrate can’t be detected by this
method. It also detects salicylates in urine.
Procedure:
1. Take 3 ml urine into a test tube.
2. Add 3 % FeCl3 solution drop by drop. Mix while adding.
3. Filter the brown- red precipitate formed.
4. Add 3 % FeCl3 solution continuously to the filtrate until a purple color appear
5. Heat and see whether the color disappear or not
Observation and results:
• If the color disappears, then acetoacetic acid is present. The acetoacetic acid on boiling loses carbon
dioxide and is converted to acetone. Acetone doesn’t react with ferric chloride.
• If the color persists, acetoacetic acid is absent. Previous color is due to salicylates.

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Figure No:17 Gerhardt’ s Test
Clinical significance of ketonuria

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3.3 Determination of protein in urine
3.3.1 3%Sulfosalicyclic acid method (SSA)
Introduction:
In the presence of 5-sulphosalicylic acid, proteins precipitate. Any turbidity that occurs will provide an
approximate estimate of the amount of protein present in the urine, which can be visually quantitated
qualitatively or more precisely using photometry. Before performing the test, the urine must be centrifuged to
remove casts and cells.
The test can detect albumin, hemoglobin, myoglobin, and Bence Jones proteins.
Procedure:
1. Add 3 ml of urine into a two-test tube
2. Label test tube as “test” and “negative control”
3. Add 3 ml of 3% sulfosalicylic acid to the test tube
4. Add 3 ml of water for control tube
5. Mix well
6. Immediately compare the test and Control by observation for turbidit
Results and interpretation:
Figure No:18
Negative: No cloudiness
1+ : definite turbidity
2+ : Heavy turbidity but no flocculation
3+ : Heavy turbidity with light flocculation
4+ ; Heavy turbidity with heavy flocculation

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Special interpretation:
This test, protein cannot be found in normal urine. In the case that the patient is using tolbutamide, penicillin,
or even other medications, a false positive result could be generated. Due to precipitation of urate in an acidic
urine, a high concentration of urates in the urine may result in a false positive test result.
Significance figures of proteinuria:
• Less than 150 mg of protein per day is considered normal in urine.
• Rapid deterioration in kidney function is linked to high protein levels in the urine.
• About 6.7 percent of Americans are impacted by it.
• More elderly folks and people with various chronic conditions are affected by it.
Causes of proteinuria:
• In many cases, proteinuria is caused by relatively benign (non-cancerous) or temporary medical
conditions.
• These include dehydration, inflammation and low blood pressure. Intense exercise or activity,
emotional stress, aspirin therapy and exposure to cold can also trigger proteinuria. In addition, kidney
stone in the urinary tract can cause proteinuria.
• Occasionally, proteinuria is an early indication of chronic kidney diseases, a gradual loss of kidney
function that may eventually require dialysis or a kidney transplant.
3.3.2 Acid heat coagulation test (premilitary step for protein)
Introduction:
Proteins like albumin and globulin can be detected in protein using a biochemical test called the heat
coagulation test. It is a typical occurrence for proteins to coagulate in reaction to heat. Denaturation and
agglutination or the separation of the denatured protein in a particular form are the two processes in which
heat coagulation of proteins takes place. When coagulable proteins are heated to their isoelectric pH, a few
changes take place to the proteins. These changes include the dissociation of subunits or the quaternary
structure, the uncoiling of the polypeptide chains, and eventually the matting together of uncoiling polypeptide
chains. Proteins undergo modifications throughout the denaturation process, which are superficially
observable in processes like coagulation and flocculation.
Proteins coagulate when their isoelectric pH is reached, which is an irreversible process. An essential clinical
test for identifying proteinuria is the heat coagulation of proteins. It takes less time and is easy. The heat
coagulation method can be used to estimate proteins both qualitatively and quantitatively. The coagulum that
has formed on the test tube can be measured to perform a quantitative examination of coagulation.
Procedure:
1. Fill 2/3rd of the test tube with urine
2. Add few drops of dilute acetic acid
3. . Heat the top half of it
4. Lower part serves as control
5. Note the appearance of the turbidity

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Results and interpretation
Figure 18 : Sulphosalicyclic Acid Test Results
Positive result: A positive result of the heat coagulation test is represented by the formation of a dense
coagulum at the upper part of the solution. The lower part of the solution acts as a control.
Negative result: A negative result of the heat coagulation test is represented by the absence of coagulum at
the upper layer. This indicates the absence of albumin and other proteins in urine.
3.4 Determination bilirubin (bile pigments in urine)
Introduction
When red blood cells degrade, a brownish-yellow pigment called bilirubin is produced. The chemical, which
is in the liver, is typically eliminated from the body during digestion. Everybody's bodies contain bilirubin
since it is a byproduct of the natural process of red blood cells degenerating and being replaced by new ones.
Both age and general health have an impact on bilirubin levels. Babies who have bilirubin levels more than
20 to 25 milligrams per deciliter risk brain damage. Because of this, it's essential to measure their bilirubin
levels. One technique to determine how much bilirubin is in your system is through a urine test.
3.4.1 Fochets Test
Two different tests are available to check for bilirubin in urine: (A) the oxidation test, in which bilirubin is
converted to green biliverdin, and (B) the diazotization test, in which bilirubin is converted to a highly colored
molecule. The reagent employed in the Fouchet test falls within the category of oxidizing reagent or test. The
sulphate radicals in urine are precipitated by barium chloride to create barium sulphate precipitate. If there are
bile pigments in the urine, they stick to these molecules.

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Principle
The reagent employed in the Fouchet test falls within the category of oxidizing reagent or test. Barium chloride
used to precipitate sulphates in urine. The sulfate radicals in urine are precipitated by ferric chloride to create
barium sulphate precipitate. If there’re bile pigments in the urine, they stick to those molecules. The
development of green color as a result of bilirubin synthesis thus denotes bilirubin (bile)in urine.
Figure No:19 Fochets Reagent
Procedure:
1. Check the pH of urine.
2. If alkaline, acidify with diluted acetic acid.
3. Add 5 ml of 10% BaCl2 solution to 10 ml urine in a test tube.
4. Mix and observe for the presence of a precipitate
5. Filter through a Whatman No:1 filter paper.
6. Dry filter paper over placing it on a second filter paper
7. Add a drop of fouchet’s reagents to the filtered precipitate
8. Observe a color
Quality control
Appearance: Yellow colored solution
Clarity: Clear without Precipitate
Results
Positive: Green color
Fochets
reagent

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Figure No 20 Fochets Results
Bilirubinuria
Bilirubin available in two main forms: conjugated bilirubin, which is common in cholestatic jaundice, and
unconjugated bilirubin, which is common in hematologic jaundice. Unconjugated bilirubin is not water soluble
and is not excreted in urine, in comparison to conjugated bilirubin, which would be both.
Homeopathic repertories published prior to the discovery of the two types of bilirubin in 1913 present an
opportunity to test the reliability of homeopathic repertories and associated Materia medica. If procedures
involved in the collecting of homeopathic observations are reliable, then in repertories published prior to 1913,
medicines listed for cholestatic jaundice should exhibit a stronger association with urine bile than medicines
listed for hematologic jaundice.
3.5 Detection of urobilinogen
A urine test detects urobilinogen quantifies the level of urobilinogen in human urine Urobilinogen is found
in some common urine. A liver condition like cirrhosis or hepatitis, as well as some anemias, may also be
identified by an excessive amount of urobilinogen in the urine.
3.5.1 Ehrlich’s test
Introduction
A biochemical test called the Ehrlich Test is used to find the amino acid tryptophan in a protein sample. Ehrlich
reagent is the source of the Ehrlich test, also known as the p-dimethylamine benzaldehyde test. It is also called
a specific amino acid test as it detects a certain amino acid, tryptophan.
This test has been used in medical testing labs, some for-drug testing and others for disease diagnosis. The
Ehrich test can also be used to diagnose liver abnormalities, carcinoid syndrome, hemolytic processes, and
obstruction of the common bile duct. The spot test for the Ehrlich test, which is commonly used to identify
psychoactive substances like tryptamine and ergoloids, is one of its variants.
A positive Ehrlich test is obtained for natural opium as it contains tryptophan. The Ehrlich reagent is also used
for the identification of indoles and urobilinogen.
Principle:
Ehrlich reagent consists of p-dimethylamine benzaldehyde. The reaction occurring in the test is based on the
principle that under acidic conditions, the Ehrlich’s reagent undergoes electrophilic substitution.

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The substitution occurs at the indole or the benzyl pyrrole ring of tryptophan to yield a blue-violet
condensation product. The condensation product formed after the reaction is further enhanced by the addition
of NaNO2.
Procedure
1. Add 0.5 ml Ehrlich’s reagent to freshly voided 2 ml urine.
2. Mix the contents and leave for 3-5 min at room temperature.
3. If no red color is produced warm the contents to 50o
C.
Reaction:
Tryptophan + Ehrlich reagent (p-dimethylamine benzaldehyde) → Blue-violet condensation product
Results:
Figure 21:Ehrlichs Test results
Positive result:
A positive result in the Ehrlich test is indicated by the appearance of red to purple or blue-violet color.
The color then changes to blue with the addition of NaNO2. This indicates that the sample contains tryptophan.
Negative result:
A negative result in the Ehrlich test is indicated by the absence of blue-violet color on the addition of the
Ehrlich reagent.
This indicates that the sample doesn’t contain any tryptophan.
3.4 Reagent strip method:
A reagent strip, a dipstick, is a narrow plastic strip with small pads attached. Each pad contains reagents for a
different reaction, thus allowing for the simultaneous determination of several tests. The colors generated on
each reagent pad vary according to the concentration of the analyte present. Colors generated by each pad are
visually compared against a range of colors on brand-specific color charts.
The complete reagent strip must be dipped into to the urine specimen and taken out in one motion while the
excess urine is wiped away by running the strip down the edge of a specimen container. This is how a reagent
strip is tested for urine manually. The activities must be read at the designated time after dipping and then

28 | P a g e
thoroughly compared with the color chart supplied by the manufacturer. Certain measures must be taken to
maintain the chemicals' reactivity in order to produce precise and trustworthy results using the dipsticks. The
strips should be kept in the original containers and must not be exposed to dampness, bright sunlight, heat, or
combustible substances. The container shouldn't be placed in the refrigerator or left out in the sun for longer
than 30o
c
Figure 22 : Reagent Strip

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4. Microscopic Examination
Microscopic Examination includes RBC, WBC, Epithelial cells, casts, Bacteria, Yeast, parasites, mucus,
spermatozoa, crystals, artifacts.
Sample for analytics
• Freshly voided urine is the best sample.
• If delayed, then refrigerate the urine.
• The best volume for the centrifuge is 10 to 12 mL.
Figure 23: Centrifuged Strip
4.1 Microscope
Figure No 24

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Handling procedure of microscope:
Microscopes are devices used to magnify small objects, allowing them to be seen by the naked eye. Most
microscopes have several different powerful lenses attached to them, allowing the viewer to inspect the
content at more than 100 times its actual size. However, microscopes are extremely expensive, so you want
to make sure you handle the device properly.
Hold the microscope with one hand around the arm of the device, and the other hand under the base. This
is the most secure way to hold and walk with the microscope.
Avoid touching the lenses of the microscope. The oil and dirt on your fingers can scratch the glass.
Clean smudges on the microscope glass with a piece of lens paper. Any other material, such as tissues or
even cotton, can scratch the glass of the microscope.
Rotate the nosepiece of the microscope all the way down to its lowest level when you have finished using
the microscope. If you leave the nosepiece up while storing it, the gears on the device can wear out. If this
happens, the nosepiece may not be able to lift and hold its position.
Cover the microscope with the dust cover to protect it from dirt and other debris.
Preparation and Examination of urine Sediment
Fresh or well preserved
It Transferred into the conical centrifuged tube
Figure No 25
4.2 Centrifugation
Centrifugation for 5 minutes at a relative centrifuged force of 400 forces an optimum amount of sediment.
The centrifuge has the least chance of damaging the elements at 1500 rpm for 5 minutes.
Centrifuged
conical tube

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Figure 26:Centrifugation
Sediment preparation
Steps in preparation of concentrated urine Sediment
Mix urine specimen well and transfer a standardized volume into a conical centrifuge tube. Use the volume
that is stated in your laboratory's procedure.
Centrifuge at the speed and spin time specified in your laboratory's procedure.
Decant the sample to a standard volume (e.g., 0.5 - 1.0 mL) and resuspend the sediment in this volume.
Pipette a specified amount of sediment into the slide and put the cover slip in place.
Figure No 27 Sediment

32 | P a g e
Examination of sediment
• Brightfield microscopy and muted lighting are typically used to investigate the sediment (some
structures will be missed if there is too much light in the field). Throughout the examination, maintain
fine focus to spot structures in various focal planes.
• Brightfield microscopy and muted lighting are typically used to investigate the sediment (some
structures will be missed if there is too much light in the field). Throughout the examination, maintain
fine focus to spot structures in various focal planes.
• Scan the slide on low power (100X total magnification) for quantification of casts, crystals and
elements that are present in only a few fields.
• Use high power (400X total magnification) to identify casts and count red blood cells (RBCs), white
blood cells (WBCs), and epithelial cells.
• Staining may be helpful in the identification of some cells and casts. Supravital stains that may be used
include Sternheimer-Malbin and 0.5% toluidine blue.
• If available, phase-contrast microscopy enhances the identification of some sediment elements,
particularly those that have a very low refractive index and may be missed using brightfield
microscopy (e.g., some casts).
• Polarized light enhances the identification of lipids and some crystals.
Figure No 28
Uri Stain
sediment test

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4.3 Microscopic contents
Red Blood cells
RBC-Appear as smooth non-nucleated, bio concave disks
Identified using high power (X40)
Reported in average number seen in 10 hpfs
The presence of RBC called Hematuria.
Differentiate in RBC
• Both air bubbles and oil droplets have a high refractivity.
• Compared to the other sediments, these can be seen at a lower level.
• RBCs are more compact than WBCs.
• Lyse the RBCs by adding acetic acid to the sediment; yeast, oil droplets, and air bubbles are left behind.
• Alternately, use supravital staining
Microscopic observations
RBC-normal appearance
RBC-Crenated RBC in concentrated urine
RBC-Dysmorphic

34 | P a g e
White blood cells
• The average diameter of WBCs, which are larger than RBCs, is 12 m. WBCs are simpler to identify
in the urine sediment than RBCs.
• Granules and multilobate nuclei are features of neutrophils' cytoplasm.
• High power imaging allows for the detection of WBCs, which are reported as the average number
detected in 10 HPF.An increased number of WBCs in the urine is called Pyuria.
Pyuria
❖ Bacterial infection of the urinary tract
❖ Acute pyelonephritis shows casts along with WBCs, protein, and bacteria.
❖ Lower urinary tract infection also shows WBCs but a small amount of protein.
Microscopic Observations
WBC Cells
Abnormal result of Urine analysis under 40X

35 | P a g e
Epithelial cells
• Mainly these cells are shed from the urethra and urinary bladder.
• Few renal epithelial cells are seen normally in the sediment.
• Epithelial cells are the following types:
1. Squamous epithelial cells
2. Transitional epithelial cells
3. Renal epithelial cells.
Squama’s Epithelial cells
Transitional

36 | P a g e
Microscopic Examination of urine deposit
Reasons of identification of urinary crystals
1. liver disease
2. inborn errors of metabolism
3. renal damage.
Amorphous urates
✓ Amorphous crystals lack any recognizable distinctive shape. Amorphous urate crystals are those seen
at an acidic pH (less than 6).
✓ They transform into amorphous phosphate crystals at a pH higher than 7.
✓ They become quite difficult to discriminate between pH 6 and pH 7.
Uric acid crystals

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✓ There are numerous distinct compounds in urine. These substances have the potential to crystallize
into salt under certain conditions. It is known as crystalluria.
✓ Healthy people can have crystals in their urine. Minor problems like a slight overabundance of protein
or vitamin C may be the root of them. Urine crystals come in a wide variety and are generally safe.
Calcium oxalate crystals
✓ There are numerous distinct compounds in urine. These substances have the potential to crystallize
into salt under certain conditions. It is known as crystalluria.
✓ Healthy people can have crystals in their urine. Minor problems like a slight overabundance of protein
or vitamin C may be the root of them. Urine crystals come in a wide variety and are generally sa
Monohydrate calcium oxalate crystals
Normal crystals seen in Alkaline urine

38 | P a g e
Amorphous phosphates
Triple phosphate crystals
Ammonium pyruvate crystals
Abnormal urine crystals

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Crystine crystals

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5.Abnormal report analysis
1. Report Number 1
The patient urine appearance is turbid, so the patient has infections and inflammations and other diseases that
affect the functioning of the urinary tract, according to this report, the normal values range from pH 4.6 to 8.0
therefore in this report pH is 6 which means that the urine pH level is normal. The usual range of specific
gravity is 1.001-1. 035.therefore the specific gravity is also in a normal range also protein is also standard.
But the glucose level is trace which shows the patient has diabetes early stage. The ketones bodies are (+) so
it's a normal range. In microscopic examination shows the patient has 02-04/hpf. A Normal result is 4 red
blood cells per high power field (RBC/HPF) or less when the sample is examined under a microscope. So, the
patient has a normal range of red blood cells.

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Report No 2
The patient urine appearance is cloudy so patient can occasionally occur as a result od mid hydration.
According to this report, the normal values range from pH 4.6 to 8.0 therefore in this report pH is 6 which
means that the urine pH level is normal. The usual range of specific gravity is 1.001-1. 035.therefore the
specific gravity is also in a normal range and protein , glucose, ketone bodies, bile pigments also normal
range in patients urine.

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Report No 3
The patient urine appearance is Slightly turbid, in most cases, cloudy urine is harmless due to natural changes
that your body goes through. And the patients. the patient urine color is pale yellow so that category of color
signals the patient is healthy and hydrated and specific gravity and pH are also in the normal range. Protein,
glucose, ketone bodies, and bile pigments are also within the normal range in the patient's
urine. In microscopic examination shows the patient has approximately 200/ HPF pus cells.
The normal range of pus cells in the urine is 0-5, but the patient has a greater than normal range, So I predict
the patient has a urinary tract infection The normal range of epithelial cells is one to five squamous epithelial
field HPF in urine, so the patient has a normal range of epithelial cells.

44 | P a g e
The patient urine color is pale yellow, that category of color signals the patient is healthy and hydrated, and
specific gravity and pH are also normal range. Protein, glucose, ketone bodies, and bile pigments are also
normal range in patients' urine. The patient urine appearance is Slightly turbid, in most cases, cloudy urine is
harmless due to natural changes that your body goes through. and the patients. In microscopic examination
shows the patient has 02-04/HPF. A Normal result is 4 red blood cells per high power field (RBC/HPF) or
less when the sample is examined under a microscope. So, the patient has a normal range of red blood cells.
Report No 5

45 | P a g e
The patient urine color is a pale yellow, that category of color signals the patient is healthy and hydrated, and
specific gravity and pH are also normal range. Protein, glucose, ketone bodies, and bile pigments are also
normal range in patients' urine. The patient urine appearance is Slightly turbid, in most cases, cloudy urine is
harmless due to natural changes that your body goes through. And the patients. In microscopic examination
shows the patient has approximately 6 to 8/ HPF pus cells. The normal range of pus cells in the urine is 0-5,
but the patient has a greater than normal range, So I predict the patient has a urinary tract infection. the patient
has a normal range of red blood cells

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6.References
1. Urine collection procedure
“Specimen Collection Procedures: Urology » Pathology Laboratories » College of Medicine
» University of Florida.” Specimen Collection Procedures: Urology » Pathology Laboratories
» College of Medicine » University of Florida, pathlabs.ufl.edu,
https://pathlabs.ufl.edu/client-services/specimen-shipping/specimen-collection-procedures-urology/.
Accessed 20 July 2022.
2. Specimen
Wilson, Michael L. “General Principles of Specimen Collection and Transport | Clinical Infectious Diseases
| Oxford Academic.” OUP Academic, academic.oup.com, 1 May 1996, https://academic.oup.com/cid/article-
abstract/22/5/766/360981.
3. Types of urine
“Specimen Collection Procedures: Urology » Pathology Laboratories » College of Medicine
» University of Florida.” Specimen Collection Procedures: Urology » Pathology Laboratories
» College of Medicine » University of Florida, pathlabs.ufl.edu,
https://pathlabs.ufl.edu/client-services/specimen-shipping/specimen-collection-procedures-urology/.
4. Urine analysis
https://uomustansiriyah.edu.iq/media/lectures/6/6_2021_10_14!12_44_08_AM.pdf
5. Urine color
“Urine Color - Symptoms and Causes.” Mayo Clinic, www.mayoclinic.org, 24 Oct. 2020,
https://www.mayoclinic.org/diseases-conditions/urine-color/symptoms-causes/syc-
20367333#:~:text=Normal%20urine%20color%20ranges%20from,likely%20to%20affect%20t.
6. Urine odor
“Smelly Urine: Causes, Other Symptoms, Treatment, and More.” Smelly Urine: Causes, Other Symptoms,
Treatment, and More, www.medicalnewstoday.com, https://www.medicalnewstoday.com/articles/smelly-
urine#:~:text=Urine%20does%20not%20usually%20have,and%20infections%20are%20also%20possible.
7. Urinometer
Shankar S, Dr Vijay. “URINOMETER | Pathology Made Simple.” URINOMETER | Pathology Made
Simple, ilovepathology.com, 18 Sept. 2017, https://ilovepathology.com/urinometer/.
8. HOD METER
“What Is Harmonic Oscillation Densitometry? Explained by FAQ Blog.” What Is Harmonic Oscillation
Densitometry? Explained by FAQ Blog, faq-blog.com, https://faq-blog.com/what-is-harmonic-oscillation-
densitometry. Accessed 20 July 2022.

47 | P a g e
9. Rothera’s test
“Rothera’s Test for Ketone Bodies: Principle, Procedure and Clinical Significances.” LaboratoryTests.Org,
Chemical Examination
10. Determination of reducing substance
“RCPA - Reducing Substances Urine.” RCPA - Reducing Substances Urine, www.rcpa.edu.au, 10 Jan. 2019,
https://www.rcpa.edu.au/Manuals/RCPA-Manual/Pathology-Tests/R/Reducing-substances-urine.
Parikh, Himil. “Physical and Chemical Examination of Urine.” Physical and Chemical Examination of Urine,
www.slideshare.net, 1 Mar. 2001, https://www.slideshare.net/100000424773559/physical-and-chemical-
examination-of-urine.
11. Benedict Test
“Benedicts Test - Principle, Procedure, Result and Limitation.” VEDANTU, www.vedantu.com,
https://www.vedantu.com/chemistry/benedicts-test. Accessed 20 July 2022.
“Benedict’s Test - Reagent Preparation, Principle, Procedure, Reaction.” BYJUS, byjus.com, 2 Mar. 2021,
https://byjus.com/chemistry/benedicts-test/.
Aryal, Sagar. “Benedict’s Test- Principle, Preparation, Procedure and Result Interpretation.” Microbiology
Info.Com, microbiologyinfo.com, 29 Oct. 2015, https://microbiologyinfo.com/benedicts-test-principle-
composition-preparation-procedure-and-result-interpretation/.
“Benedict Test.” Benedict Test, www.pdfdrive.com,
https://www.pdfdrive.com/search?q=benedict+test&pagecount=&pubyear=&searchin=&em=. Accessed 20
July 2022.
laboratorytests.org, 5 Nov. 2018, https://laboratorytests.org/rotheras-test/.
12. Garhads test
“Rothera’s Test for Ketone Bodies: Principle, Procedure and Clinical Significances.” LaboratoryTests.Org,
laboratorytests.org, 5 Nov. 2018, https://laboratorytests.org/rotheras-test/.
https://www.labce.com/spg961108_microscopic_examination_of_urine_sediment.aspx
“Urine Test: Microscopic Urinalysis (for Parents) - Nemours KidsHealth.” Urine Test: Microscopic
Urinalysis (for Parents) - Nemours KidsHealth, kidshealth.org, https://kidshealth.org/en/parents/test-
urinalysis.html#:~:text=After%20a%20urine%20(pee)%20sample,then%20viewed%20under%20a%20micr
oscope. Accessed 21 July 2022.
14. Microscope
“How to Handle a Microscope.” Sciencing, sciencing.com, 10 Mar. 2018, https://sciencing.com/handle-
microscope-5650017.html.

48 | P a g e
“Urine Analysis:- Microscopic Examination, and Interpretations (Part 2) - Labpedia.Net.” Labpedia.Net,
labpedia.net, 25 Jan. 2020, https://labpedia.net/urine-microscopic-examination-and-interpretations-part-
2/.
16. Urine Report Analysis
“RBC Urine Test | Multimedia Encyclopedia | Health Information | St. Luke’s Hospital.” RBC Urine
Test | Multimedia Encyclopedia | Health Information | St. Luke’s Hospital, www.stlukes-stl.com, 29
Aug. 2015, https://www.stlukes-stl.com/health-content/health-ency-multimedia/1/003582.htm.
“Urine Glucose Test: Purpose, Procedure, and Results.” Urine Glucose Test: Purpose, Procedure, and
Results, www.healthline.com, https://www.healthline.com/health/glucose-test-urine. Accessed 22 July
2022.
17. Additional Reference
Multistix© 10 SG reagent strips [Color chart]. Tarrytown, NY: Bayer Corporation Diagnostics Division,
1996.
. Multistix© 10 SG reagent strips [Package insert]. Tarrytown, NY: Siemens Healthcare Diagnostics Inc,
2008.
de Wardener HE. The Kidney. 3rd Ed. Boston: Little, Brown & Co, 1967.
Lecture Notes

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Area of evaluation Weightage Marks awarded
Accuracy of language 20 marks
Relevance of content 30 marks
Appropriateness of style 15 marks
Following assignment guidelines 15 marks
References 20 marks
Final marks evaluation
Overall comment by the examiner

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