AN EXERCISE IN THE DETERMINATION OF URINE SPECIFIC
GRAVITY AND LONG'S COEFFICIENT
INTRODUCTION
The specific gravity of a liquid is the ratio of the density of the liquid to the density of pure water (1.0 gram/ml).
The closer the specific gravity is to 1.000, the closer it is to pure water. When solutes are added to a liquid, the
specific gravity increases. Therefore, the specific gravity of a urine sample is indicative of the amount of solutes
present in the urine. The higher the value, the more solutes are dissolved in the urine. The use of Long's
coefficient is a relative mathematical means of comparing the specific gravity of a urine sample to the actual
amount of solute present in the sample. The normal specific gravity for various urine samples are given below:
Adults: Random Specimen = 1.010-1.025
24 hour Specimen* = 1.015-1.018
* total urine collected during a 24 hour period
Newborn babies Random Specimen = 1.002-1.004
There is a progressive decrease in the urine specific gravity as one reaches middle age and over.
When using the urinometer to measure the specific gravity of a sample, it is important to note that the scale on the
urinometer is calibrated only for a temperature of 15.5 degrees C. In order to obtain a correct reading of any
given urine sample the temperature of the sample must be known and corrected to the calibrated temperature.
MATERIALS NEEDED
The kit in the metal cabinet contains the following:
(1) urinometer (special hydrometer for urine specific gravity)
(1) urinometer cylinder
(1) bottle of sodium chloride crystals
Thermometers are available on the front table.
PROCEDURES
1. Before actually measuring the specific gravity of a urine sample, you should practice reading the urinometer
2.
3.
4.
5.
scale in different solutions. Do this by filling the cylinder about 3/4 full of deionized water (white DI-on
laboratory sink). Add a small pinch of sodium chloride to give some solute to the pure water. Read the
temperature of the water sample using the centigrade thermometer. Grasp the urinometer float at its top and
slowly insert into the cylinder (insert weighted end first - see diagram). Avoid wetting the float stem above
the liquid line. Excessive wetting of the stem will cause the float to sink below the true test reading. Be very
careful of the urinometer float because they can break easily.
Using your thumb and index finger, impart a slight spin to the float as it is released into the sample.
Remember that the urinometer must be able to float in the liquid, add more liquid if necessary to insure a free
floating urinometer. This spinning helps keep the float in the middle of the sample and not against the
cylinder sides.
Read the scale of the urinometer at the lowest portion of the meniscus (see diagram).
To correct the reading for the temperature variation, use the following method: Take the recorded
temperature of the sample in either C or F degree value. If you work with F (Fahrenheit) degrees, for every
5.4°F above 60°F add 0.001 to the float reading. For sample temperatures below 60°F, subtract 0.001 for
every 5.4°F.
If you are working with C (Centigrade or Celsius) degrees, for every 3.0°C above 15.5°C, add 0.001 to the
float reading and for every 3.0°C below 15.5°C subtract 0.001 from the reading.
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For example:
The urinometer float shows a specific gravity of 1.015 and the measured temperature is 82°F. Subtract: 82°F 60°F = 22°F. Divide 22 degrees by 5.4 degrees = 4.07 = 4.00 (rounded). Multiply 4.00 x 0.001 = 0.004.
Corrected specific gravity =1.015 + 0.004 = 1.019
6. After the above procedure, clean out the cylinder. If you like, add a little more or less sodium chloride than in
the previous procedure and determine the temperature corrected specific gravity. Practice this until you
understand the procedure well.
7. In the bin marked "Urine Containers" in the front of the laboratory, there are some clean urine cups. Use them
to collect some real urine and determine the corrected specific gravity of the urine sample.
8. Long's coefficient may be used to show the relationship between the specific gravity of a sample and the
amount of solute dissolved in the sample. Take the last two figures of the corrected specific gravity sample
and multiply by 2.6 (Long's coefficient). This will give you the approximate grams of dissolved solute in a
liter of the sample.
9. For example: 1.019: 19 x 2.6 = 49.4 grams/liter
10. Calculate the gram/liter values of your various samples you tested in the laboratory. Record all of the values
here:
11. Be sure to clean all equipment well and return to the metal cabinets. Rinse out the urine cup in the laboratory
sinks and discard in the wastebasket.
Discussion and Study Questions:
 How did you perform this experiment?
 What is specific gravity of a liquid?
 What does a specific gravity of 1.0 gram/ml mean?
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 If a solution has a higher specific gravity what does this mean?
 Why would a urine sample be collected over 24 hour period for a urine specific gravity (USG)? Why
not just one sample?
 How do we have to adjust for temperature? In other words, why would a change in temperature cause
the specific gravity to change?
 How do we correct for temperature fluctuations above 15.5 degrees Celsius or 60 degrees F?
 What does the Long’s coefficient (2.6) tell you?
 How would the following conditions alter USG?

Sweating?

Glucosuria?

Dehydration?

Excessive fluid intake?

Diabetes insipidus?
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AN EXERCISE IN THE METHODS OF URINE ANALYSISCHEMICAL ANALYSIS
INTRODUCTION
The chemical analysis of a urine sample is usually done using "dip-stixs." These include CLINISTIXS,
KETOSTIXS, MULTISTIXS and COMBISTIXS. Also, a tablet called CLINITEST is used. These are chemically
treated paper sticks and tablets that are placed into a urine sample. Each "stix" has a particular chemical test to
detect the presence or absence of a substance in the urine. For example: CLINSTIX tests for glucose, KETOSTIX
tests for ketone bodies such as acetoacetic acid, MULTISTIX tests for pH, protein, glucose, ketones, bilirubin,
blood and urobilinogen, and COMBISTIX tests for pH, glucose and protein in the urine. The CLINITEST tablets
are used to test for the presence of glucose in a urine sample.
The Hay's test is for bile salts in the urine and uses the detergent action of the bile salts in the urine to lower the
surface tension of the water in the urine. If a small amount of sulfur is added to the urine sample, the presence of
bile salts will cause the sulfur to sink.
MATERIALS NEEDED (It is not necessary to try every dip stick version, as long as you obtain results for most
urine products. They are redundant.)
The kit in the metal cabinets will contain the following:
(1) Multistix bottle
(1) Ketostix bottle
(1) Combistix bottle
(1) Clinistix bottle
(1) Clinitest bottle (this may not be available check with instructor)
(1) small bottle of sulfur powder
(1) metal scoop for sulfur powder
(1) medicine dropper
(1) test tube
(1) forceps
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PROCEDURES
1. Obtain one of the urine cups from the bin marked "Urine Containers" and get a fresh real
urine sample.
2. Take the bottles of Multistix, Combistix, Ketostix and Clinistix and read the instructions
on the side of the bottles. Each instruction sheet is complete with a color chart that
explains the chemical test. Be sure you can read each of the "stix" tests. Be sure that you
replace the cap on the bottle after you have removed the strip for the test. The cap should
be replaced "promptly" and "tightly". Use the urine in the urine container for your tests
and record the results in your laboratory book. You might note here that a high level of
ascorbic acid (vitamin C) in the diet may alter all of the tests for the presence of glucose.
Complete this chart to document your test results.
Stix used
Substance tested
results
3. To perform the Clinitest for the presence of glucose follow the following procedure:
a. Place 10 drops of water and 5 drops of fresh urine in the test tube.
b. Using the forceps in the kit, add one Clinitest tablet to the test tube. Be careful-the
tablet contains sodium hydroxide and may cause burns. The sodium hydroxide in the
tablet generates heat so that the liquid will boil in the test tube.
c. A high concentration of phosphates may produce a white precipitate. If glucose is
present in the sample, a reddish-colored precipitate (cuprous oxide) is formed. The
solution may be turned green, yellow, orange, or red depending on the relative
concentration of glucose in the urine sample. The following code is usually used for
recording the results:
SOLUTION COLOR:
Blue
Green
Yellow
Orange
Red
Test is Negative
Record "+"
Record "++"
Record "+++"
Record "++++"
The relative number of "+" indicates the degree of glucose concentration, green is lowest and red
being the greatest amount of glucose in the urine. Be able to relate these findings with the
concept of the Tm for glucose as discussed in the lecture. Be sure to clean the test tube with soap
and water.
Record the results here:
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4. Use the following procedure for the Hay's test for the presence of bile salts in the urine:
i. fill the clean test tube 1/2 full of urine. Sprinkle a small amount of sulfur
powder (use the metal scoop) on the surface of the cool urine.
ii. if the sulfur sinks at once, bile salts are present in the amount of 0.01% or
more. If the sulfur sinks only after gentle agitation, bile salts are present in
the amount of 0.0025% or more. If the sulfur remains floating even after
gentle shaking, bile salts are absent.
5. Be sure to wash and dry the test tube before returning it to the kit. Discard urine down the
sink. Clean the urine cup in the sink and discard in the trash.
Record your findings for bile salts here:
Discussion and Study Questions:
 How did you perform this experiment?
 Why are the “dip-stixs” clinically important?
 What did the CLINITEST tablets test for and what did the MULTISTIX test for? What
did the Hay’s test tell you?
 Is glucose normally found in the urine?
Urobilinogen? Bile salts?
Protein?
Ketones?
Blood? Bilirubin?
 If these items (above) are found in the urine and SHOULD NOT be, what does this tell
you about the patient (specifically)?
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 What is the pH of urine?
 What is the relevance of the colors for the Clinitest? If the color is red, what would this
suggest about the patients Tm for glucose?
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AN EXERCISE IN THE METHODS OF URINE ANALYSISMICROSCOPIC SEDIMENTS
INTRODUCTION
The normal sediment in urine varies considerably with the sex of the subject, the time of the
day when the specimen was taken, the specimen pH and how the specimen was collected and
stored. The urine sediment usually will contain some squamous epithelial cells. These originate
in the mucosa of the bladder and urethra, but may also originate in the vaginal mucosa of
females. Various sediment crystals may also be present. Calcium oxalate and yellow uric acid
crystals are common in acidic urine. Often, a few leukocytes are seen; however, amounts greater
than 6 to 10 per high power field IN THE MALE are regarded with suspicion and may be
diagnostic of a Urinary Tract Infection (UTI). Slightly higher numbers of leukocytes are
considered normal IN FEMALES because of them originating from the vaginal mucosa.
Erythrocytes are usually not present; however they may be present after exercising or perhaps
following menses.
Bacteria are not normally present in freshly voided, properly handled urine. However,
because of contamination, bacteria may multiply rapidly over a relatively short period of time.
Large numbers of bacteria in freshly voided urine may be indicative of UTI.
Casts are results of solidification of albumin that has entered the uriniferous tubule. Casts are
usually hollow and they retain the shape of the tubule. The casts may contain any material which
was lying free in the tubule or loosely attached to its wall. Types of casts include: granular, RBC,
WBC, epithelial and hyaline (clear). Casts may be recognized by their parallel sides and rounded
ends. Large numbers of casts are usually associated with albuminuria and may indicate the
presence of renal disease.
Ova of certain parasites, sperm, trichimonads, yeast, hair and tissue paper fibers may also be
found in a urine specimen.
Slides are normally prepared from fresh urine samples. A 10 ml urine sample is centrifuged for
5 minutes to collect sediment. The urine supernatant is poured off, and the sediment is stained
with 2 drops of Sedi-stain, a dye which allows one to visualize the cells and other elements. A
drop of stained sediment is placed on a glass slide, covered with a coverslip and examined
directly under a compound microscope. You will be examining a prepared slide of a fixed,
stained urine sample.
MATERIALS NEEDED
Obtain a prepared urine slide and a compound microscope.
PROCEDURES
1. Examine the specimen under both low and high power. Be sure to adjust the light, the
condenser and the iris diaphragm to maximize your view.
2. Refer to the diagram and explanation sheets below and on the following pages to identify the
various sediments.
3. Sketch and identify each of the observed sediments below in the space provided.
4. Return the slide to the slide tray. Replace the compound microscope in its proper location.
Slides: Normal Urine Sediments, unstained, Triole phosphate, cystine, Celesium oxalate,
Calcium phosphate, Hippuric acid, Uric acid
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Urinary
Sediments
Refer to
http://www.ec.upstate.edu/path
/urinalysis/frame.htm
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Diagram Key Number
Description and Pathology
1,2
Epithelial cells are the covering cells from the urethra and prepuce in
males and the external genitalia in females.
If leukocytes occur in large numbers this may indicate a renal infection
Squamous cells are considered the same as epithelial cells
Mucus threads are present in small numbers normally. Increased
numbers are found in chronic inflammation of the urethra and bladder
Casts are cylindrical structures with parallel sides and blunt rounded
ends. They are formed in the renal tubules. Casts consist of
mucoprotein matrix with formed elements imbedded. If the formed
elements are RBC’s, this would indicate glomerular lesions. If the
formed elements are leukocytes, this would indicate renal infection.
Epithelial casts have epithelial cells in the mucoprotein matrix.
Granular casts in the urine usually indicate serious renal damage. There
is a high concentration of protein in the matrix.
Hyaline casts usually indicate mild renal damage. There are clear casts
with little imbedded materials in the matrix.
Uric acid crystals are rhombic, six-sided plates, often arranged in
rosette-like clusters.
A presence of cholesterol crystals would indicate the level of saturated
fats in the body. The presence of these crystals is rare.
The presence of calcium phosphate crystals would indicate an alkaline
urine.
Cystine crystals are flat, hexagonal plates with unequal sides. The
presence of these crystals may indicate cystinuria. Cystinuria is a
congenital defect involving the reabsorption of cystine by the renal
tubules.
The presence of tyrosine crystals indicates serious liver damage. The
crystals appear as fine needles arranged in radiating sheaves.
Calcium oxalate crystals are colorless, octahedral crystals with a highly
refractile cross connecting the corners. They are present in acidic urine
but occasionally in alkaline urine.
Hipuuric acid crystals will occur seldom in urine samples.
Ammonium –magnesium phosphate crystals are prism shaped crystals
presenting three, four or six sides. These crystals are found in alkaline
urine.
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9,10
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Study Questions:
 What objects would you expect to see in urine? Sketch them here.
 Which objects would be a sign of pathology?
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