URINE ANALYSIS & MICROSCOPY, SPECIFIC GRAVITY & OSMOLALITY

Objectives :

At the end of the practical, the student should be able to,1. Describe how to collect a sample of urine for analysis2. Interpret the results of macroscopic examination of a sample of urine3. Identify the following in a sample of urine, under the microscope

a. RBCb. WBCc. Epithelial cellsd. Casts (Hyaline, cellular, granular)e. Crystals

4. Explain the physiological significance of the following, a. Observation of RBCs, WBCs, Casts or Crystals in a sample of

urineb. Polyuria, Oliguria and Anuria.

5. Measure the specific gravity of a sample of urine.6. Describe the technique of measuring serum and urine osmolality.7. Student Exercise : Interpret the physiological basis of the changes observed in

serum and urine after drinking solutions of varying osmolality.

Urine Analysis

Macroscopic Microscopic Chemical Bacteriological

Quantity Quality Cells Sugar Gram stain Casts Protein Culture & ABST

Crystals

Colour Appearance Smell Precipitate

Practical No : 17

Collection of a sample of urine

Choose a wide mouthed,

screw capped, sterile bottle

Male Female Retract foreskin Wash with water Do not contaminate the rim of the

bottle Collect midstream urine Immediately close the lid of the

bottle

Separate labial folds Wash with water Do not contaminate the rim of the

bottle Collect midstream urine Immediately close the lid of the

bottle

Label the bottle accurately with the patient's name and BHT number Fill the request form Send to the lab immediately, or if delayed > 1 hr, put in the refrigerator

Macroscopic appearance of urine

Quantity : Volume, normally 700 - 2500 ml/day (0.5 - 1 ml/kg/hr)

Volume - fluid intakeConsumption of AlcoholEating salty foods

Volume - fluid intakeDehydration

Anuria is the absence of a urine output and is seen in,Acute renal failureUrinary tract obstruction

Oliguria is a urine output of < 400 ml/day in an adult. Is seen in,Severe dehydrationAcute renal failureAcute glomerulonephritis

Polyuria is a urine output of > 2500 ml/day and is seen in,Diabetes MellitusDiabetes InsipidusChronic Renal Failure

Quality : Colour

Characteristic colour of urine is due to the pigments, urochrome and uroerythrin

Normally, is a light straw colour, which darkens on standing due to oxidation of urobilinogen to urobilin.

Tea coloured urine - due to urobilin (obstr. Jaundice) Dark Red urine - due to Hb in urine (intravascular h'lysis) Bright Red urine - due to large number of red cells Black colour - phenylketonuria Colour due to foods and drugs

o Brown - metronidazoleo Orange - Rifampicino Yellow - Vit. B

Appearance Smoky appearance due to a little amount of red cells in urine Turbid appearance due to proteins Frothy urine due to proteins and bile salts

Smell A fishy smell in ketonuria (diabetic ketoacidosis)

Precipitate Whitish precipitate seen in heavy proteinuria

Microscopic appearance of urine

Cells

Red Blood Cells White Blood Cells Epithelial Cells

- Are small circular cells with a yellowish center

- 2 -3 cells/mm3 in females

- normally no cells in males

- Increased amounts seen in,

Glomerulonephritis UTI Urinary calculi

- 'Pus cells'- Larger than RBC- Round shaped with lobed

nuclei and a granular cytoplasm- Normally <10cells/mm3

- Increased amounts in,CystitisUrethritisNephritisProstatitis

- Are nucleated, flat or columnar cells

- Normally, only little amounts seen.

- If large amounts, there may be some tubular damage

Casts

- Are cylindrical structures formed by precipitation of muco-proteins (Tamm - Horsefal proteins) in the tubular lumen.

- Hyaline Casts - Pale, transparent and homogenous apperance- Cellular Casts - Cells are embedded on the hyaline casts, to form RBC,

WBC, Epithelial cell casts- Granular Casts - Disintegrated cells on the cellular casts give the

granular Appearance

Crystals

- Are formed by precipitation of chemicals in urine.- Ammonium, Magnesium, Calcium phosphate crystals (formed in alkaline urine)

also known as Triple Phosphate crystals have a "coffin lid" shape.- Calcium Oxalate crystals are formed in acid urine and have an envelope shape.- Uric acid crystals maybe normal, but are significantly increased in

hyperuricaemia (gout).

1 3 4

Measuring the Specific gravity of a sample of urine

Specific gravity is measured by means of a urinometer calibrated from 1.000 to 1.060. (The specific Gravity of Distilled water is 1.000 at a temperature of 20oC).

1 Leucocyte and pus cell2,3,4 Polygonal, epithelial cells5,6 Squamous epithelial cells7 Hyaline casts8 Pseudohyaline9 Epithelial cell cast10 Erythrocyte cast11 Leucocyte cast and leucocytes12 Granular casts (coarse granules)13 Waxy cast (fine granules)14 Fibres15 Bacterial casts and bacteria

16 Spermatozoa17 ,18 Triple phosphate crystals19 Calcium hydrogen phosphate20 Uric acid crystals21 Calcium carbonate crystals22 Ammonium urate crystals23 Leucine crystals24 Tyrosine crystals25 Calcium crystals26 Calcium urate crystals27 Cysteine crystals

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89

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24 25 26 27

The temperature of urine must also be measured for correct calculation of the specific gravity.

Value of the test :-The Specific Gravity of urine varies according to kidney function.

Concentrated urine = High SGDilute urine = Low SG

Materials :-1 urinometer1 Thermometer (0 - 50oC)1 Measuring Cylinder (50 ml)Sample of urine (approx. 40 ml)

Method :-- Put the sample of urine into the

measuring cylinder (if urine sample is small, dilute with water at a known percentage)

- The urinometer is then gently floated in the urine.

- Wait for it to settle.- The urinometer must not be in contact

with the sides of the measuring cylinder.- The depth to which it sinks, is observed

directly from the calibration, and recorded.- Using a thermometer, the temperature of

that sample of urine, is measured.

Observation :-- Urinometer reading - 1.020- Temperature - 32oC

Calculation :-- Add to the Specific Gravity recorded, 0.001 for every 3oC that the urine

temperature is higher than the calibration temperature- Subtract from the Specific Gravity recorded, 0.001 for every 3oC below the

calibration temperature.- Therefore,

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Temperature = (measured temperature - 20 o C) x 0.001 correction 3

Temperature = (32 o C - 20 o C) x 0.001 correction 3

= 0.004

Specific gravity = urinometer reading + temp. correction1.020 + 0.0041.024

Measuring cylinder

Thermometer

Urine sample

Urinometer

Interpretation :

Normal Specific Gravity 1.020 (1.010 - 1.025)Low Specific Gravity Below 1.010 (kidney or Endocrine disorder)High Specific Gravity Above 1.025 (Glycosuria, Proteinuria)

Measuring serum osmolality

Method :-

- A blood sample is collected from the patient, allowed to clot and the serum separated.

- Using a micropipette, 150 l of serum is measured and put into a small tube.- Attach the tube into the head of the Osmometer and place in the cooling cavity.- First, reduce the temperature upto zero Celsius and then further reduce it upto

freezing point.- Osmolality is read at the freezing point.- Within the cooling cavity, the sample will solidify within 2 - 3 minutes.- The indicator of the instrument moves when cooling and when comes to the

mark, press the vibrator button.- Read the osmolality directly.- Before putting another sample, the de-freezing button has to be switched on and

the sample de-freezed.- As there are three scales in the machine, the scale relevant to the sample should be

used. (Approximate osmolality)-

0 - 400 0 - 800 0 - 1600

Normal serum osmolality : 280 - 296 mosm/kg of water

Urine osmolality is measured in a similar manner to the above method.

Questions :

1. What is the normal Serum Osmolality?2. What is the normal Urine Osmolality

3. What is the normal Urine Specific Gravity

Exercise : Serum and urine changes after dehydration and different fluid loads

Method :-

- Seven volunteers who had not had a significant meal or drink for some time, are selected.

- Their urine and serum osmolality, urine output and specific gravity are measured.-- Then 6 of them are given one each of the following solutions to drink.

1 L of water 1 L of Oral Rehydration Solution 1 L of 5% glucose 1 L of King Coconut water 1 L of water + Frusemide 1 L of water + Lime juice

- One of these volunteers, is not allowed to drink any type of fluid (i.e. dehydrated)

- After 45 minutes of drinking their respective solutions, another urine sample is obtained, and the specific gravity and osmolality of each is measured and recorded.

- After 1 1/2 hours of drinking, another urine sample as well as a blood sample is obtained from each subject, and the serum and urine osmolality, and urine specific gravity is measured.

- Fill the chart on the following page and give the physiological basis for the changes observed.

SituationOsmolality

of fluid (mosm/l)

At 0 hours After 45 minutes

After 1 1/2 hours

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After Dehydration

1 L water

1 L ORS

1 L 5% glucose

1 L King Coconut water

1 L H2O + frusemide

1 L H2O + lime juice