Urine Analysis

Sample for urine analysis

1. The urine may be random or morning samples.
2. Urine must be tested within 1 to 2 hours of collection.
3. It may need 24 hours of urine collection.

Precautions for urine analysis

1. Urine can be refrigerated for 6 to 8 hours.
   1. At low temperatures, there will be precipitation of amorphous urates or phosphates.
2. Bring the refrigerated urine to room temperature before testing.
3. Urine collected for more than 2 hours may be rejected.
4. The decomposition of urine begins in 30 minutes at room temperature and 4 hours in refrigeration.
5. For longer periods, boric acid can be used as a preservative.

Factors that will interfere with the urine analysis:

1. Certain foods will color the urine, like:
   1. Carrots will change their urine color to dark yellow.
   2. Beets may cause red color urine.
   3. Rhubarb will change its color to red or brown.
2. Asparagus produces a typical urine odor.
3. If you keep urine on the table, bacteria will change the urine pH, make it alkaline because of urea splitting bacteria and there is the production of the ammonia.
4. Effect of the food on the urine pH:
   1. Eating citrus fruits, dairy products, and vegetables makes urine alkaline.
   2. A high protein diet and some foods (cranberries) make urine acidic.
   3. A high protein, a fat diet without the intake of carbohydrates also causes ketonuria.
5. When radiopaque contrast media is given to the patient will give false-positive urine protein for at least 3 days.
6. Females may have contamination from the vaginal secretions and show an increased number of WBCs.
7. Traumatic catheterization may cause hematuria and false RBCs in the urine.
8. Vaginal secretions contamination may cause false-positive leucocyte esterase.
9. False proteinuria is seen when urine is contaminated with vaginal secretions.
10. Urine culture gives false-positive results when contaminated with stool, vaginal secretions, and contaminated test tubes or hands and clothes.

Complete Urine Analysis

The volume of urine:

1. It is 600 to 2500 ml /24 hours, and the average volume is 1200 ml/24 hours.

Polyuria:

1. It occurs when urine amount exceeds 2000 mL/24 hours.
   1. This is seen in diabetes mellitus.
   2. In diabetes Insipidus.
   3. Excessive fluid intake.
   4. Drugs like diuretic therapy.

Oliguria:

1. It is seen when urine volume is less than 500 mL/ 24 hours. This is seen in the following conditions.
   1. Renal tubular dysfunction.
   2. Edema.
   3. End-stage kidneys.
   4. Obstructive uropathy.
   5. Dehydration.
   6. Vomiting.
   7. Shock.
   8. Diarrhea.

Anuria: 

1. It occurs when there is no urine output. This is seen in the following conditions:
2. 1. Acute renal failure.
   2. Heart failure.
   3. Acute Glomerulonephritis.
   4. Obstructive uropathy.
   5. Shock.

Urine classification based on volume

urine output can be calculated as follows:

1. Adult = >0.5 mL/Kg/hour.
2. Child = >1 mL/Kg/hour.
3. Neonate  (< 1-year-old) = >2 mL/Kg/hour.

Urine volume factors:

1. It varies according to age.
2. Urine volume is 600 to 2500 mL/day, and the average is 1200 mL/day.
3. At night urine volume is usually <700 mL, and specific gravity are more than 1.018.
4. The ratio of the night-to-day urine volume is 1:2 to 1:4.
5. The range of 600 to 2000 mL/day is considered normal.

Table showing the variation of the urine volume in various age groups:

Color of urine:

1. The normal color is Pale to dark yellow.

Abnormal Color:

1. It may be due to some of the foods and medicines.
2. Urine with no color may be caused by long-term kidney disease or uncontrolled diabetes.
3. Dark yellow urine can be caused by dehydration.
4. Red urine can be caused by blood in the urine.
5. The Orange color is due to bilirubin.
6. The dark yellow color may be seen due to bilirubin and urobilin.
7. Brown and black colors may be due to melanin, homogentisic acid, and biliary pigments.
8. Green-colored urine may be seen due to Biliverdin, pseudomonas infection, and oxidized bilirubin.

Cloudy urine can be caused by:

1. Pus (white blood cells).
2. Blood (red blood cells).
3. Sperm, Bacteria, and yeast.
4. Crystals.
5. Mucus.
6. Parasite infection, such as trichomoniasis.

Red color urine may be:

1. When clear, this will indicate the following:
   1. Hemoglobinuria.
   2. Myoglobinuria.
2. When cloudy red urine indicates:
   1. Hematuria.

Various colors of the urine

Various colors of urine in different conditions:

Drugs that can change the color of the urine:

Clarity of urine:

1. Normally it is Clear. No visible particles are seen.
   1. Hazy urine is where there are few particulates, and through urine, print can be seen easily.
   2. Cloudy when there are many particulates, and through urine, the print is blurred.
   3. Turbid urine, when through urine print, cannot be seen.
   4. Milky where there are many particulates, and it may be clotted.

Urine turbidity:

1. It is seen in nonpathological conditions like:
   1. With the presence of squamous epithelial cells.
   2. The presence of mucus.
   3. The presence of spermatozoa.
   4. Amorphous phosphates, urates, and carbonates.
   5. In the case of contrast media.
   6. In the case of fecal contamination.

Pathological urine turbidity:

1. Urine turbidity is seen in pathological conditions like:
   1. White blood cells.
   2. Red blood cells.
   3. Presence of bacteria.
   4. Presence of yeast.
   5. Abnormal crystals.
   6. Lymph fluids and lipids.

Turbid urine solubility in various conditions:

1. In Alkaline urine, amorphous phosphate and carbonates are seen.
2. In acidic urine, amorphous urates, and radiographic contrast media.
3. Soluble by heat are amorphous urates and uric acid crystals.

The odor of urine:

1. The odor is not clinically significant.  This is not part of the urine analysis.
2. Normally odor is slightly “nutty” (faint, aromatic odor).
   1. On keeping the urine, the ammonia odor is more prominent. This is because of the breakdown of the urea.

Abnormal Odor may be due to the following:

1. Some foods, such as asparagus, onions, and garlic, cause a pungent odor.
2. Vitamins and antibiotics (such as penicillin) can cause urine to have a different odor.
3. A sweet, fruity odor may be caused by uncontrolled diabetes mellitus.
4. A urinary tract infection (UTI) can cause a bad, unpleasant odor.
5. Urine that smells like maple syrup means maple syrup urine disease when the body cannot break down certain amino acids.
6. Phenylketonuria gives a mousy smell.
7. Bleach-like odor is due to contamination.

Various causes of the odor of urine:

The specific gravity of urine:

1. The kidneys’ ability to selectively reabsorb essential chemicals and water from the glomerular filtrate is one of the most important functions.
2. If the urine is refrigerated, then bring it to room temperature for measurement of specific gravity.
3. Specific gravity is the density of a substance relative to the density of distle water.
4. The normal range of urine specific gravity = 1.005 to 1.030
   1. The normal urine sample ranges between 1.010 to 1.025.
   2. Specific gravity is higher in the first-morning urine specimen (>1.020).

5. The normal range of urine-specific gravity = 1.005 – 1.030.
   1. Neonates = 1.012.
   2. Infants = 1.002 to 1.006.
   3. Adults = 1.003 to 1.030.
6. The urinometer measures specific gravity.
   1. To check the urinometer’s reliability, put this into the water, and it should read 1.000.
   2. It can check by using the known specific gravity solution.

Urine-specific gravity measurement by urinometer

A urine refractometer:

1. It measures the refractive index of the urine.
2. Its principle is that the refractive index compares the velocity of the light in air with the velocity of light in a solution.
3. The dissolved particle concentration in the solution will determine the velocity and angle at which the light will pass through the solution.
4. The refractometer’s advantage is that it uses only 1 to 2 drops of urine when the urinometer needs more.

Urine refractometer

Abnormal Specific Gravity classification:

The pH of the urine:

1. Normal pH = 4.6  to  8.0.
2. The average person has a pH of 5 to 6.

Urine analysis pH normal range

3. pH is helpful for the identification of crystals.
4. pH is helpful for the acid-base status.

Urine pH, the role of kidneys and lungs

Urine pH in acidosis/alkalosis

Urine pH control mechanism

Factors affecting the urine pH:

1. Acidic urine is produced in the following ways:
   1. High protein diet.
   2. Lung diseases occur when CO2 is retained in the body.
   3. Diabetes mellitus in uncontrolled diabetics.
2. Conditions producing alkaline urine:
   1. Urinary tract infection.
   2. Bacterial contamination of the urine.
   3. A diet high in vegetables, citrus fruits,  or dairy products makes more alkaline urine.
   4. Hyperventilation and loss of CO2.
   5. When treated with NaHCO3 and K citrate and antibiotics in treating the urinary tract infection.

Abnormal pH.

1. Some foods (citrus fruit and dairy products) and medicines (such as antacids) can affect urine pH.

Significance of pH:

1. Maintenance of acidic urine can be good for treating urinary tract infections caused by urea-splitting microbes because these bacteria can not grow in an acid medium.
2. People on a high-protein diet produce acidic urine, and people on a vegetarian diet produce alkaline urine.
3. The exception is cranberry produces acidic urine, and it can be used as a home remedy for urinary tract infections.
4. Alkaline urine is seen in patients with renal calculi made of calcium phosphate or magnesium-ammonium phosphate.

Measurement of urine pH:

1. There are various commercially available urine reagent strips to measure pH; these measure pH between 5 to 9.
2. Indicators are methyl red and bromthymol blue.
3. Methyl red changes color from red to yellow with a pH change of 4 to 6.
4. Bromthymol blue changes from yellow to blue with the change in pH from 6 to 9.

Proteins in the urine:

1. Normally urine is negative.
   1. It is <10 mg/dL or
   2. Around 100 mg/24 hours.
2. The protein consists of primarily low molecular weight serum proteins filtered by the glomeruli.
   1. Albumin is the major protein found in normal urine.
   2. The tubules reabsorb the majority of the albumin.
   3. Proteinuria may be seen after vigorous exercise, dehydration, and fever.
   4. Benign proteinuria is postural and intermittent; this accounts for 5% of people.
3. Proteinuria may be classified based on the origin of the protein into:

Prerenal proteinuria:

1. This is not indicative of kidney disease.
2. This is due to the increased level of low-molecular-weight plasma proteins such as hemoglobin, myoglobin, and acute-phase protein.
3. This is caused by  nonrenal diseases and is transient; it is seen in:
   1. Hemoglobinuria.
   2. Myoglobinuria.
   3. Acute phase proteinuria.
   4. This is usually not detected by the routine urine reagent strips.

Renal proteinuria:

1.  This is due to renal diseases involving glomeruli or tubules.
2. In glomerular damage, selective filtration is impaired, and an increased amount of albumin and red blood cells and white cells are excreted in the urine.
3. It is seen in the following:
   1. SLE.
   2. Streptococcal glomerulonephritis.
   3. Strenuous exercise  (reversible condition).
   4. Pre-eclampsia and hypertension. (reversible condition).
   5. Toxic heavy metals.
   6. Severe viral infection.
   7. Microalbuminuria is seen in diabetic nephropathy indicating kidney damage in diabetes mellitus.

Postrenal proteinuria:

1.  Proteins can be added as the urine passes through the ureter, urinary bladder, urethra, prostate, and vagina.
2. Bacterial and fungal infection of the lower urinary tract,
3. Menstrual contamination also contains proteins.
4. Prostatic fluid and spermatozoa.
5.  Bacterial and fungal infections and inflammation produce exudate-containing protein from the interstitial fluids.
6. The presence of blood resulting from injury or menstrual contamination contributes to protein, as does the presence of prostatic fluid and a large number of spermatozoa.

Orthostatic or postural proteinuria:

1.  This is a persistent benign condition frequently seen in young patients.
2. It appears when the person is upright and disappears when the patient lies down.
3. Procedure to confirm the diagnosis:
   1. These patients are advised to empty their bladders before going to bed.
   2. Take the first urine sample when patients get up.
   3. Take another sample when patients are upright for several hours.
      1. The first sample will be negative, and the second sample will be positive in orthostatic proteinuria.

Microalbuminuria:

1. This is seen in diabetic nephro[athy.
   
2. This microalbuminuria indicates diabetic complications.
3. The further complication of diabetes can be prevented by controlling diabetes.
4. This is also associated with an increased risk of cardiovascular diseases.
5. Microalbuminuria is reported as albumin excreted as µg/min or in mg/24 hours.
   1. The level is significant from 20 to 200 µg/min Or 30 to 300 mg/24 hours.
   2. The albumin/creatinine ratio is >3.4 mg/mmol in these cases.
   3. The first-morning specimen is recommended.

Bence Jones proteinuria:

1. It is seen in multiple myeloma patients.
2. These are monoclonal light-chain immunoglobulins.
3. This is a low molecular weight protein and is excreted in the urine.
4. Procedure to detect BJ protein:
   1. Heat the urine, and these proteins coagulate at 40 to 60 °C.
   2. Dissolve when the temperature reaches 100 °C.
      1. Reading the result: Urine turbid at 40 to 60 °C and clear at 100 °C is considered positive.

Summary of the proteinuria:

Method to detect proteinuria:

The urine dipstick:

1. The urine dipstick reagents detect urine protein. This is a semiquantitative test.
2. Trace positive reaction = 15 to 30 mg/dL.
3. When 4+ positive reaction = 2.0g/dL.
4. Trace proteinuria = 10 to 30 mg/dL.
   1. 1+ = 30 mg/dL.
   2. 2+ = 100 mg/dL
   3. 3+ = 300 mg/dL
   4. 4+ = >1000 mg/dL

The sulfosalicylic acid method:

1. It is a precipitation method.
2. Procedure:
3. Transfer 3 mL of centrifuged urine into a test tube.
4. Add 3 drops of sulfosalicylic acid (250 g/L) along the side of the test tube.
5. The acid will form a layer underneath the urine.
6. Do not mix, and let it remain for one minute.
7. Reading:
8. There is barely perceptible turbidity = reported as traces or ± (roughly 5 mg/dL).
9. The heavier turbidity graded = 1+ to 4+.
   1. No turbidity = 0 mg/dL (<6 mg/dL)
   2. Slightly turbid = Traces, 20 mg/dL (6 to 30 mg/dL)
   3. 1+ = 50 mg/dL (Print visible through the specimen, 30 to 100 mg/dL).
   4. 2+ = 200 mg/dL  (Print invisible, 100 to 200 mg/dL).
   5. 3+ = 500 mg/dL (Flocculation, 200 to 400 mg/dL).
   6. 4+ = >1000 mg/dL (Dense precipitate, >400 mg/dL).

Trichloroacetic acids (TCA):

1. This is also a precipitation method. This method is used in automation.
2. TCA precipitates urine proteins as a fine suspension. The turbidity of the urine suspension is proportional to the amount of proteins in the urine.
3. Procedure:
   1. Take TCA 12.5 g/dL.
   2. Dissolve it in the distle water and make up to 100 mL.
   3. Prepare the solution fresh every month.
   4. Make the standard protein solution in sodium chloride.
4. Take the urine clear of the turbidity; if needed, you can centrifuge the urine.
   1. Take 4 mL of urine, 4 mL of the standard, and 4 ml of urine as blank.
   2. Add TCA 1.0 mL.
   3. Add 1.0 mL of NaCl (9 g/L) to all three test tubes.
   4. After 5 to 10 minutes, read all three tubes at 420 nm.

Calculation formula:

Heat coagulation test:

1. Centrifuge the urine, and make urine clear.
2. Take 7.5 mL of urine in heat resistant test tube.
3. Add 3  drops of 5 M acetate buffer (pH 4.0 ).
   1. Make this buffer by mixing 24 ml of glacial acetic acid and 7.5 g of anhydrous sodium acetate in 100 ml of distle water and mixing well.
4. Reading the result:
   1. If precipitate forms at this stage will indicate mucin (a mucoprotein).
   2. Centrifuge and remove this precipitate.
      1. Now heat the upper half of the urine, and bring it to boiling.
         

         Urine heat test for the protein

   3. Now examine for turbidity, comparing the upper half to the lower half.
      1. Hardly visible turbidity = 1+ (10 to 30mg/dL) or  ±.
      2. Moderate turbidity = 2+ (40 to 100 mg/dL).
      3. Heavy turbidity = 3+ (200 to 500 mg/dL).
      4. A flocculant precipitate = 4+ (>500 mg/dL).

Postural (Orthostatic) proteinuria test:

1. Procedure:
2. Ask the patient to empty the bladder before going to bed.
3. Collect the urine immediately after getting up in the morning.
4. After the patient is in the vertical position for several hours, get a second sample.
5. Check the protein in both samples.
6. Postural (Orthostatic) proteinuria-positive cases will show:
   1. First-morning sample =   negative for urine proteins.
   2. Second sample             = Positive for urine protein.

 Table showing proteinuria grading: 

1. The degree of proteinuria	Protein excreted per 24 hours
   Mild	<1.0 g /24 hours
   Moderate	1.0 to 3 or 4 g /24 hours
   Severe	> 3 g / 24 hours

Various urine tests for proteins and their characteristics:

Classification of the proteinuria:

Protein in the urine may be due to the following conditions:

1. Kidney damage.
2. Infection.
3. Cancer.
4. High blood pressure.
5. Diabetes.
6. The Systemic lupus erythematosus (SLE).
7. Glomerulonephritis.
8. Heart failure.
9. Leukemia.
10. Poison (lead or mercury poisoning).
11. High blood pressure during pregnancy (preeclampsia).
12. Protein in the urine may mean :
13. Kidney damage.
14. Infection.
15. Cancer.
16. High blood pressure.
17. Diabetes.
18. The Systemic lupus erythematosus (SLE).
19. Glomerulonephritis.
20. Heart failure.
21. Leukemia.
22. Poison (lead or mercury poisoning).
23. High blood pressure during pregnancy (preeclampsia).
24. Proteinuria can be classified into different grades according to the amount of protein in the urine.

Beta-2-Microglobulin:

1. It is structurally related to Immunoglobulin. It is used to differentiate Glomerular from tubular dysfunction.

Glucose in urine:

1. The healthy individual has a blood glucose of 100 mg/dL or less.
2. This condition may be benign due to emotional stress or ingesting a large meal.
3. When due to a pathological condition, then it is seen in diabetes.
4. Normally urine is negative.
5. Urine glucose is detected by:
   1. The dipstick.
   2. The copper sulfate method (Clinitest).
6. Under normal circumstances, almost all glucose filtered by the glomerulus is reabsorbed in the proximal convoluted tubules.

The renal threshold of glucose:

1. The level at which the tubular reabsorption is stopped for glucose is 160 to 180 mg/dL, called the renal threshold. But mostly,
   it is considered around 180 mg/dL.

2. The main function of insulin is to convert glucose to glycogen for storage (glycogenesis).
3. The opposing hormone causes glycogen’s breakdown into glucose (glycogenolysis), leading to an increased circulating glucose level in the blood.
4. Epinephrine is a strong inhibitor of insulin secretion.
   1. Epinephrine is secreted in response to severe stress, cerebrovascular trauma, and myocardial infarction.
5. Renal glycosuria may be seen when the renal tubule absorption of glucose  is compromised; this may be seen in:
   1. End-stage renal disease.
   2. Osteomalacia.
   3. Fanconi’s syndrome.

Method to detect glucose in the urine:

Copper reduction method:

1. This test was used in the early days.
2. This test is based on the reduction of copper sulfate (CuSO4) to cuprous oxide in the presence of alkali and heat.
3. Reading: The color changes from blue (negative = CuSO4) to green, yellow, orange/red (CuO).
4. Urine glucose benedict reaction:

Glucose-oxidase reagent strips:

1. Glucose-oxidase detects specific glucose in the urine only, while copper reduction detects other reducing substances.
2. The result is reported as 1+, 2+, 3+, and 4+, and these colors also provide quantitative measurements ranging from 100 mg/dL to 2000 mg/dL or 0.1% to 2%.

Comparison of Benedict reaction and Oxidase method:

 Glucose in urine is seen in the following:

1. Intravenous (IV) fluids.
2. Uncontrolled diabetes.
3. Adrenal gland diseases.
4. Liver damage.
5. Brain Injury.
6. Poisoning.
7. Kidney diseases.
8. Pregnant women can have glucose in their urine, which is normal during pregnancy.

Hyperglycemia may be seen in nondiabetic cases where hormones act against the insulin, seen in conditions like:

1. Pancreatitis.
2. Pancreatic cancers.
3. Cushing’s syndrome.
4. Acromegaly.
5. Hyperthyroidism.
6. Pheochromocytoma.

Ketones in urine:

1. Ketones bodies are mainly three intermediate products from lipolysis, and these are excreted in the urine:
   1. Acetone 2%.
   2. Acetoacetate  20%.
   3. β-hydroxybutyric acid 78%.

Ketone bodies in diabetic patients

2. Normally the measurable amount of ketones doesn’t appear in the urine. All metabolized fat is broken down into carbon dioxide (CO2) and water (H2O).
3. When there is no carbohydrate for the body’s energy, fat stores are metabolized to supply energy, and ketone bodies appear in the urine.
4. The reason for the fat metabolism is the inability to metabolize carbohydrates, as seen in the following:
   1. Diabetes Mellitus.
   2. Vomiting.
   3. Inadequate intake of carbohydrates as in starvation.
5. Ketonuria indicates a deficiency of insulin and can regulate the dose of insulin.
   1. Ketones are excreted in combination with basic ions (Na⁺, K⁺. and Ca⁺⁺), so ketonuria produces ketoacidosis.

6. These patients have a fruity odor on their breath because acetone is very volatile and blown off from the lungs.
   1. Normally urine is negative.
7. It has importance in Diabetes Mellitus and particularly in uncontrolled diabetes mellitus patients.

Procedure to detect ketones bodies:

Ketone test strips or tablets:

1. These are used based on sodium nitroprusside (nitroferricyanide) reaction to measure ketone bodies.
2. This will detect acetoacetic acid or acetone in alkaline conditions.
3. β-hydroxybutyric acid does not react with this reagent.
4. Reporting of the result:
   1. Color changes from lavender to maroon (purple).
   2. Acetoacetate + acetone is expressed as:
      1. Negative = 0 mg/dL
      2. Small amount =  10 mg/dL
      3. Moderate amount = 30 mg/dL
      4. Large amount = 80 mg/dL

Rothera’s test:

1. Grind fine powder of :
   1. 10 g Na2CO3 (anhydrous).
   2.  20 g (NH4)2SO4.
   3. 0.5 g sodium nitroprusside.
   4. Mix the powder completely and stopper it tightly.
2. Procedure:  Take about 0.5 g of the above powder on a flat surface.
   1. Add 2 drops of urine to the powder.
   2. Ketone-positive urine will change color from lavender to purple in 30 seconds.

Abnormal Ketonuria

1. Uncontrolled diabetes.
2. Very low-carbohydrate diet.
3. Starvation.
4. Eating disorders, e.g., anorexia nervosa or bulimia.
5. Alcoholism.
6. Poisoning from drinking rubbing alcohol (isopropanol).
7. Ketones are often found in the urine during fasting for 18 hours or longer.
8. The sick person cannot eat or vomits for several days.
9. Low levels of ketones are sometimes found in the urine of healthy pregnant women.

Bilirubin in urine:

1. Bilirubin is a yellow-orange bile pigment compound formed when RBCs are hemolysed or ruptured.
2. The appearance of bilirubin in the urine indicates liver disease and is called bilirubinuria.
3. Bilirubin is a pigmented yellow compound, a degradation product of hemoglobin.
   1. It is detected long before jaundice clinically appears.
4. It is negative in normal urine.
   1. Negative in hemolytic jaundice.
5. Increased bilirubinuria is seen in the hepatic and obstructive jaundice.

Bilirubin metabolism and excretion in the urine

Conjugated bilirubin appears in the urine:

1. Conjugated bilirubin appears in the urine when the normal degradation cycle is disrupted by obstruction of the bile duct-like gall stones or cancer.
   1. Another possibility is when the liver cells are damaged and allow the leakage of bilirubin directly into the circulation.
   2. Hepatitis and cirrhosis are common causes of liver cell damage and result in bilirubinuria.
   3. Increased hemolysis does not produce bilirubinuria.
2. Bilirubin in the urine is detected only if the urine sample is fresh because bilirubin decomposes rapidly in bright light.
3. If the urine is left in the lab, then bilirubin is converted into biliverdin, a green compound by the light not detected by the bilirubin strips.

Clinical importance of the bilirubin in the urine (bilirubinuria):

1. Hepatitis (infectious).
2. Liver diseases.
3. Cirrhosis.
4. Gallbladder stones  (cholelithiasis) or cancer-causing obstruction.
5. Normal:
   1. Urine bilirubin is negative (0 to 0.2 mg/dL (0 to 0.34 µmol/L).
   2. Bilirubin can be detected in urine by the Foam test.

Procedure for the foam test:

1. Place a small volume of the urine in the test tube, cap it, and shake it vigorously.
2. Result:
   1. The foam at the top is white means bilirubin is negative.
   2. If the foam is orange, the color means bilirubin is present.
3. Other methods to detect bilirubin in the urine are:
   1. Diazo reaction in the form of tablets or Dipsticks.

Fouchet’s test (Ferric chloride test.): This is basically an oxidizing reaction.

Another modification of Fouchet’s test for urine bilirubin:

1. Composition of Fouchet’s reagent:
   1. Trichloracetic acid = 25 g
   2. Ferric chloride = 10% aqueous solution.
   3. Distle water = 100 mL
2. procedure: Take 10 mL of urine.
   1. Add 1 g barium chloride.
   2. Mix and filter.
   3. Leave the precipitate on filter paper and let it dry.
   4. Add Fouchet’s reagent or yellow nitric acid to the precipitate.

Comparison of Fouchet’s, Ferric chloride, and Diazo reaction for Bilirubin:

Urobilinogen in urine:

1. Urobilinogen is like bilirubin bile pigment, resulting from the hemoglobin degradation product.
2. Urobilinogen is a colorless compound derived from bilirubin. It is excreted into the bile and partially hydrogenated by bacteria in the intestine.
3. As shown in the above diagram, it is produced in the intestine by bacterial action on the bilirubin.
   1. The intestine reabsorbs approximately 50% of urobilinogen.
   2. Rest excreted in the feces.
   3. A small amount in enterohepatic circulation passed from the kidney and excreted in the urine as urobilin.
   4. This is why urobilinogen is positive normally in the urine.
4. Urobilirubin excreted from 0.5 to 4 mg /24 hours.
   1. <1 mg/dL is normally found in the urine.
   2. At least 0.1 mg/dL is needed for the detection.

Urine analysis:: Urobilinogen metabolism and excretion into stool and urine

How to check Urobilinogen in the urine:

1. The reagent strip for urobilinogen involves the Ehrlich aldehyde reaction. This will form the red azo dye.
2. This test must be done on the fresh sample because urobilinogen is an unstable compound and breakdown into urobilin on standing.

Significance of urobilinogen in the urine:

1. It helps in the early detection of liver diseases.
2. It is an increase in hemolytic disorder.
3. In hepatic jaundice, liver disorder, hepatitis, cirrhosis, and cancers.
4. It is absent in obstructive jaundice.

The normal level of urobilinogen:

Increased level of urobilinogen is seen in:

1. Hemolytic anemia.
2. Pernicious (megaloblstic) anemia.
3. Malarial attack.
4. Excessive bruising.
5. Pulmonary infarction.
6. Cirrhosis.
7. Acute hepatitis.
8. Cholangitis.

A decreased level of urobilinogen is seen in:

1. Complete or partial obstruction of the biliary tract.
2. Cholelithiasis.
3. Biliary duct inflammation.
4. Cancer of the head of the pancreas.
5. Antibiotic therapy will suppress intestinal bacterial flora.

The difference between Bilirubin and urobilinogen in the urine:

Microscopic Urine analysis:

Normal  microscopic findings:

1. Normally Very few or no red or white blood cells or casts are seen.
2. No bacteria, yeast cells, or parasites are present.
3. A few crystals are normally seen.

RBCs in urine:

1. The positive blood test indicates the presence of RBCs.

Hematuria:

1. It is a disorder of renal or genitourinary origin where the bleeding is due to trauma or damage to the organs.
2. The main causes are:
   1. Renal calculi.
   2. Glomerular diseases.
   3. Malignancies.
   4. Pyelonephritis.
   5. Trauma.

Hemoglobinuria:

1. This may result from the lysis of RBCs. There will be mixed hemoglobinuria and hematuria.
   1. There may be lysis of RBCs in the urinary tract in alkaline urine.
   2. In intravascular hemolysis, no RBCs are seen in the urine.

Myoglobinuria:

1. It is a heme-containing protein found in the muscles. This is seen in Rhabdomyolysis like :
   1. Trauma.
   2. Crush syndrome.
   3. Convulsions.
   4. Muscle-wasting disease.
   5. Heroin abuse.
   6. Prolonged come.
   7. Extensive exertion.

Red blood cells in the urine may be caused by the following:

1. Kidney or bladder injury.
2. Kidney stones.
3. Urinary tract infection (UTI).
4. Inflammation of the kidneys (glomerulonephritis).
5. Kidney or bladder tumor.
6. An autoimmune disease like systemic lupus erythematosus (SLE).

White blood cells in urine:

Pus cells:

Pus cells in the urine may be caused by :

1. Urinary tract infection.
2. Urinary bladder tumor.
3. Inflammation of the kidneys.
4. The Systemic lupus erythematosus (SLE).
5. Inflammation in the vagina or under the foreskin of the penis.

CAST in urine:

1. Urine sediment after the centrifugation shows the crystals and casts.
2. Casts are formed in the collecting tubules.
3. Various types of urine casts have different significance.
4. Depending upon the various reason, there are the following types of casts:
5. Granular cast:
   1. Granular casts appear homogenous, coarsely granular, and very dense.
   2. These casts are colorless.
   3. Granular casts are found in:
   4. Acute tubular necrosis.
   5. Pyelonephritis.
   6. Advanced glomerulonephritis.
6. Hyline cast:
   1. These are clear and colorless and form when the tubules contain precipitated proteins.
   2. Hyaline casts may be seen in physiologic states like strenuous exercise and mild renal disease.
   3. These are not associated with any particular disease.
   4. Hyaline casts are found in:
   5. Malignant hypertension.
   6. Chronic renal diseases.
   7. Diabetic nephropathy.
   8. Congestive heart failure.
   9. Pyelonephritis.
   10. Glomerulonephritis.
7. Cellular cast:
   1. These are WBC cell cast.
   2. WBC casts can originate from any place in the urogenital tract.
   3. WBC casts always come from the renal tubules.
   4. WBC casts are found in:
   5. Lupus nephritis.
   6. Acute glomerulonephritis.
   7. Pyelonephritis.
   8. Acute glomerulonephritis.
8. Epithelial cast:
   1. Renal epithelial cell casts formed from the shedding of the renal tubular cells.
   2. These shed cells undergo degenerative changes.
   3. These casts are first coarse and then become fine granular.
   4. Epithelial cell casts are found in:
   5. Acute tubular necrosis.
   6. Pyelonephritis.
   7. Acute glomerulonephritis.
   8. Viral infection like CMV.
   9. Drugs intoxication like salicylates overdose.
9. Waxy cast:
   1. Waxy casts formed from the degeneration of the granular cast.
   2. Waxy casts are found in:
   3. Nephrotic syndrome.
   4. Severe renal failure.
   5. Renal amyloidosis.
   6. Malignant hypertension.
   7. Diabetic nephropathy.

Crystals in urine:

1. Crystals in the urine are identified by their specific shapes.
2. These crystals are also identified by their solubility.
3. Mostly there are no symptoms due to these crystals.
4. Sometimes these crystals may be associated with renal stone formation, leading to obstruction.
5. The presence of large amounts of crystals or certain types of crystals indicates :
   1. Kidney stones.
   2. Damaged kidneys.
   3. Problems with metabolism.
   4. Medicines.
   5. Some types of urinary tract infections can also increase the number of crystals in the urine.

Bacteria in urine:

1. A urine nitrite test is done to confirm the bacteria present in the urine.
2. Nitrie is a rapid and indirect method of detecting urinary infections.
3. Gram-negative bacteria produce enzymes that reduce the nitrate into nitrite.
4. To check the bacterial infection, a nitrate test is advised:
   1. Pregnant women.
   2. School-going children, especially girls.
   3. Old people.
   4. Diabetic nephropathy.
   5. Patient with recurrent urinary infection.
   6. Mostly the infection starts from the urinary bladder and goes upwards to the kidneys.
5. Bacteria positive test in the urine mean a urinary tract infection (UTI).

Yeast cells or parasites

• Such as the parasite that causes trichomoniasis, which means an infection of the urinary tract.

Urine analysis summary

Normal urine picture:

Questions and answers:

Note: The details of urine microscopy are seen in urine part 2.