PATHOLOGICAL OR ABNORMAL URINE CONSTITUENTS I- Proteinuria : - Proteinuria is a condition in which urine contains an abnormal amount of protein. - The majority of proteins found in the urine arise from the blood. - As blood passes through healthy kidneys, they filter the waste products out and leave in the things the body needs, like proteins. Most proteins are too big to pass through the kidneys' filters into the urine unless the kidneys are damaged. The main protein that is most likely to appear in urine is albumin. Proteins from the blood can escape into the urine when the filters of the kidney, called glomeruli, are damaged. Sometimes the term albuminuria is used when a urine test detects albumin specifically.
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PATHOLOGICAL OR ABNORMAL URINE
CONSTITUENTS
I- Proteinuria :
- Proteinuria is a condition in which urine contains an abnormal amount of protein.
- The majority of proteins found in the urine arise from the blood.
- As blood passes through healthy kidneys, they filter the waste products out and
leave in the things the body needs, like proteins. Most proteins are too big to pass
through the kidneys' filters into the urine unless the kidneys are damaged. The main
protein that is most likely to appear in urine is albumin. Proteins from the blood can
escape into the urine when the filters of the kidney, called glomeruli, are damaged.
Sometimes the term albuminuria is used when a urine test detects albumin
specifically.
- Large amounts of protein in the urine may cause it to look foamy. Also, because the
protein has left the body, the blood can no longer soak up enough fluid leading to
swelling in the hands, feet, abdomen, or face. These are signs of very large protein
loss.
- In a healthy renal and urinary tract system, the urine contains no protein or only
traces amounts.
- Because albumin is filtered more readily than the globulins, it is usually abundant in
pathological conditions. Therefore, the term albuminuria is ~~~= proteinuria.
- The qualitative tests for proteinuria are most commonly performed using a reagent
test strip. These methods depend on the change in the response of an indicator dye in
the presence of protein. (See the figure)
- Quantitative examination of urine protein requires considerable attention to
the volume or time of urine collection because the concentration may vary with
time and volume.
- Most assays are performed on urine sample of 12-24h.
- Reference Value (RV) for 24-h urine Male = 1-4 mg/dl
Female = 3-10mg/dl
Child = 1-10mg/dl
-Reference value – Qualitative Normal = Negative
Qualitative Tests for Proteins
-Before performing a test for albumin, the urine should be filtered or centrifuged.
1 - Heat Coagulation Test :
Fill a test tube one third full of urine and gently heat the upper half of the fluid to
boiling, being careful that this fluid does not mix with the lower half which serves as
a control. Turbidity indicates proteins or calcium or magnesium phosphate. Acidify
the urine slightly by the addition of 3-5 drops of dilute acetic acid where upon the
turbidity if due to phosphate will disappear.
2 - Heller's Test :
1 -Place 3ml of conc. HNO3 in test tube.
2 -Tie the tube to about a 30 degree angle. Using a pipette allow the urine to flow
slowly down the side of the tube.
3 -If albumin is present, within a few minutes a white ring will form at the
junction between the two liquids.
II- Glucosuria:
-Glucosuria is a condition in which urine contains an abnormal amount of glucose.
-Glucose is present in glomerular filtrate and is reabsorbed by the proximal tubule.
- If the blood glucose level exceeds the reabsorption capacity of the tubules, glucose
will appear in the urine.
-Tubular reabsorption of glucose is by active transport in response to the body's
need to maintain an adequate concentration of glucose .
-The blood level at which tubular reabsorption stops is termed renal threshold
which for glucose is between 160-180 mg/ dl .
-Glucosuria occurs in diabetes mellitus, which characterized by abnormal blood
glucose level (hyperglycemia) and usually increased volume of urine. The urine may
be light in color and have a high specific gravity .
Qualitative Tests for Glucose
-Before testing for sugar the urine should be free of albumin. If present, it must be
removed by heat coagulation. Firstly, acidify the urine with dilute acetic acid, boil,
cool and filter .
Principles of Fehling' and Benedict's reagent :
I- Fehling's Test
-In this test the presence of aldehydes but not ketones is detected by reduction of
the deep blue solution of copper (II) to a red precipitate of insoluble copper oxide.
The test is commonly used for reducing sugars but is known to be NOT specific for
aldehydes. For example, fructose gives a positive test with Fehling's solution as does
acetone.
Two solutions are required:
-Fehling's "A" uses 7 g CuSO4.5H2O dissolved in distilled water containing 2 drops
of dilute sulfuric acid.
-Fehling's "B" uses 35g of potassium tartrate and 12g of NaOH in 100 ml of
distilled water.
-These two solutions should be stoppered and stored until needed.
Method:
-Mix 1ml of solution-"A" with 1ml of solution-"B"
-Add 1 ml of this mixture to an empty test tube.
- Add 1ml of the urine in test tube. If the sample is from patient the sample must
first boil l to liberate ammonia which may have a destructive effect on the reagent.
-Mix urine with Fehling's reagent .
-Place the tube in a water-bath at 60°C .
-A positive test is indicated by a green suspension and a red precipitate.
-The test is sensitive enough that even 1 mg of glucose will produce the
characteristic red colour of the compound.
Copper-tartrate-complex
II- Benedict's reagent
- Also called Benedict's solution or Benedict's test is a reagent used as a test for the
presence of reducing sugars (such as glucose, lactose, and fructose, but not sucrose),
or more generally for the presence of aldehydes, in a solution.
- Benedict's reagent contains blue copper(II) sulfate (Cu S O 4) · 5H2O which is
reduced to red copper(I) oxide by aldehydes, thus oxidizing the aldehydes to
conc. ammonia (1-2ml) =====>>>>mix ===>>>> Deep permanganate color after
few minutes indicate the presence of acetone.
-NV – random sample -Qualitative Normal = Negative
IV- Hematuria and Hemoglobinuria :
- Hematuria is the presence of red blood cells (RBCs) in the urine. In microscopic
hematuria, the urine appears normal to the naked eye, but examination under a
microscope shows a high number of RBCs (see the figure). Gross hematuria can be
seen with the naked eye—the urine is red or the color of cola.
- Several conditions can cause hematuria, most of them not serious. For example,
exercise may cause hematuria that goes away in 24 hours. Many people have
hematuria without any other related problems. Often no specific cause can be
found. But because hematuria may be the result of a tumor or other serious
problem, a doctor should be consulted.
To find the cause of hematuria, or to rule out certain causes, the doctor may order a
series of tests, including urinalysis, blood tests or others.
- Urinalysis is the examination of urine for various cells and chemicals. In addition
to finding RBCs, the doctor may find white blood cells that signal a urinary tract
infection or casts (groups of cells molded together in the shape of the kidneys' tiny
filtering tubes) that signal kidney disease. Excessive protein in the urine also signals
kidney disease.
- Blood tests may reveal kidney disease if the blood contains high levels of wastes
that the kidneys are supposed to remove.
Hemoglobinuria: results from hemolysis i.e. the rupturing of erythrocyte and the
liberation of hemoglobin. This may occur in malaria, typhoid, yellow fever,
hemolytic jaundice and other diseases.
Benzidine reaction:
- This reaction depends on the Fe ion found in hemoglobin.
Method:
- 0.5ml urine + 1.5ml benzidine, then add 0.5ml H2O2 >>>>>> Blue color.
V- Bilirubin (Bile):
- Bilirubin is the waste product that results from the breakdown of hemoglobin
molecules from worn out red blood cells. Ordinarily, it is excreted from the body as
the chief component of bile. Excessive levels of bilirubin stain the fatty tissues in the
skin yellow; this condition is called jaundice.
- Bilirubin is found in the dark urine of obstructive jaundice and later in the course
of hepatitis. Urobilinogen is found in early hepatitis. It is colorless but darkens on
standing. Bile pigments are included in the routine strip used in urinalysis. Positive
results are an indication for measuring s. bilirubin and liver enzymes.
Test for bile pigments:
Method:
- 1ml urine + 5 drops 1% iodine in alcohol are added >>>>>> Green ring indicates
the presence of bile
pigments.
Note: In all clinical labs qualitative urinalysis depends on using dipstick (strip)
to determine the normal and abnormal constituents of urine (see the figure).
RESULTS
-You are supplied with the sample of urine, carry on routine UA .
-Detect the presence of any pathological constituents.
-According to what you studied in the lab, is this urine normal or not?
-Present your results in a good and full lab report .
III- Microscopic Examination
- A sample of well-mixed urine (usually 10-15 ml) is centrifuged in a test tube
at relatively low speed (about 2-3,000 r.p.m) for 5-10 minutes until a moderately
cohesive button is produced at the bottom of the tube.
- The supernatant is decanted and a volume of 0.2 to 0.5 ml is left inside the
tube.
- The sediment is resuspended in the remaining supernatant by flicking the
bottom of the tube several times.
- A drop of resuspended sediment is poured onto a glass slide and cover-
slipped.
- The sediment is first examined under low power to identify most crystals,
casts, squamous cells, and other large objects. Next, examination is carried out
at high power to identify crystals, cells, and bacteria.
Examination
The sediment is first examined under low power to identify most crystals, casts,
squamous cells, and other large objects. Next, examination is carried out at high
power to identify crystals, cells, and bacteria.
White Blood Cells
Pyuria refers to the presence of abnormal numbers of leukocytes that may appear with
infection in either the upper or lower urinary tract or with acute glomerulonephritis.
Usually, the WBC's are granulocytes. White cells from the vagina, especially in the
presence of vaginal and cervical infections, or the external urethral meatus in men and
women may contaminate the urine.
Epithelial Cells
Renal tubular epithelial cells, usually larger than granulocytes, contain a large round or oval nucleus and normally slough into the urine in small numbers. However, with
nephrotic syndrome and in conditions leading to tubular degeneration, the number sloughed is increased.
Casts
Urinary casts are formed only in the distal convoluted tubule (DCT) or the collecting duct
(distal nephron). The proximal convoluted tubule (PCT) and loop of Henle are not
locations for cast formation. Hyaline casts are composed primarily of a mucoprotein
(Tamm-Horsfall protein) secreted by tubule cells. The Tamm-Horsfall protein secretion
(green dots) is illustrated in the diagram below, forming a hyaline cast in the collecting
duct:
Red blood cells may stick together and form red blood cell casts. Such casts are
indicative of glomerulonephritis, with leakage of RBC's from glomeruli, or severe
tubular damage.
White blood cell casts are most typical for acute pyelonephritis, but they may also
be present with glomerulonephritis. Their presence indicates inflammation of the
kidney, because such casts will not form except in the kidney.
Bacteria
Bacteria are common in urine specimens because of the abundant normal microbial
flora of the vagina or external urethra and because of their ability to rapidly
multiply in urine standing at room temperature. Therefore, microbial organisms
found in all but the most scrupulously collected urines should be interpreted in view
of clinical symptoms.
Diagnosis of bacteriuria in a case of suspected urinary tract infection requires
culture.
Yeast
Yeast cells may be contaminants or represent a true yeast infection. They are often
difficult to distinguish from red cells and amorphous crystals but are distinguished
by their tendency to bud. Most often they are Candida, which may colonize bladder,
urethra, or vagina.
Crystals
Common crystals seen even in healthy patients include calcium oxalate, triple phosphate crystals and amorphous phosphates.
Very uncommon crystals include: cystine crystals in urine of neonates with
congenital cystinuria or severe liver disease, tyrosine crystals with congenital
tyrosinosis or marked liver impairment, or leucine crystals in patients with severe