Lab 10 Routine Urine Analysis Microscopic Analysis of

Lab (10): Routine Urine Analysis Microscopic Analysis of Urine KAU-Faculty of Science- Biochemistry department Clinical biochemistry lab (Bioc 416) 2012 Lecturer: Nouf Alshareef nf. shareef@hotmail. com

Microscopic Analysis of Urine Crystals Casts Cells • • • RBCs WBCs Epithelial cells Bacteria Yeast

• • • Microscopic analysis. In this test, urine is spun in a special machine (centrifuge) so the solid materials (sediment) settle at the bottom. The sediment is spread on a slide and looked at under a microscope. Things that may be seen on the slide include: Red or white blood cells. Blood cells are not found in urine normally. Inflammation, disease, or injury to the kidneys, ureters, bladder, or urethra can cause blood in urine. Strenuous exercise, such as running a marathon, can also cause blood in the urine. White blood cells may be a sign of infection or kidney disease. Casts. Some types of kidney disease can cause plugs of material (called casts) to form in tiny tubes in the kidneys. The casts then get flushed out in the urine. Casts can be made of red or white blood cells, waxy or fatty substances, or protein. The type of cast in the urine can help show what type of kidney disease may be present. Crystals. Healthy people often have only a few crystals in their urine. A large number of crystals, or certain types of crystals, may mean kidney stones are present or there is a problem with how the body is using food (metabolism). Bacteria, yeast cells, or parasites. There are no bacteria, yeast cells, or parasites in urine normally. If these are present, it can mean you have an infection. Squamous cells. The presence of squamous cells may mean that the sample is not as pure as it needs to be. These cells do not mean there is a medical problem, but your doctor may ask that you give another urine sample.

1 - Crystals • Not usually present in fresh urine • Appear if urine is left for several hours before examination. • Type of crystal depends on: p. H and constituents of urine (conc. ) • Diagnostically significant crystals: § if present in high conc. for long time • May indicate the presence of: § metabolic disorder (congenital disorder or sever liver disease) § renal calculi formation (stones) § provide information that can be used to regulate medications

Identification of urine crystals • Can identified by morphology alone but … • Knowing urine p. H is important (provide supporting information)

Urinary Crystals Normal crystals (common) • Found in healthy individual • Include: q q q Uric acid, Calcium oxalate Amorphous urates or phosphates Triple phosphate Ammonium biurate Calcium carbonate. Pathological crystals (uncommon) • Abnormally found in urine • Due to certain disorders • Include: q q q Amino acid crystals Bilirubin crystals Cholesterol

Normal Crystals “most common” 1. Normal Acid Urine 2. Normal Alkaline Uric acid Triple phosphate Calcium oxalate Amorphous phosphates Amorphous urates Calcium oxalate

Crystals in Normal Acid Urine 1 - Oxalate crystals • occur in urine at any p. H. Calcium oxalate crystals are square shape with a characteristic “X” mark

Crystals in Normal Acid Urine 2 - Uric acid crystals uric acid crystals: lemon or star needle shaped Note: • Presence of high conc. Of Uric acid crystal may indicates Gout

Crystals in Normal Acid Urine 3 - Amorphous urates crystals • aggregates of finely granular material without any defining shape. • (Na, K, Mg, or Ca salts) urate • Has a pink precipitate of urate salts

Crystals in Normal Alkaline Urine 1 - Phosphate crystals • • found if urine p. H > 6. 5 Its formation depends on ammonia conc. usually associated with bacterial growth. can indicate urinary tract infection.

Crystals in Normal Alkaline Urine 2 - Amorphous phosphates • aggregates of finely granular material without any defining shape. • Has a white precipitate of calcium phosphate • Alkaline p. H of urine may be due to: 1 - Caused by diet (vegetarian, rich in phosphates) 2 - Pathology

Pathological Crystals “most common “ 1 - Cystine crystals cystine crystals are hexagon in shape 3 - Bilirubin Crystals 2 - Tyrosine Crystals:

Differentiation between amorphous urates and amorphous phosphates : • Urine p. H • Centrifugation

3 - Urinary Casts

Urinary Casts • They are cylindrical structures found in urine and composed mainly of mucoprotein (Tamm-Horsefall mucoprotein) which is secreted by epithelial cells of kidney tubules Why these proteins are precipitated? • factors responsible are not fully understood • but may relate to the concentration and p. H of urine in these areas. • Some casts may contain cells (epithelial cells, RBC, WBC) so, • Casts can be made up of Protein, lipid, cells or mixed.

• If casts are present in large numbers, the urine is almost sure to be positive for albumin.

Types of Urinary cast • • Hyaline Cellular Granular Waxy casts • They represent different stages of degeneration of cells in a cast. Clinical indications • Casts in the urine always indicate some form of kidney disorder.

1 - Hyaline casts • Is the most common type of cast • They are solidified Tamm-Horsfall mucoprotein secreted from the tubular epithelial cells of each nephron Causes: • Most commonly seen in normal individuals due to: – Dehydration, low urine flow, concentrated urine, or acidic urine – or vigorous exercise. Morphology: • Hyaline casts are cylindrical and clear, colorless, homogeneous, transparent, and usually have rounded ends.

Hyaline casts

2 - Cellular casts Red blood cell casts • Aggregation of RBCs • Always pathological. • Found in urine due to: Glomerular damage, or urinary tract injury (bleeding) (Glomerular damage allows RBCs to pass through glomerular membrane) Red blood cell cast in urine

White blood cell casts • Indicate inflammation or infection of the kidnies White blood cell cast in urine This renal tubular cell cast suggests injury to the tubular epithelium. http: //library. med. utah. edu/Web. Path/TUTORIAL/URINE. html#6

Epithelial cell casts • Is rarely seen in urine because renal disease that affects the tubules is infrequent. Found in urine due to: • Degeneration of nephron tubule due to acute tubular necrosis and toxic ingestion • distinguished from WBCs by its large size, round nuclei

3 - Granular casts • The second-most common type of cast • result either from the breakdown of cellular casts or the inclusion of aggregates of coagulated proteins Causes: • Indicates significant chronic renal disease • can also be seen for a short time following exercise Morphology: • contain fine granules may appear grey or pale yellow in color or appear black because of the density of the granules.

Casts which persist may break down, so that the cells forming it are degenerated into granular debris, as has occurred in this granular cast.

4 - Waxy casts • represent the end product (final step) of cast degeneration • result from the degeneration of granular casts. Causes: • usually indicate renal failure. • malignant hypertension and diabetic Morphology: • larger than hyaline casts, more rigid, sharp edges, fractures, and broken-off ends. • appear yellow, grey, or colorless.

This is a broad, waxy cast. Note that the edges are sharp and there are "cracks" in this cast.

(A) Hyaline cast (C) leukocyte cast granular cast B) erythrocyte cast

3 - Cells Microscopic analysis • RBCs: RBCs WBCs

1 - Epithelial cells • Little amount normally appear in female due to reproductive period.

2 - RBCs: • RBCs up to 5/HPF are commonly accepted as normal. • High RBCs in urine is called hematuria, may be due to: - hemorrhage, - inflammation, nephratitis - necrosis, - trauma, or neoplasia in urinary tract.

3 - WBCs • presence of WBCs suggests infection. • WBC up to 5/HPF are commonly accepted as normal. • Higher numbers (pyuria) indicate presence of inflammation somewhere in urinary tract.

4 - Bacteria and yeast • Bacteria are the commonest organism seen • commonly due to contaminants. Bacteria yeast

Others Sperm Schistosoma & Hematobium Tricochomonas vaginal

Exercises:

Oxalate

Phosphate

Phosphate crystals

Microscopic Examination: • Urine sedimentation may contain cells, casts and crystals and is examined microscopically after centrifugation of a urine sample. • A very small amount of all of the above sediments is normal. • Concern begins when any of these components is significantly elevated.

Procedure: 1. centrifuged well-mixed urine 10 ml at 3000 r. p. m for 10 min until precipitate. 2. supernatant is decanted and 0. 2 -0. 5 ml is left inside the tube. 3. The sediment is resuspended in the remaining supernatant. 4. drop of sediment is poured in slide and cover-slipped. 5. The sediment is first examined under low power to identify most crystals, casts, cells and other large objects. 6. Next, examination is carried out at high power to identify crystals, cells, and bacteria.