Clinical Urinalysis Review

1. Clinical Urinalysis Review Austin Community College Medical Laboratory Technology Clinical II Spring 07

2. Urine Blood Testing • http://library.med.utah.edu/WebPath/TUTORIAL/URINE/URINE.html

3. Chemical Exam of Urine

4. Chemical Exam of Urine

5. Chemical Exam of Urine • Reagent strip manufactures • Bayer Corporation- Diagnostics Division (formerly Ames) produces Multistix • Boehringer-Mannheim Corporation which produces Chemstrip • Behring Diagnostics which produces Rapignost

6. Chemical Exam of Urine

7. Chemical Exam of Urine • Reagent strip precautions and source of errors • Normal dipstick procedure: • Dip strip briefly, but completely into well mixed, room temperature urine sample. • Withdraw strip, blot briefly on its side • Keeping the strip flat, read results at the appropriate times by comparing the color to the appropriate color on the chart provided.

8. Chemical Exam of Urine • Sources of error (& preventions) • Testing cold specimens • would result in a slowing down of reactions; test specimens when fresh or bring them to RT before testing • Inadequate mixing of specimen • could result in false reduced or negative reactions to blood and leukocyte tests; mix specimens well before dipping • Over-dipping of reagent strip • will result in leaching of reagents out of pads; briefly, but completely dip the reagent strip into the urine

9. Chemical Exam of Urine • Inadequate blotting & Failure to keep strip horizontal • will result in over-run or mixing of reagents between the different reaction pads; blot excess urine off the strip and keep strip horizontal. If dipping from the tube, can run the side of the strip along the rim to remove excess urine. • Improper timing of tests • over development of reagent pad colors leading to falsely increased results; follow manufacturer’s recommendations

10. Chemical Exam of Urine • Inadequate light • misinterpretation of results; use good lighting • Mis-using color chart • misinterpretation of results; hold strip just over color chart and match colors as close as possible, consider use of back-up tests, if needed, especially if urine’s color masks reaction colors.

11. Chemical Exam of Urine • Handling and Storage • Keep strips in original container, stored at RT • Protect from moisture and volatile fumes • Use before expiration date • Do not touch reagent pad areas

12. Chemical Exam of Urine • Quality Control - use appropriate, commercially prepared positive and negative controls. • Use commercially prepared pos and neg controls, at least once per 24 hours, and anytime a new bottle is opened, or question of validity of results. Readings should agree with published results ± one color block.

13. Urine Glucose Testing • Normal : no glucose detected • Clinical significance of abnormal results (Glucosuria) • Plasma glucose level exceeds renal threshold (160-189 mg/dL) • Diabetes mellitus • Renal tubular dysfunction • Filtered glucose not being reabsorbed in tubules

14. Urine Glucose Testing • Dipstick Testing Method • Glucose initiates reaction • Coupled reaction • Glucose oxidase – oxidizes glucose to gluconic acid and concurrently reduces oxygen to hydrogen peroxide. • Hydrogen peroxide in presence of the enzyme peroxidase will oxidize an indicator, giving a colored reaction. • Chromogens • Potassium iodide or • Tetramethylbenzindine

15. Urine Glucose Testing • Sensitivity • @ 50-100 mg (compared to Clinitest’s 250) SO- Can have a positive dipstick but a neg Clinitest • Specificity - is specific for glucose only. • not affected by other sugars or reducing substances.

16. Urine Glucose Testing • Interfering substances • High specific gravity and high pH may depress color. • Ascorbic acid-false neg • Bleach or peroxide may give false positive

17. Urine Bilirubin Testing • Normal : no bilirubin detected • Clinical significance of abnormal results (Bilirubinuria) • Jaundice - Condition when serum bilirubin becomes greater than the liver can handle, and there is an abnormal collection of bilirubin in the tissues giving them a yellow color

18. Urine Bilirubin Testing • Prehepatic / Hemolytic jaundice • Excessive hemolysis of RBC; beyond what the liver can process • Type of bilirubin? • Is bilirubin found in the urine? YES/NO? Explain.

19. Urine Bilirubin Testing • Prehepatic / Hemolytic jaundice • Type of bilirubin? – indirect, insoluble, unconjugated • Is bilirubin found in the urine? – No, the bilirubin is not water soluble

20. Urine Bilirubin Testing • Hepatic jaundice • Liver’s cells malfunctioning • Ie. viral hepatitis, cirrhosis etc. • Both (direct) bilirubin and urobilinogen found in urine.

21. Urine Bilirubin Testing • Post hepatic (regurgative or obstructive) hepatitis • Obstruction to outflow of bile – some type of blockage • Gall stones • Tumor • Edema • Conjugated bilirubin backed up into blood (Bilirubinuria) and passes into urine

22. Urine Bilirubin Testing • Testing method • Urine dipsticks for bilirubin – a diazo reaction • Impregnated with stabilized diazotized 2,4 dichloraniline • Color goes from buff to brown also shades of pink – violet • If urine is strongly colored, look for change in pad color after dipping. Use Ictotest for backup.

23. Urine Bilirubin Testing • Interfering substances • Medication metabolites, pigments and indican may obscure readings • False negatives due to aged specimens, especially those exposed to light and oxidation.

24. Urine Ketone Testing • Ketone Bodies • Origin - not normally present • Products of fat catabolism - breakdown of fat into CO2 and H2O • What are the 3 ketone bodies?

25. Urine Ketone Testing • Acetone • 2%. -Acetone is volatile, & excreted primarily through the lungs • Diacetic Acid (Acetoacetic) • the first formed, • 20 % of the total • the form detected by most ketone test procedures • Beta hydroxybutyric Acid • majority formed, but not detected by routine tests. • Only Hart’s test, an old ‘wet chemical’ test will detect this one.

26. Urine Ketone Testing • Definitions • Ketonuria - ketones in the urine • Ketonemia - ketones in the blood • Ketosis - disease state, when patient has increased amount of ketones. • Acidosis - state when blood pH is decreased, an accumulation of acids; commonly occurs as a result of ketosis

27. Urine Ketone Testing • Clinical significance • Health – formed in liver and completely metabolized • Disease – excessive formation and accumulation • Disturbance of carbohydrate metabolism • when there is a decrease of carbohydrate metabolism, then the body stores of fat must be metabolized to supply energy. • As a result of this increased fat metabolism ketones will be found in the urine. Ex. low carbohydrate diets, diabetes • Starvation • Vomiting and diarrhea in children • Van Gierke's Disease – glycogen storage disease • High fat diet

28. Urine Ketone Testing • Clinical effects • Metabolic acidosis • Lowering of blood & urine pH • Brain toxicity

29. Urine Ketone Testing • Testing • most use nitroprusside • detects diacetic acid and a small amount of acetone, but does not detect β-hydroxybutyric acid. • Produces purple color • Can be used on urine or blood

30. Urine Specific Gravity Testing • The specific gravity is a measure of the weight of urine compared to an equal amount of water.  • Specific gravity it proportional to urine osmolality which is a measure of concentration.

31. Urine Specific Gravity Testing • The specific gravity will always be greater than 1.000 and will increase as more materials are dissolved in the urine.  • The value changes throughout the day depending on fluid intake.

32. Urine Specific Gravity Testing • Specific gravity between 1.002 & 1.035 on a random sample is normal if kidney function is normal. • Specific gravity in Bowman’s capsule fluid is @ 1.007 • Any reading below this indicates hydration • Any reading above this indicates some degree of dehydration

33. Urine Specific Gravity Testing • Again dilute urine will have values less than 1.010.  • Fixed specific gravity = 1.010; isothenuria • Diabetes insipidus • End-stage renal disease • And concentrated urine will have values usually over 1.020. • Usually due to dehydration and can be seen in well population as well as sick.

34. Urine Specific Gravity Testing • Increased urine specific gravity may indicate / be seen in: • * Dehydration • * Diarrhea • * Excessive sweating • * Glucosuria • * Heart failure (related to decreased blood flow to the kidneys) • * Renal arterial stenosis • * Syndrome of inappropriate antidiuretic hormone secretion (SIADH) • * Vomiting • * Water restriction

35. Urine Specific Gravity Testing • Decreased urine specific gravity may indicate / be seen in: • * Excessive fluid intake • * Diabetes insipidus – central or nephrogenic • * Renal failure (that is, loss of ability to reabsorb water) • * Pyelonephritis

36. Urine Specific Gravity Testing • Specific gravity > 1.035 (refractometer) • Could have very high glucose levels • Could contain radiographic dye

37. Urine Specific Gravity Testing • Testing • Polyelectrolytes , pH indicator (bromthymol blue measures the pH change), and alkaline buffer

38. Urine Specific Gravity Testing • Interfering substances • False elevation of results may be seen in samples with increased protein concentration. • Some reports of reduced specific gravity results on alkaline specimens. • Lipids may also effect results

39. Urine Blood Testing • Normally not found in urine • Hemoglobinuria – free hemoglobin in urine • Circulating free hemoglobin normally picked up by haptoglobin preventing loss in urine • When serum levels of hemoglobin > 100 mg/dL threshold is exceeded • Hematuria – RBCs in the urine • Trauma / irritation of renal organs

40. Urine Blood Testing

41. Urine Blood Testing • Testing dipstick reaction

42. Urine Blood Testing • ‘Blood’ test detects • Free Hemoglobin • RBCs – get lysed on the pad & their hemoglobin reacts • Myoglobin – muscle hemoglobin • Principle based on the peroxidase-like activity of the heme portion of the molecule

43. Urine Blood Testing • Sensitivity – can detect at levels of 5-10 cells/uL • Interfering substances • Ascorbic acid • Nitrates • Oxidizing agents (ie bleach) • Contaminate blood (menstrual)

44. Urine pH Testing • Normal: kidneys capable of 4.5 – 8.0 • Factors effecting pH • Diet – general & specific foods • Time of day • Metabolic disorders • Drugs / medications • Dipstick capable: 4.5 – 9.0

45. Urine pH Testing • Test method • Dipstick indicators – methyl red and bromthymol blue • Range 4.5-9.0 • Caution – other chemicals on dipstick can effect pH reading

46. Urine Protein Testing • Normally not found in measurable amounts on dipstick (<150 mg/dL /day) • Permeability of glomerulus • Damage to glom capularies • Changes in glom blood flow • Albumin excretions may be increased temporarily due to exercise, uti, and acute illness with fever. • Dipstick results of >@ 1+ (30mg/dL) would equal to @ 500 mg/dL (clinical proteinuria)

47. Urine Protein Testing • Only albumin detectable by dipstick • Sensitivity (@15-30 ml/dL)

48. Urine Protein Testing • New testing for microalbumin & creatinine • Results: • Protein 20-200 mg/dL (30-300 mg/dL /24 hr) • Creatinine 10-300 mg/dL • Albumin/creatinine ratio • Normally albumin in the urine is less than 30 mg/ gram creatinine

49. Urine Protein Testing • Principle - Protein error of indicators • at fixed pH, certain indicators show one color in the presence of protein and another in absence of protein - the “error” of the indicator. • Indicator – tetrabromphenol blue - can be hard to read at the trace end • Citrate Buffer – maintains pH 3 -quite acid

50. Urine Protein Testing • Sources of error • Sensitive only to albumin • Urine with strong / unusual color makes reading difficult • Highly alkaline or buffered urine will neutralize acid buffer and lead to increased erroneous results. • Urine container contamination would interfere