AUBF (Chapter 5) Chemical Examination of Urine PDF

ANALYSIS OF URINE AND BODY FLUIDS (CHAPTER 5: CHEMICAL EXAMINATION OF URINE) Outline Quality Control of Reagent Strips I. Reagent Strips II. Confirmatory Testing Run positive and...

ANALYSIS OF URINE AND BODY FLUIDS (CHAPTER 5: CHEMICAL EXAMINATION OF URINE) Outline Quality Control of Reagent Strips I. Reagent Strips II. Confirmatory Testing Run positive and negative control at least once per III. Urine pH 24 hours (at the beginning of each shift) IV. Urine Protein Run additional controls V. Microalbuminuria When a new bottle of strips is opened VI. Creatinine When results are questionable VII. Glucose When there are concerns over strip integrity VIII. Ketones Record control results IX. Blood X. Bilirubin Manufactured positive and negative controls are XI. Urobilinogen available XII. Nitrite Do not use distilled water as a negative control XIV. Leukocyte Esterase because reagent strip chemical reactions are XV. Specific Gravity designed to perform at ionic concentrations similar to urine Reagent Strip All negative control reading should be negative All positive control reading should agree with Provide a simple, rapid means of performing routine published control values chemical test on urine. Be aware of manufacturer-stated limitations and Single and multi test strips are available. interfering substances The brand and number of tests used are a matter of Correlate chemical readings to each other (physical laboratory preference and microscopic readings) Specified by urinalysis instrumentation manufacturers Reporting of Bacteria and Amorphous Crystals Strips consist of chemical-impregnated absorbent pads on a plastic strip Light A color-producing chemical reaction takes place Moderate when the absorbent pad comes in contact with Heavy urine Mucus Threads = Turbid The reactions are interpreted by comparing the color produced on the pad within the required time Confirmatory Testing frame with a chart supplied by the manufacturers Color comparison charts are supplied by the Confirmatory test use different reagents or manufacturer methodologies to detect the same substances as Several degrees of color are shown to provide reagent strips with the same or greater sensitivity or semiquantitative reading of negative, trace, 1+, 2+, specificity 3+, or 4+ can be reported Non-reagent strip testing procedures using tablets Estimate of mg/dL are also provided for many of the and liquid chemicals may be available when test areas questionable results are obtained Automated reagent strip readers also provide SI Chemical reliability of these procedures also must units be checked using positive and negative controls Reagent Strip Technique Urine pH Dip strip briefly into well-mixed specimen at room Lungs and kidneys are major regulators of the temperature acid-base content in the body Remove excess urine by touching edge of strip to A healthy individual produces a first morning container as strip is withdrawn specimen slightly acidic at 5.0 to 6.0 Blot edge of strip on an absorbent pad Postprandial specimen : more alkaline Wait for the specified amount of time Normal pH range : 4.5 to 8.0 Read using a good light source No absolute values are assigned Considerations include: Improper Technique Errors ➔ Acid-base content of blood ➔ Patient’s renal function Formed elements such as red and white blood cells ➔ Presence of UTI sink to the bottom of the specimen and will be ➔ Patient’s dietary intake undetected in an unmixed specimen. ➔ Age of the specimen A good light source is essential for accurate ❖ Standing urine: alkaline interpretation of color reactions The strip must be held close to the color chart A pH above 8.5 is associated with an aged or without actually being placed on the chart improperly preserved specimen, so a fresh specimen should be obtained Reagent strips and color charts from different manufacturers Clinical Significance of pH Specimens that have been refrigerated must be allowed to return to room temperature prior to Respiratory or metabolic acidosis/ketosis reagent strip testing, as the enzymatic reactions on the strips are temperature dependent Respiratory or metabolic alkalosis Aid in determining the existence of systemic Handling and Storing Reagent Strips acid-base disorders of metabolic or respiratory acidosis/alkalosis Store with desiccant in an opaque, tightly sealed Defect in renal tubular secretion and reabsorption container of acid and bases Remove strips immediately prior to use/testing ➔ Renal Tubular Acidosis Do not expose to volatile fumes Renal calculi formation ➔ Kidney stones Strips should be stored below 30oC ➔ Nephrolithiasis Do not use past the expiration date Treatment of UTI Visually inspect for discoloration/deterioration ➔ pH will tell you if a patient is taking UTI medications  Precipitation and identification of amorphous Prerenal Proteinuria materials ➔ Urates Conditions affecting the plasma not the kidney ➔ Phosphates Not indicative of actual renal disease Determination of unsatisfactory specimen Transient, increase levels of low-molecular-weight pH = 8.5 (reject) plasma proteins such as: ➔ Hemoglobin ➔ Myoglobin pH Crystals ➔ Acute phase reactants Rarely seen on reagent strips Caox ➔ ALBUMIN dumbbell-shaped (or x) The increased filtration of these proteins exceeds the normal reabsorptive capacity of the renal tubules, resulting in an overflow of the proteins into the urine Reagent strips detect primarily albumin, prerenal Triple phosphates proteinuria is usually not discovered in a routine (coffin-lid shape) urinalysis CRP - C-Reactive Protein hsCRP Ammonium biurate Alpha 1-antitrypsin (“thorn-apples”) Fibrinogen Ceruloplasmin Markers of Inflammation: pH-Reagent Strip Reactions Brand Multistix and Chemstrip measure urine pH Acute phase reactants units in 0.5- or 1-unit increments between pH 5 to 9 CRP - increase if there’s an inflammation ➔ Needed to measure between 5.0 to 9.0 in one half or one unit increments Bence Jones Protein (BJP) Use of double-indicator system (methyl red and Example of proteinuria due to increased serum bromothymol blue) to differentiates pH units protein levels is the excretion of Bence Jones Methyl Red: produces a color change from Protein by persons with multiple myeloma red/orange to yellow in pH range 4 to 6 Multiple myeloma, a proliferative disorder of the immunoglobulin producing plasma cells, the serum Bromothymol Blue: turns from green to blue in the contains markedly elevated levels of monoclonal pH range 6 to 9 immunoglobulin light chains (Bence Jones protein) Multiple myeloma confirmation Methyl red + H+ → Bromothymol blue - H+ ➔ Serum electrophoresis (Red/Orange → Yellow ) (Green → Blue) Renal Proteinuria Therefore, in the pH range 5 to 9 measured by the reagent strips, one sees colors progressing from 1. Glomerular or tubular damage orange at pH 5 through yellow and green to a final a. Glomerular proteinuria deep blue at pH 9 b. Microalbuminuria No known substances interfere with urinary pH c. Orthostatic Proteinuria measurements performed by reagent strips d. Tubular proteinuria Protein Proteinuria associated with true renal disease may be the result of either glomerular of tubular damage Most indicative of renal proteinuria ➔ Proteinuria seen in early renal disease a. Glomerular Proteinuria Normal urine contains very little protein Damage to glomerular membrane ➔ Less than 10 mg/dL Impaired selective filtration causes increased ➔ 100 mg per 24 hours protein filtration leading to cellular excretion Low-molecular-weight serum proteins are filtered; Abnormal substances deposit on the membrane many are reabsorbed ➔ Primarily immune disorders result in immune Albumin is primary protein of concern complex formation ➔ It holds water in place ❖ Lupus erythematosus ➔ Water in blood vessels will try to leak outside the ❖ Streptococcal glomerulonephritis (large blood vessel and go to tissue = edema immune complexes) Other proteins ➔ Amyloid material ➔ Vaginal, prostatic, and seminal ➔ Toxic substances ➔ Tamm Horsfall (Uromodulin) Benign proteinuria (transient) Clinical Significance ➔ Can be produced by conditions such as: ❖ Strenuous exercise Presence of protein requires determination whether ❖ High fever the protein represents a normal or a pathologic ❖ Dehydration condition ❖ Exposure to cold Clinical proteinuria = 30 mg/dL or 300 mg/L Increase in pressure on the filtration mechanism Proteinuria are varied and can be grouped into three ➔ Hypertension major categories: ➔ Strenuous exercise ➔ Prerenal ➔ Dehydration ➔ Renal ➔ Pregnancy ➔ Postrenal ❖ Preeclampsia ★ Increase in blood pressure ★ Complete bed rest b. Microalbuminuria Reaction Interference Micral Test Highly buffered alkaline urine overrides acid buffer Diabetic nephropathy with type 1 and type 2 diabetes system (color change unrelated to protein concentration) mellitus ➔ Leaving reagent pad in urine too long removes ➔ Reduced glomerular filtration buffer ➔ Eventual renal failure Also associated with an increased risk of cardiovascular disease False-positive c. Orthostatic (Postural) Proteinuria Highly pigmented urine High SG Quaternary ammonium compounds, detergents, Increase pressure on the renal vein when in the antiseptics, chlorhexidine vertical position Occurs in vertical position, disappears in horizontal position Sulfosalicylic Acid Precipitation Collection instructions ➔ Empty bladder before bed Confirmatory test for protein ➔ Collect specimen immediately on arising Cold precipitation test that reacts equally with all ❖ Negative reading will be seen on the forms of protein first morning specimen Must be performed on centrifuged specimens to ❖ Positive result will be found on the remove any extraneous contamination second specimen Microalbuminuria d. Tubular Proteinuria Semiquantitative testing for patients at risk for Tubular damage affecting reabsorptive ability renal disease ➔ Acute tubular necrosis Immunochemical assays for albumin or ❖ Toxic substance albumin-specific reagent strips ❖ Heavy metals Measure creatinine to produce an ❖ Viral infections albumin:creatinine ratio ❖ Fanconi syndrome (generalized proximal First morning specimens are recommended convoluted tubule defect) Amount of protein Micral Test ➔ Glomerular disorder: up to 4g/day ➔ Tubular disorders: much lower levels Gold-labeled antihuman antibody-enzyme conjugate Postrenal Proteinuria Dip strip in urine to marked level for 5 seconds Albumin binds to antibody Protein added in the lower urinary and genitourinary Bound and unbound conjugates move up strip tract Microbial infections causing inflammations and Unbound removed in captive zone containing release of interstitial fluid protein albumin;bound continues up strip Menstrual contamination Reaches enzyme substrate, reacts Semen/Prostatic fluid Colors from white (neg) to red (varying degrees) Vaginal secretions Compare color to chart Traumatic injury Results read from 0 to 10 mg/dL Protein-Reagent Strip Reactions Immunodip Test Traditional principle Immunochromatographic technique ➔ Protein error of indicators Specially designed container for strip ➔ Certain indicators change color in the presence Place container in controlled amount of specimen of protein at a constant pH for 3 min, urine enters container ➔ Protein accepts H+ from the indicator, Albumin binds to blue latex particles coated with increased sensitivity to albumin due to more antihuman albumin antibody amino groups to accepts H+ than other proteins Bound and unbound migrate up strip Tetrabromophenol blue or tetrachlorophenol Unbound encounters area of immobilized albumin on tetrabromo sulfonephthalein and an acid buffer strip — forms blue band pH level 3 both indicators are yellow Bound continues migrating to an area of immobilized Color progresses through green to blue antibody and forms blue band Report: Color of band is compared with chart Negative 30 mg/dL Reagent Strip Microalbumin Test Trace 100 mg/dL Commercially available strips 1+ 3+ 300 mg/dL 1. Clinitek microalbumin reagent strips 2+ 4+ 2000 mg/dL 2. Multistix Pro reagent strips Simultaneous measurement of albumin and Trace values are