Introduction to Urinalysis PDF

Introduction to Urinalysis MLT 108 Goal of Effective Quality Assessment Program To obtain consistently accurate and reproducible results Test results should reflect patient’s condition, and not be modified due to procedural or personnel variations An action has not been pe...

Introduction to Urinalysis MLT 108 Goal of Effective Quality Assessment Program To obtain consistently accurate and reproducible results Test results should reflect patient’s condition, and not be modified due to procedural or personnel variations An action has not been performed if it has not been documented – lab techs must document EVERYTHING  Biological Hazards Standard Precautions and transmission-based precautions Established in 1996 All body fluids, secretions, and excretions are considered potentially infectious and capable of disease transmission Infection prevention practices are applied to all patients in healthcare settings Appropriate PPE Proper disinfectant Spill kits Post-exposure treatment  Global Harmonization Program Safety Data Sheets (SDS) Universal symbols understood by all cultures & languages Safety Data Sheet (SDS)  QA - Preanalytical Components Includes all procedures occurring before testing: Specimen collection, transport, storage, and handling Patient education, preparation, and instructions Procedures for handling inappropriate or unacceptable specimens Documentation of any problems or unusual situations Accurate, up-to-date, and available procedure manuals Adequate personnel training and supervision Proper labeling: Always label the urine cup, not the lid The lid comes off and can be switched out causing mislabels The cup contains the sample and cannot be exchanged Proper Mixing: Always mix urine samples before performing any test to unsure a homogenous solution  Renal Medical Terminology Kidneys nephr/o, ren/o Renal Pelvis pyel/o Glomerulus glomerul/o Urine ur/o, urin/o Ureters ureter/o Urinary Bladder cyst/o Urethra urethr/o Stone -lith  Where does urine come from? The average adult has around 6 liters of blood The kidneys filter the Plasma volume (about 3 liters) around 60 times a day This is almost 200 liters a day! The kidneys take water and waste products out of the blood and send it to the bladder for storage until excretion Nerve impulse triggers urination once the bladder receives around 150 mL Urine is a protein-free ultrafiltrate of plasma  Functions of the Kidneys Maintains homeostasis through the balance of water, salts, and acids Water is mostly reabsorbed in the Distal Convoluted Tubule Sodium reabsorbed at 65% in the Proximal Convoluted Tubule, 25% in the Loop of Henle Kidneys reabsorb bicarbonate (HCO3−) and secrete hydrogen (H+) to maintain acid-base equilibrium of the blood Removal of urea, creatinine, uric acid, urochrome, and other wastes Creatinine and Urea are exclusively excreted by kidneys, specific to kidney function Remaining urea in the blood can be measured via the Blood Urea Nitrogen (BUN) Maintain blood pressure, erythrocyte production, activation of Vitamin D, and production of hormones 65% 25%  Regulation of Blood pH Between 7.35 and 7.45 3 body systems involved in the maintenance of pH: 1. Blood-Bicarbonate Buffer System Buffers prevent pH from changing 2. Pulmonary System Lungs exhale or retain carbon dioxide 3. Renal System Increases or Decreases the excretion of Hydrogen (H+) ions Increases or Decreases the formation of Ammonia (NH3) Increases or Decreases the reabsorption of Bicarbonate (HCO3−)  Hormones in the Urinary System Liver produces Angiotensinogen Kidneys produce Renin which splits Angiotensinogen into Angiotensin I Renin production triggered by low blood pressure, low Na+/K+, hemorrhage Lungs and the Kidneys produce Angiotensin Converting Enzyme (ACE) which converts Angiotensin I to Angiotensin II Angiotensin II has many functions: Stimulates Aldosterone from the Adrenal Glands Aldosterone causes the kidneys to retain Sodium and Water Constricts blood vessels Triggers thirst from the Hypothalamus Stimulates the release of Antidiuretic Hormone (vasopressin) from the Pituitary Gland (ADH is produced in the Hypothalamus, released from the Posterior Pituitary Gland) All of these functions help control blood pressure and thus filtration rate  Nephrons Functional units of the kidneys About 1.3 million nephrons in each kidney Site of urine production through filtration, reabsorption, and secretion Contain Glomeruli (clusters of capillaries) where filtration takes place 99% of the total filtered volume is reabsorbed Renal artery supplies blood to the nephrons Renal vein carries filtered blood from the kidney back into the blood  Urine Volumes and Terminology Normal urine production is 600-1,800 mL Enuresis - involuntary discharge of urine Anuria - absence of urine formation Dysuria - difficult / painful urination Oliguria - less than 400 mL per day Urinary Hesitancy - difficulty in starting Diuresis - increased output of urine urinary stream Polyuria - excessive urination Urinary Retention - inability to completely empty the bladder which can lead to stones and UTIs Incontinence - inability to control excretion of wastes  Renal Failure Terminology Azotemia – elevated levels of nonprotein nitrogen compounds in the blood, usually urea, caused by: congestive heart failure, hemorrhage, dehydration, renal failure, glomerulonephritis, stones, or prostatitis Uremia – toxic levels of urea in the blood Acute Renal Failure – sudden onset of uremia which may become fatal Chronic Kidney Disease – progressive loss of renal function End-Stage Renal Disease – final stage of chronic kidney disease requiring dialysis or transplant, usually from Protein and/or Glucose Nephritis - inflammation of the kidneys Glomerulonephritis - inflammation of the glomeruli, usually caused by infection (Strep throat) or autoimmune (lupus or Goodpasture's syndrome) Pyelonephritis – inflammation of the renal pelvis, usually caused by urinary tract infections or sexually transmitted infections (pelvic inflammatory disease - PID) Renal Failure leads to Hyperproteinuria and Hypoproteinemia  Stones A stone is also known as a calculus Stones are caused by stationary minerals Nephrolith - a stone located in the kidney Ureterolith - a stone located in the ureter Cystolith - a stone located in the bladder Shockwave Lithotripsy - breaking of stones into fragments using x-rays Percutaneous Nephrolithotomy - surgical removal of nephrolith through a small opening in the back  History of Urinalysis Clay tablets show the first documented evaluations of urine to be over 6,000 years old The Pisse Prophecy, written by the Pisse prophets, correlated urine color, smell, and clarity to a diagnosis Early practitioners would taste their patient’s urine samples The invention of the microscope in the late 1500’s added microscopy to the evaluation of urine In 1956, the chemical dipstick allowed for quick, concise evaluations In 1982, the KOVA system made microscopic urinalysis more consistent The 1990s brought automation to the urinalysis department  Significance of Urinalysis Urine collection is usually non-invasive – even less so than blood collection Urine samples are easily obtainable as the average, hydrated person urinates around 7 times a day Urinalysis is a quick, cheap, and informative laboratory test Urine samples are a great screening test for a wide range of diseases: urinary tract infection, yeast infection, kidney disease, liver disease, diabetes, dehydration, sexually transmitted infections, kidney stones, kidney transplant rejection, blood transfusion reactions, multiple myeloma, ketoacidosis, crush injuries, glomerulonephritis, pyelonephritis, cancer, Fanconi’s syndrome, cystinosis and cystinuria, maple syrup urine disease, phenylketonuria, porphyrias, pelvic inflammatory disease, lupus, Goodpasture's syndrome  Urinary Laboratory Tests Urinalysis – the physical, chemical, and microscopic analysis of a urine sample for diagnostic purposes – requires at least 12 mL of urine Urine Osmolality – number of solute particles in the urine, often compared to the serum osmolality Urine Sodium – for Congestive Heart Failure or Renal Failure Serum / Urine Electrophoresis – separation of proteins Glomerular Filtration Rate – volume of plasma processed per minute by the Glomeruli Serum Creatinine – creatinine level in the blood, assesses GFR, more specific to the kidney than BUN Blood Urea Nitrogen (BUN) – waste product remaining in the blood, Urea is an end product of protein breakdown (protein -> ammonia -> urea), excess is either caused by an increase in protein break down, or a decrease in kidney function Blood pH – important when evaluating renal issues / renal (metabolic) compensation Total Protein / Creatinine Ratio – excess urine protein can detect renal stress Creatinine Clearance - calculation based on urine creatinine, serum creatinine, body mass, and urine output  Review The kidneys produce urine (ultrafiltrate) by filtering the blood, removing waste, controlling blood pressure, controlling electrolytes Water is reabsorbed by the Distal Convoluted Tubule Sodium is reabsorbed by the Proximal Convoluted Tubule Kidneys help maintain a 7.35 - 7.45 blood pH Creatinine and Urea are exclusively excreted by the kidneys Liver produces Angiotensinogen, Kidneys produce Renin ACE (lungs and kidneys) converts Angiotensin I to Angiotensin II Nephrons are the functional unit at about 2.5 million Normal urine production is 600 – 1,800 mL Always label the urine cup, not the lid Urinalysis includes physical, chemical, and microscopic Urinalysis is a great screening test for many diseases because it is quick, cheap, and informative Osmolality MLT 108  Osmolality The concentration of solute particles in a solution Affected by solute number only, not size or weight Osmolality = φ * n * C φ is the osmotic coefficient, n is the number of ions per molecule, and C is the concentration in moles per kilogram of solvent The osmotic coefficient accounts for molecules that are not ions Can be measured through 4 colligative properties of a solution: Osmotic Pressure, Vapor Pressure, Boiling Point, or Freezing Point Depression Clinical Osmometers use either freezing point depression (most common) or vapor pressure and provide units milliosmoles/kilogram of water The higher the osmolality, the more concentrated the solution (more solutes)  Freezing Point Depression Sample is added to the osmometer chamber which contains a wire needle and a thermometer Sample is cooled below its freezing point (supercooled) Vigorous agitation (the freeze pulse) disrupts the supercooled solution resulting in crystallization and heat of fusion The heat of fusion causes a rise in temperature that plateaus The plateau represents the true freezing point of the solution, which is proportionate to the osmolality The lower the plateau, the higher the osmolality / more concentrated Freezing Point Depression  Osmolality of Urine and Serum Urine osmolality helps determine renal concentrating ability – especially after 10 hours of fluid deprivation – and can detect early signs of renal disease Serum osmolality differentiates many diseases – especially involving Na+ The 3 most prevalent solutes in a normal urine are: Urea, Chloride, and Sodium The 3 most prevalent solutes in normal serum are: Sodium (about 90%), Glucose, and BUN Urine osmolality should be 1 to 3 times that of Serum Osmoreceptors in the Hypothalamus trigger the thirst sensation and produce antidiuretic hormone to decrease osmolality as needed  Calculated Osmolality and Osmol Gap The Calculated Serum Osmolality includes only the 3 normal, predominant solutes found in the Serum 2 equations may be used for the Calculated Serum Osmolality (2*Na) + (Glucose/20) + (BUN/3) less accurate, easy to remember (1.86*Na) + (Glucose/18) + (BUN/2.8) + 9 more accurate, harder to remember Osmol Gap is the difference between the measured serum osmolality (from the Osmometer) and the calculated serum osmolality A large Osmol Gap is usually caused by alcohol, lactate, or ketones The Osmol Gap is used in toxicology – especially for alcohol poisoning Note: alcohol may be measured if the automated methodology is not Freezing Point Depression (because alcohol does not freeze)  Remember, the EtOH (alcohol) is only included in the measured osmolality if it was performed using a methodology other than Freezing Point Depression, which is the most common methodology  Normal Ranges Random Urine Osmolality 50 – 1,300 mOsm/kg Urine Osmolality after 10 Greater than 850 mOsm/kg hours of Fluid Deprivation Serum Osmolality 275 – 300 mOsm/kg Urine/Serum Ratio 1.0 – 3.0 Serum Osmol Gap 5 – 10 mOsm/kg  Comparing Urine and Serum Osmolality Note the wide range for random urine osmolality (50-1,300 mOsm/kg) This is because the concentration of urine varies greatly throughout the day secondary to diet, water intake, and exercise It is very important to collect the urine and serum sample at the same time for compassion It is also preferred to have fasting samples (urine osmo after 10 hours of fluid deprivation)  Disease States – Increased Serum Osmolality Diabetes Mellitus (hyperglycemia) Renal Tubular Disorder Severe Burns Primary Aldosteronism (Hyperaldosteronism) Dehydration Sepsis Alcohol Poisoning Uremia (glomerulonephritis, polycystic kidney disease, end-stage renal) Profuse Sweating (hyperthyroidism, menopause, obesity, Parkinson’s disease, heart failure, tuberculosis infection, medications) Note: most of these disease states are caused by Hypernatremia (high sodium levels in the blood)  Disease States – Decreased Serum Osmolality Potassium Deficiency Overuse of Diuretics Ketonuria Severe Burns Renal Failure Congestive Heart Failure Cystic Fibrosis with increased sweat Nephrotic Syndrome (losing unfiltered solutes in the urine) Hepatic Cirrhosis (decreased albumin production) Prolonged Vomiting and/or Diarrhea (gastrointestinal infections, eating disorders) Note: most of these disease states are caused by Hyponatremia (low sodium levels in the blood)  The Connection Between Serum Sodium Sodium makes up about 90% of the serum osmolality Around 99.5% of the serum sodium is reabsorbed by healthy kidneys (mostly in the Proximal Convoluted Tubule) This is over a pound of salt being reabsorbed, and less than 2 grams of salt being lost per day (replaced by the diet) Various disease states that affect blood pressure, blood volume, albumin levels, antidiuretic hormone, and renal reabsorption will ultimately affect salt levels which changes the serum osmolality value  Disease States – Diabetes Insipidus Diabetes insipidus results in polyuria & extreme thirst Caused by a lack of antidiuretic hormone, or resistance to antidiuretic hormone Lack of ADH may be caused by the hypothalamus (production) or the posterior pituitary (secretion) This type will actually have a normal urine osmolality and ratio Resistance to ADH is known as Nephrogenic DI This type causes a decreased urine osmolality and borderline low ratio  Panic / Alert / Critical Values Serum Osmolality less than 240 mOsm/kg Serum Osmolality greater than 322 mOsm/kg Serum Osmolality greater than 395 mOsm/kg puts the patient at risk for grand mal seizures Values greater than 420 mOsm/kg are lethal  Other Uses for Freezing Point Depression The Freezing Point Depression methodology can be compared to other methodologies – or even the Calculated Serum Osmolality – to screen for antifreeze poisoning in suicide and homicide patients Freezing Point Depression is also used in pharmaceuticals, in vitro fertilization, and as a QA check for beverages, chemicals, and waste water treatment  Advanced Instruments 3320 Uses Freezing Point Depression 20uL sample size 60 second testing time 290 mOsm/kg Standard 2 or 3 levels of QC for both Serum and Urine 3 Calibrators 2000 mOsm/kg 850 mOsm/kg 50 mOsm/kg Advanced Instruments 3320 Procedure Remove chamber cleaner Add a tip to the pipette Aspirate 20uL of sample (don’t forget to mix the urine sample first) Wipe the pipette tip to remove excess Insert the pipette on the blue track Press the Start button on the Osmometer Slide the blue track to insert the pipette Wait 60 seconds, record the result Remove the pipette and clean the chamber  Review Osmolality is the concentration of solute particles in a solution Osmolality is not affected by solute size or weight Freezing Point Depression is the most common methodology in the clinical lab Units are milliosmoles/kilogram of water (mOm/kg) 10-hour fluid deprivation samples are preferred Osmol Gap is the difference between the measured serum osmolality and the calculated serum osmolality The calculated serum osmolality is based on Sodium, Glucose, and BUN A large Osmol Gap (above 10) is usually caused by alcohol, lactate, or ketones Many disease states affect urine osmolality, serum osmolality, and/or the ratio Serum Sodium levels (Hypernatremia and Hyponatremia) can significantly change a patient's osmolality as 90% of the serum osmolality is caused by Sodium ions Most Osmometers require Calibrators, Standards, and multiple levels of Controls Specific Gravity MLT 108  Applications of Specific Gravity Wine and Beer Servers and Computers The alcohol by volume (ABV) level can be To assess the purity of immersion cooling solutions determined by measuring the density before and after the fermentation process (a type of liquid cooling in which the hardware is Agriculture directly submerged into a non-conductive liquid) Engineering The specific gravity of the soil is important in determining its potential crop yield Aids in material selection when weight is a concern Construction Especially for bridges and skyscrapers Manufacturing Determining the concentration of cement mixtures before pouring Used as a quality assurance check for many solutions Geology such as glue or paint (dilution may be required) Differentiating and calculating the Clinical Laboratory composition of rocks and gems Hospital labs measure the Specific Gravity of urine to assess kidney function and/or patient hydration  Specific Gravity Often abbreviated SG The ratio in weight of a given solution to an equal volume of water at a specified temperature Comparison of the density from one liquid to that of pure water (mass of all solutes present per volume of solution) Depends on the number and size of solutes in the solution There are no units because it is a ratio The linearity of Specific Gravity is 1.000 (pure water) to 1.999 However, most instruments cannot read above 1.040 The greater the density/concentration, the higher the Specific Gravity  Solutes Affecting Specific Gravity Glucose White Blood Cells Protein Bacteria Cells Bilirubin Crystals Red Blood Cells Casts Salt is not listed as it has a small impact on SG, unlike osmolality This is because SG is influenced by size and weight of solutes Osmolality is based on the number of solutes only This, in part, is why we rarely test serum Specific Gravity  Refractometry Indirect method based on the refractive index of light When light passes from air into a solution at an angle, it refracts and slows the direction of the beam The ratio of light refraction in two differing mediums is called the refractive index As the number of solutes increases, light velocity decreases, and the angle decreases This can easily be measured via a Refractometer Using a Refractometer  Normal Urine Specific Gravity The Normal Specific Gravity of a random urine sample is between 1.005 and 1.030 depending on hydration, diet, and exercise The more hydrated a patient is, the more dilute their urine sample is, and the lower their Specific Gravity result (closer to water which is 1.000) The less hydrated a patient is, the more concentrated their urine sample is, and the higher their Specific Gravity result (further away from that of pure water) Because of this connection, we can use the Specific Gravity to assess a normal, healthy patient’s hydration status Because the normal range is so dynamic, physicians often request a fluid deprivation sample, or corelate the SG to other urine results + signs and symptoms  Decreased Specific Gravity Results Low SG is known as hyposthenuria and may be caused by: Nephrogenic Diabetes Insipidus (resistance to ADH, increased water output) Pyelonephritis (inflammation of the renal pelvis) Renal Failure (inability to concentration urine) Acute Tubular Necrosis (inability to concentration urine) Preanalytical Complications Excess exposure to light (bilirubin break down) Delay in processing time (cell degradation) Exposure to extreme temperatures (protein degradation)  Elevated Specific Gravity Results Increased SG is known as hypersthenuria and may be caused by: Diabetes Mellitus (increased glucose in the urine) Urinary Tract Infection (falsely elevated by the bacteria cells in the urine sample) Heart Failure (decreased blood flow to the kidney, concentrated urine) Diarrhea/Vomiting (fluid loss) Excessive Sweating (fluid loss) Early Stages Of Renal Damage (protein/glucose escaping the nephrons)  Consistent Specific Gravity Results When the kidneys start to fail, they lose their ability to concentrate urine Although this may be confirmed by a fluid deprivation Osmolality test, the Specific Gravity may be a screening test (other UA results to correlate) If a patient is unable to concentrate urine secondary to renal disease, their urine will consistently have a Specific Gravity equal to that of the plasma ultrafiltrate which is 1.010 This repetitive SG of 1.010, usually seen at ESRD, is termed isosthenuric or isosthenuria  Is it Urine? Patients trying to pass a Urine Drug Test may water down their urine sample to generate false negatives for drug metabolites A Specific Gravity below 1.003 is questionable, and is probably not urine The sample should be recollected until a higher SG is obtained  Specific Gravity Dilutions If the Specific Gravity of a solution exceeds the readable limit of the Refractometer being used, a dilution with DI water must be made The dilution factor is multiplied by the values proceeding the decimal place Example: if 1mL of pure water and 1mL of solution were mixed together, the dilution factor would be 2 (1 over 1+1 = 1/2) If the Specific Gravity from the dilution was 1.030, then we would multiply the numbers after the decimal place (.030) by dilution factor 2 to get 1.060 Example #2: 2mL pure water, 1mL urine would give a dilution factor of 3 If the Specific Gravity obtained was 1.031, the corrected SG would be 1.093  SG Calibration and Standards Most Refractometers have a calibration screw which adjusts the angle in which the light bends within the eyepiece Because DI water has a SG of 1.000, we can use it as a Standard to Calibrate the instrument Additional Standards may also be purchased from reagent manufacturers  SG Quality Controls At least 2 levels of QC are ran at least once a day on the Refractometer If multiple instruments are used, all Refractometers should be QC’d “Home-brewed” controls include a 5% salt solution in DI water which should yield a SG of 1.022 +/- 0.001, and a 9% sucrose solution in DI water which should yield a SG of 1.034 +/- 0.001 These home-made Quality Controls must be validated by another means Reagent-grade QC may be purchased, in which case the acceptable range would be provided by the manufacture Most manufactures offer a “Normal” and “Abnormal” urine control which may be used for osmolality, SG, Pregnancy, Drug Testing, and Urinalysis  Other Methodologies The Urinalysis Dipstick, the Chemical portion of the 3-part Urinalysis, now includes a test pad for Specific Gravity Because of this, the manual testing of SG on a Refractometer is often used as a backup methodology in modern clinical labs The test pad on the dipstick changes from Blue to shades of either Green or Yellow based on the ionization of polyelectrolytes within the test pad which results in a pH change from alkaline to acidic The intensity of the pH change is proportionate to the color change and can be compared to a scale for interpretation  Review Specific Gravity & Osmolality are both indicative of a solution's concentration compared to DI water Osmolality is based on solute number only, SG is influenced by the size and weight of the solutes Urine osmolality is often compared to serum osmolality, SG is not Osmolality easily detects Sodium imbalance (hypernatremia and hyponatremia), SG does not Specific Gravity is a great test for evaluating dehydration, kidney function, and the presence of cells Hyposthenuria , Hypersthenuria , Isosthenuric 1.010 Refractometers utilize the refractive index of light The calibration screw on the Refractometer is adjusted to correlate with Standard(s) Dilutions are made with DI water, and the result is multiplied by the dilution factor At least 2 levels of QC are ran daily for each Refractometer The Chemical portion of a Urinalysis often includes a SG pad which has mostly replaced the Refractometer Urine Collection MLT 108  Urine Containers Urine containers should be: Clean Dry Leak-proof Disposable Easy to open and close Sterile (if used for cultures) Large enough to support the minimum volume of 12 mL  Sample Rejection Urine samples should be rejected if they are: Not labeled Improperly labeled (missing info / lid instead of cup labeled) Leaking Insufficient in volume (less than 12 mL) Contaminated with stool, toilet paper, cleansing pad, etc. Not properly transported Protected from light Temperature Regulated Transported or preserved within 1 hour  Sample Rejection Always document rejected samples If it isn’t documented, then it didn’t happen Most hospitals want the lab to: call the patient’s nurse, document the conversation via a comment in the computer system, reorder the test, and send the new label Example: “Unlabeled urine cup was sent to the lab at 0927 on 05/14/2025 from ED West. Lab called Kayla Smith, RN at 0931 who stated the sample was from patient 0248669. New label (accession number 14-915278-12) was sent to ED West at 0935 by Derrick, MLT.” Don’t feel bad about rejecting urine samples as the average person voids multiple times a day, and obtaining a new sample is usually quick, easy, and noninvasive Making a nurse mad for an hour is better than killing a patient!  Importance of Transport Time Urine samples that are not sent to the lab within 1 hour will: change color, clarity, odor, and pH have falsely decreased/negative glucose, ketones, bilirubin, urobilinogen have increased bacterial growth have degraded red blood cells, white blood cells, parasites, crystals, and casts have altered Osmolality and Specific Gravity These changes are caused by bacterial growth and/or solute degradation/alteration Any of these things may drastically change the patient’s diagnosis  Unable to Meet the Transport Time If the transport time of 1 hour is not possible, the sample should be preserved before transport Refrigeration (2-8oC) is the easiest, most common form of preservation Chemical preservatives may be used for certain tests / if refrigeration is not an option Boric Acid – used for Urine Cultures, cannot be used for drugs or hormones Formalin – used to preserve cells and casts, cannot be used for Chemical UA Sodium Fluoride – used for urine drug screens, cannot be used for Chemical UA  Urine Culture Tubes (Boric Acid) If a patient has a Urinary Tract Infection, the doctor needs to know which bacteria species is causing the infection This is important as different antibiotics work for different species Boric Acid is the most common preservative for Urine Cultures (the growth of bacteria from a urine sample to determine the species causing infection) This is because Boric Acid prevents bacterial growth and metabolism  Urine Specimen Types Random Timed Most common sample Collected over/at a specified time of day, or timed alongside a specific procedure Collected at anytime without restrictions For quantitative urine testing, comparisons, For routine screening daily estimates, or hormone cycles Affected by excess fluid intake or exercise May be used to compare to the 24-hour sample First Morning / Fluid Deprivation 24-Hour Preferred sample All voids in the same container for 24 hours Most concentrated Must contain only 1 First Morning sample, Better chance of detecting trace Protein preferably the latter for best recovery of cells, Not as common because inconvenient crystals, and casts For quantitative or “clearance” testing - especially Total Protein, Creatinine, & Glucose  24-Hour Urine Samples All voids collected in a calibrated Urine Jug for a 24-hour period Should contain one First Morning sample at the end of the 24-hour cycle Patient should be given very detailed instructions for collection Sample should be refrigerated or placed on ice throughout the 24 hours After verifying orders, labels, and patient identification, the very first thing the lab should do is notate the Total Volume of the Urine Jug The Total Volume is necessary for many calculations, and cannot be measured after a sample is poured off After documenting Total Volume, the next step is to fully mix the sample Some tests require centrifugation to concentrate the sample, or to prevent contamination/interference from cells  Examples of 24-Hour Urine Tests Creatinine Cortisol – Cushing’s Syndrome Glucose – Gestational Diabetes Total Protein Cystine Level - Cystinuria and Cystinosis Protein/Creatinine Ratio Sodium – Congestive Heart Failure and Renal Failure Albumin Urine Urea Nitrogen – detect Nitrogen imbalances Albumin/Creatinine Ratio Creatine – elevated in both the Urine and the Serum Protein Electrophoresis in Muscular Dystrophy and Hyperthyroidism Uric Acid – evaluates kidney function Elevated leads to Gout, Decreased mean Liver Disease  24-Hour Urine Creatinine Creatinine is formed from Creatine and Creatine Phosphate from muscle metabolism, and is excreted at a constant rate based on muscle mass Plasma Creatinine is inversely related to Glomerular Filtration Rate (GFR) Creatinine should account for only about 5% of the Non-protein Nitrogen in both the Urine and the Serum Creatinine is a better indicator of renal function than BUN or uric acid Total Creatinine is tested for via Spectrophotometery using the Jaffe Reaction Remember to document the total volume, mix the sample, aliquot, and centrifuge  24-Hour Urine Creatinine Clearance Volume of plasma (in mL) processed per minute by the glomeruli is called the Glomerular Filtration Rate (GFR) Creatinine Clearance is the average clearance rate of creatinine at the glomeruli per day Calculated from: minutes/day, body mass, Urine Creatinine, and the Serum Creatinine Normal: Males 97-137 mL / minute / 1.73m2 Females 88-128 mL / minute / 1.73m2 Elevated concentrations associated with abnormal renal function, especially at the Glomeruli (24-Hour Urine Creatinine x 24-Hour Total Volume) 1.73 m2 X (Serum Creatinine x 1440 minutes) Body Surface Area There are 1440 minutes in a day 1.73 m2 is the ideal body surface area, we divide this by the (24 hours x 60 minutes = 1440) patient’s actual body surface area to get a corrected value  24-Hour Urine Total Protein Record total volume, mix sample, pour off an aliquot, and centrifuge Increased levels may be associated with uncontrolled diabetes, high blood pressure, lupus, or preeclampsia Protein/Creatinine Ratio may help differentiate the diagnosis 24-Hour Urine Albumin may be ordered to differentiate Albumin from other proteins Trace amounts of Albumin, often called “Prealbumin” is usually indicative of diabetes Protein Electrophoresis may be ordered to differentiate all other proteins  Preeclampsia Pathology Preeclampsia is a condition of hypertension during gestation Preeclampsia is associated with high blood pressure, edema, and proteinuria The mother releases stress signals and increases blood pressure to provide enough nutrients to the baby (especially if the spiral arteries are not appropriately dilated) These changes damage the Endothelial Cells within the blood vessels Once the Endothelial Cells in the Nephrons get damaged, Protein slips through If untreated, can lead to Eclampsia which may result in stillbirth, seizures, coma  Preeclampsia Lab Results Greater than 1+ positive reaction for Protein on the UA Chemical Dipstick Greater than 300 mg of Total Protein from the 24-Hour Urine Greater than 0.3 mg/dL Creatinine/Protein Ratio from the 24-Hour Urine Mild Hemolytic Anemia Low Platelet Count Elevated Liver Enzymes in the Serum (especially ALT and AST) May lead to DIC  Urine Protein Electrophoresis Protein in the Urine is abnormal, but figuring out which Protein(s) are present can be diagnostic, this information is obtained through Urine Protein Electrophoresis (UPEP) UPEP should be performed with a Serum Electrophoresis (SPEP) for comparison UPEP can be performed on a Random, First Morning, or 24-Hour sample First Morning and 24-Hour samples are preferred Sample should be mixed prior to testing Samples should be centrifuged to concentrate the Proteins (minimum volume of 12 mL) Samples are added to agarose gel which is then shocked with electrical current The proteins migrate various lengths based on their size and charge The presence/absence of proteins are noted in addition to a semi-quantitative value  Urine Protein Electrophoresis Patterns Multiple Myeloma – huge Gamma protein spike in both the Serum and the Urine Tubular Necrosis – decrease in Gamma proteins in the Serum, all proteins present in the Urine with Gamma proteins dominating Diabetes – normal Serum, trace Albumin in the Urine Nephrotic Syndrome – decreased Albumin and Alpha 2 proteins in the Serum, all proteins in Urine Hypogammaglobulinemia – decreased Gamma proteins in the Serum, no proteins in the Urine  24-Hour Urine Cortisol – Cushing’s Syndrome Cortisol is a lipid-soluble hormone responsible for many functions Cushing’s Syndrome is an Endocrine disorder resulting in elevated levels of cortisol Cushing's Disease is a specific type of Cushing’s Syndrome caused by an adenoma of the pituitary gland producing excess ACTH which in turn leads to increased cortisol levels Results in weight gain, a fatty hump between the shoulders (buffalo hump), stretch marks, bruises, high blood pressure, bone loss, muscle breakdown, hyperglycemia, and, eventually, type 2 diabetes Random Plasma Cortisol levels are not beneficial, & Timed Plasma Cortisol levels – between 11PM and midnight – are not a viable option for outpatient clinics 24-Hour Urine Cortisol is a great and convenient test for diagnosing Cushing’s Syndrome After documenting total volume, an aliquot should be poured off and spun down before testing 24-Hour Urine Cortisol is typically tested for using Tandem Mass Spectrophotometry (TMS)  Glucose and Gestational Diabetes Gestational Diabetes is a temporary form of Diabetes that takes place during pregnancy Gestational Diabetes may be screened for by Serum Glucose, Random Urine, or A1C Gestational Diabetes may be confirmed by Fasting Serum Glucose, Glucose Tolerance Test (GTT), Timed Urine, and/or 24-Hour Urine The 24-Hour Urine provides 2 vital pieces of information – Urine Volume (polyuria) and Total Urine Glucose – an aliquot should be poured off and centrifuged before testing A Timed Urine may be collected to correlate with the 24-Hour Urine or the GTT  24-Hour Urine Cystine Cystinuria Autosomal Recessive defect in the amino acid transport system (SLC3A1 or SLC7A9 gene) Amino Acids are Nitrogen-containing amines with a carboxylic acid functional group DNA is transcribed to mRNA, the mRNA nucleotides (codons) bind to tRNA (anticodons) which carry amino acids (translation), the amino acids bind together by peptide bonds to build polypeptides and eventually proteins after a Stop Codon Because of the inherited defect, excess amounts of amino acids Cystine, Arginine, Lysine, and Ornithine are produced by the body Cystine, unlike the others, is not water soluble and therefore starts to accumulate in the kidneys/bladder ultimately forming Cystine Crystals and Cystine Kidney Stones After documenting total volume, an aliquot should be poured off & centrifuged before testing  24-Hour Urine Cystine Cystinosis Caused by 100+ different Autosomal Recessive mutations of the CTNS gene The CTNS gene codes for the production of Cystinosin which is a transporter protein responsible for removing Cystine from lysosomes (lysosomes hold oligopeptides which are broken down into amino acids) Nephropathic (also known as Infantile) Cystinosis is the most common type, patients often develop Fanconi Syndrome in which vital nutrients are lost in the urine Cystinosis is often more severe than Cystinuria and may cause buildup of cystine crystals in the eyes and white blood cells in addition to the urine  Managing Cystinuria and Cystinosis Limiting Salt and Animal Proteins from the diet helps decrease oligopeptides Staying hydrated helps dilute out the urine and prevent stones from forming Medications to increase urine pH, increase cystine solubility, or cleave cystine from the body Genetic counseling to understand the risks of passing the gene on Shockwave lithotripsy or surgical removal of stones as needed  Urine Collection Types Midstream Clean-catch Most common, requires cleaning of the genitalia Suprapubic Aspiration Removal of urine from the bladder using a needle Catheterized Catheter tubing from the urethra to the bladder Bagged Collections External bag attached to the skin in order to collect urine  Midstream Clean-Catch Most common sample Requires cleansing pads, patient instructions, and a sterile container The glans penis and/or urethral meatus must be thoroughly cleaned with cleansing pads prior to collection – “clean-catch” After cleaning, initial urination should be discarded in the toilet, the middle portion of the void should be collected into a sterile cup, and the final volume discarded into the toilet – “midstream” The cleansing pad prevents normal skin flora from entering the sample Collecting just the midstream portion prevents normal flora, epithelial cells, and sperm cells of the urethra from entering the sample This process is needed for bacterial cultures and to prevent vaginal / sperm contamination The patient MUST properly collect the sample for valid results – important to educate the patient Midstream Clean-Catch  Suprapubic Aspiration Sterile collection by medical personnel Puncture of abdominal wall and bladder by using a needle attached to a syringe Sample aspirated directly from the bladder Mostly used for infants or adults that cannot void or be catheterized secondary to blockage Suprapubic Aspiration and Missing the Target When collecting urine via suprapubic aspiration, there is always the possibility of missing the bladder and extracting something other than urine, usually abdominal fluid To ensure the sample is urine, the lab can run a Creatinine level (waste product of protein breakdown) Creatinine concentrations are around 50 times higher in the urine compared every other fluid in the body Most suprapubic aspirations are now ultrasound guided to prevent missing  Catheterized Specimen Sterile, plastic tubing inserted through the urethra into the bladder Urine flows directly into the collection container from the catheter tubing Used when voiding is painful, not possible, or not incomplete (stones, surgery, unconscious, medications, enlarged prostate, urinary retention)  Types of Catheters Indwelling Catheter (brand name Foley Catheter) (2 or 3 way) is used in clinical settings by healthcare professionals and remain inside the patient for weeks to months Intermittent Catheter (also known as Straight Catheter) is designed to be used only once, and may be administered by the patient themselves after instruction External Catheter (Condom Catheter) are for males only, attach to the head of the penis, and are non invasive collecting devices Suprapubic Catheter is inserted into the abdomen instead of the urethra Although catheters are sterile and should be inserted using sterile technique, once contaminated, catheters may become a source of urinary tract infections – especially if the port site is not cleaned before and after, if the tip touches a nonsterile surface, or if the patient is immunocompromised In which case, the tip of the urine catheter or a sterile sample from the catheter itself (and not the bag) may be sent to the lab for bacterial culture  Bagged Collections Mostly used on infants / pediatrics Plastic urine collection bag attached with a hypoallergenic skin adhesive External genitalia should be cleaned prior to attachment Collects urine externally, noninvasive Specimen should be removed soon after collection Appropriate for routine testing only  Review Urine samples should be dry, leak-proof, disposable, 12+mL, and sterile Urine samples should be tested within 1 hour or preserved for best results Refrigeration is the most common preservation, Boric Acid (gray top) for Culture Random Urine is most common for convenience, First Morning is ideal & most concentrated 24-Hour Urine Samples are great for Clearance testing and Renal Function Note the Total Volume of 24-Hour Urine Jugs, mix, pour off, and centrifuge if needed Creatinine Clearance requires a calculation and is tested via Spec using Jaffe Reaction TP, Albumin, and UPEP are useful in differentiating proteinuria disease states Cystine Crystals (hexagon) are seen in Cystinuria and Cystinosis Midstream Clean-Catch is the most common collection type and requires patient instructions Physical Urinalysis MLT 108  Urinalysis Urinalysis is the evaluation of urine for screening and diagnostic purposes There are 3 parts to a Urinalysis: Physical, Chemical, and Microscopic It is important to correlate results between each portion of the urinalysis, as well as other lab tests and patient signs and symptoms The Physical properties of urine include: Color Clarity Specific Gravity* Osmolality Foam Odor Remember, before testing urine samples for any analyte, it must be a well-mixed, homogenous solution  Normal Urine Color Normal color is Yellow due to urochrome Urochrome is a lipid-soluble pigment in plasma excreted at a constant rate Dark yellow Concentrated urine Higher Specific Gravity Dehydration / Burns Pale yellow Dilute urine Lower Specific Gravity Overhydrated – maybe Diabetes Insipidus Urine color should always be evaluated by holding the sample up to a light source  Substances That Change Urine Color Blood or Myoglobin – Hemolytic Anemia, Transfusion Reaction, Renal Disease… Bilirubin – Liver Disease Amber Color Biliverdin – Liver Disease Green Exotoxins – Pseudomonas UTI Porphyrins – Porphyrias Port Wine Color Melanin – Melanoma Normal Medications, Dyes, Vitamins, Pigmented foods Yellow --------------------------------- Abnormal Colors --------------------------------- Urochrome Remember, before testing urine for any analyte, it must be well-mixed Remember, urine color should always be evaluated by holding the sample up to a light source  Urine Clarity Describes the cloudiness / turbidity of urine caused by suspended particulate matter which scatters light Normal urine is Clear Other options include: Hazy, Cloudy, and Turbid Causes of turbidity: Contamination (baby powder, lotion, toilet paper, cleansing pad…) X-ray Contrast Media Red Blood Cells, White Blood Cells, Epithelial Cells Bacteria Cells, Yeast Cells Mucus Casts and/or Crystals Lipids / Cholesterol Urine Clarity should always be evaluated by placing the sample in front of lined paper or text  Urine Clarity Clear Hazy Cloudy Turbid Clear Hazy Cloudy Turbid No particulate matter, Some particulate Significant particulate matter, Gross particulate matter, Lines are clearly visible matter, Lines are visible Traces of lines are visible Lines are not visible Remember, before testing urine for any analyte, it must be well-mixed Remember, Urine Clarity should always be evaluated by placing the sample in front of a background  Normal, Hazy Urine? Although normal urine should be Clear, a Hazy urine may be normal when caused by: Squamous Epithelia Cells* Mucus* Sperm Cells Contaminants Amorphous Phosphates / Amorphous Urates To prevent a falsely-hazy samples, patients should be given clear, detailed instructions for collection *Excessive presence of Squamous Epithelia Cells and/or Mucus may be pathogenic  Abnormal Urine Clarity A Cloudy or Turbid urine sample is never normal The presence of Bacteria Cells, Yeast Cells, Casts, or Lipids are never normal White Blood Cells greater than 3/HPF is never normal Any amount of Red Blood Cells (unless during menstruation) is never normal Some Crystals are pathogenic, and some are non-pathogenic  Correlating Red Urine Color and Clarity in Red Urine Clear Cloudy Clarity Clarity Pink Clear Hematuria Plasma Plasma (intact Red Blood Cells) Hemoglobinuria Crush Injury Myoglobinuria Hemolytic Anemia Renal Failure Myopathy Transfusion Reaction Menses  Transfusion Reactions Giving a patient just 15mL of incompatible ABO blood can be lethal If a patient receives “bad blood” and develops a transfusion reaction, the Red Blood Cells will be marked for destruction by their immune system This ultimately leads to Hemolysis which can be seen in both the urine and the blood Hemolysis will turn a patient’s samples Clear and Red as the lysed RBCs release Hgb Normal Hemolyzed Hemolyzed Normal Plasma (but clear) Plasma Urine Urine  Foam in the Urine Normal urine when shaken will produce small amounts of white foam (bubbles) that rapidly dissipates Stable White Foam indicates large amounts of Albumin in the urine Stable Yellow Foam indicates large amounts of Bilirubin in the urine The color should be Amber  Urine Odor Normal urine has an Aromatic odor Ingestion of certain foods or drugs can change the odor Other odors are associated with disease states: Foul / Ammonia – Bacterial Infections (UTI) Clarity would be Hazy-Turbid from the intact Bacteria Cells Sweet, Fruity – Ketones (diabetes, starvation, keto diet) Maple Syrup – Maple Syrup Urine Disease (MSUD) Mousy / Moist / Moldy – Phenylketonuria (PKU) Bleach – Contamination  Review Urine samples should be well-mixed before testing The Physical portion of the Urinalysis includes Color, Clarity, Foam, Odor, SG, and Osmo Color should be evaluated by holding the sample up to a light source Urine is normally Yellow due to Urochrome Bilirubin is Amber, Blood & Myoglobin are Red/Pink, Porphyrins are Port Wine, Melanin is Black, Green Exotoxins from a UTI caused by Pseudomonas is Green, Biliverdin is Blue-Green Clarity should be evaluated by holding the sample up to a background (lines / text) Clarity is most affected by intact cells, crystals, and casts White Foam = Albumin and Yellow Foam = Bilirubin Foul Odor = Bacteria, Sweet = Ketones, Maple = MSUD, Mousy = PKU, Bleach = contaminant All results should correlate with each other to form a diagnosis