Medical Technology Licensure Exam Review - Analysis of Urine and Other Body Fluids PDF

MEDICAL TECHNOLOGY LICENSURE EXAM REVIEW – ANALYSIS OF URINE AND OTHER BODY FLUIDS CLINICAL MICROSCOPY Lecturer: Sir Errol Coderes, RMT Notes by: Xiao - The Conqueror of Demons, The Vigilant Yaksha, & Alatus, the Golden-Winged King...

MEDICAL TECHNOLOGY LICENSURE EXAM REVIEW – ANALYSIS OF URINE AND OTHER BODY FLUIDS CLINICAL MICROSCOPY Lecturer: Sir Errol Coderes, RMT Notes by: Xiao - The Conqueror of Demons, The Vigilant Yaksha, & Alatus, the Golden-Winged King SAFETY IN THE CLINICAL LABORATORY B. SHARP HAZARDS - Sharp objects (needles, lancets, broken glassware) A. BIOLOGICAL HAZARDS - Disposed of in puncture-resistant containers - Potentially harmful organisms - Phlebotomists should carry these red, puncture-resistant - Chain of infection containers in their collection trays o Process of how microorganisms are transmitted from one person to another - An accidental needle-stick must be reported to the o Essential in preventing the spread of infection supervisor o Requires a continuous link between:  Source C. RADIOACTIVE HAZARDS  Mode of transmission - When procedures using radioisotopes are performed  Susceptible host - Exposure to radiation during pregnancy presents a danger to the o Routes of infection fetus  Inhalation - The best method of radioactive waste disposal is to store the used  Ingestion radioactive material in a locked, marked room until the background  Direct inoculation or skin contact count is down to 10 half-lives for radioiodine 6 COMPONENTS OF THE CHAIN OF INFECTION D. CHEMICAL HAZARDS 1. Infectious agent 2. Reservoir Chemical Spills 3. Exit (Portal of Exit) 4. Mode of transmission - BEST FIRST AID: Flush the area with amounts of water for at least 5. Entry (Portal of Entry) 15 minutes then seek medical attention. 6. Susceptible host - For alkali or acid burns in the eye, wash out eye thoroughly with running water for 15 minutes (Turgeon) - DO NOT NEUTRALIZE CHEMICALS that come in contact with the skin. PREVENTION - Acid spills on floors can be neutralized and then soaked up with wet rags or spill 1. PERSONAL PROTECTIVE EQUIPMENT pillows - Gloves Chemical Handling o For every patient, use new set of gloves (not applicable in the Philippines) - ALWAYS ADD ACID TO WATER!! - Fluid-resistant laboratory gowns o To avoid sudden splashing o An EXPLOSION can occur if water is added to acid - Eye and face shields - Countertop shields NATIONAL FIRE PROTECTION ASSOCIATION (NFPA) 2. PROPER HANDWASHING HAZARDOUS MATERIALS CLASSIFICATIONN - Hand contact is the primary method of infection transmission - Handwashing = best way to break the chain of infection - Uses numbers from 0 to 4 to classify hazard severity, with 4 representing - Handwashing procedure extremely hazardous o Wet hands with warm water - Mnemonic: You Were Born Right: Yellow, White, Blue Red (starting rightmost) o Apply antimicrobial soap o Rub to form lather, create friction, and loosen debris o Thoroughly clean between fingers, including thumbs, under fingernails and rings, and up to the wrist for at least 15 (or 20) seconds o Rinse hands in a downward position o Dry with a paper towel o Turn off faucets with a clean paper towel to prevent contaminations o And don’t forget the handwashing song: Happy Birthday (sung twice) KEEP IN MIND! Hand Hygiene - When the hands are visibly soiled: wash hands with soap and water - When the hands are not visibly soiled: apply alcohol-based hand-rub (ex. sanitizer) 3. DISPOSAL OF BIOLOGICAL WASTES - All biological waste, except urine, must be placed in appropriate containers labeled with the biohazard symbol. - The accepted "BIOHAZARD" label is fluorescent orange. (Henry) - Discard urine by pouring it into a laboratory sink, avoid splashing, and then flush with water. KEEP IN MIND! - Empty urine containers can be discarded as non-biologically hazardous Degree of Hazards (Hazards Index) waste - 0 = No/Minimal Hazard - Disinfection of the sink using a 1:5 or 1:10 dilution of sodium hypochlorite - 1 = Slight Hazard should be performed daily. - 2 = Moderate Hazard - Disinfection eliminates many or all pathogenic microorganisms, except - 3 = Serious hazard bacterial spores - 4 = Extreme/Severe Hazard - 1:10 dilution of sodium hypochlorite is prepared by adding 1part of sodium hypochlorite to 9 parts of water (effective for 1 month; used for disinfecting Mnemonic: “No SMS Ex’s” countertops & spills.) E. ELECTRICAL HAZARDS - DO NOT OPERATE equipment with wet hands. KEEP IN MIND! - All electrical equipment is grounded in a 3-pronged plug to avoid - The basic outline of the biohazard symbol is a plain trefoil, which is three circles electric shock overlapping each other equally like in a triple Venn diagram with the - If electrical shock occurs, never touch the person or the equipment overlapping parts erased. The diameter of the overlapping part is equal to half involved. the radius of the three circles. o Turn off the circuit breaker o Unplug the equipment o Move the equipment using a nonconductive glass or wood object F. FIRE /EXPLOSIVE HAZARDS RENAL FUNCTION - Flammable chemicals should be stored in safety cabinets and explosion-proof refrigerators in a remote area. URINARY SYSTEM - Persons with burning clothes should be wrapped in the blanket to - Kidney: weighs approximately smother the flames. 150g & measures 12.5 cm x 6 cm - All laboratory personnel must be involved in laboratory fire drills at x 2.5 cm (length, width, depth) least annually - Ureter: 25 cm long - Bladder: when approximately 150 WHEN A FIRE IS DISCOVERED… mL urine accumulates, a nerve - R (Rescue) = Rescue anyone in immediate danger reflex is initiated - A (Alarm) = Activate the institutional fire alarm system - Urethra: 4 cm long in women and - C (Contain) = Close all doors to potentially affected areas 24 cm long in men - E (Extinguish/Evacuate) = Attempt to extinguish the fire, if possible; exit the - About every 10-15 seconds, area small amounts of urine are emptied into the bladder from the TO OPERATE A FIRE EXTINGUISHER… ureters - P = Pull the pin - Urine is actually a fluid biopsy of - A = Aim at the base of the fire the kidney - S = Squeeze handles - S = Sweep nozzle side to side TYPES OF FIRE AND FIRE EXTINGUISHER Fire Type of Hazard Type of Extinguisher type NEPHRON A Ordinary combustibles: Water, dry chemical, loaded steam - Basic structural & functional unit of the kidney paper, cloth, rubbish, plastic, - 1 to 1.5 million nephrons per kidney wood - Consists of glomerulus and renal tubules B Flammable liquids: grease, Dry chemical, carbon dioxide, halon foam gasoline, paints, oil ORDER OF URINE FORMATION C Electrical equipment and Dry chemical, carbon dioxide, halon 1. Glomerulus motor switches 2. Proximal convoluted tubule (PCT D Flammable metals: mercury, Metal X, sand; dry powder; fought by fire 3. Loop of Henle (LH) magnesium, sodium, lithium fighters only 4. Distal convoluted tubule (DCT) E Detonation (Arsenal fire) Allowed to burn out and nearby materials 5. Collecting duct (CD) K Cooking media: grease, oils, Liquid designed to prevent splashing and --------------------------------------------- fats cool the fire 6. Renal Calyx - Water (A) | Dry chemicals (ABC) | Carbon dioxide (BC) | Halon (BC) 7. Renal Pelvis - Class D and E fires should be handled only by trained personnel - Dry chemical extinguishers (ABC) are the most common all-purpose extinguishers G. PHYSICAL HAZARDS GENERAL PRECAUTIONS: - Avoid running in rooms and hallways - Watch for wet floors - Bend knees when lifting heavy objects - Keep long hair pulled back - Avoid dangling jewelry - Maintain clean, organized work area - Wear closed-toe shoes MISCELLANEOUS HAZARD INFORMATION - Ergonomic hazards are work-related and include strain due to repeated positions - Cryogenic hazards are hazards due to extremely low temperatures - Mechanical hazards include centrifuges, refrigerators, autoclaves, homogenizers and glasswares - Centrifuge accidents or improper removal of rubber stopper from test tubes may produce aerosols 1. RENAL BLOOD FLOW - The kidneys receive 25% of the total cardiac output REVIEW!!! ORDER: 1. Clean fingers and wrist for at least: 4. Degree of hazard number 3: - Renal artery (blood in) a. 15 seconds a. Slight hazard - Afferent arteriole (“Approaching”) b. 30 seconds b. Serious hazard - Glomerulus c. 45 seconds c. Moderate hazard - Efferent arteriole (“Exiting) d. 60 seconds d. Extreme hazard - Peritubular capillaries (“Surrounding the renal tubules”) - Vasa recta 2. When hands are not visibly soiled: 5. Type of fire for flammable - Renal vein (blood out) a. Wash hands with soap and liquids water a. Type A - Total Renal Blood Flow: 1200 mL/min b. Apply sanitizer b. Type B - Total Renal Plasma Flow: 600 to 700 mL/min c. Apply Benzalkonium chloride c. Type C d. Wipe hands with paper towel d. Type D 2. GLOMERULAR FILTRATION e. Type E 3. Yellow quadrant in the NFPA f. Type K GLOMERULUS hazard classification: a. Health hazard - The "working portion" of the kidney b. Fire hazard - Coil of approx. 8 capillary lobes (capillary tuft) located within the Bowman's c. Specific hazard capsule d. Reactivity hazard - Attached to the glomerular basement membrane are the podocytes (epithelial cells) - Resembles a sieve - Non-selective filter of plasma substances with MW of 200 d. Many  RTE cells from the DCT may be mistaken for WBCs  >2 RTE/hpf indicates tubular injury 2. Nine (9) bacteria were seen per HPF. How do  VARIATIONS you report this finding?  Oval Fat Body (Renal tubular fat bodies) a. Rare o Lipid containing RTE cell (may also be a b. Few monocyte/macrophage) c. Moderate o Seen in lipiduria (Ex. nephrotic syndrome) d. Many o Identified by:  Lipid stains (TAG and neutral fats) Answer key: C, B, B  Polarizing microscope (Cholesterol - "MALTESE CROSS" formation) CASTS  Bubble cell o RTE cell with non-lipid vacuoles - Excretion is termed cylindruria o Injured cells in which the endoplasmic reticulum has dilated - Unique to the kidney prior to cell death - Represents a biopsy of an individual tubule o Seen in acute tubular necrosis - The most difficult & the most important urinary sediment constituent - Primarily formed in the = DCT and collecting duct 4. Bacteria - Major constituent = Uromodulin/THP (produced by RTE cells) o True UTI = Bacteria + WBCs (If bacteria only = contamination or old - Other proteins such as albumin & immunoglobulins are also incorporated into the specimen) cast matrix o Enterobacteriaceae (Ex. E. coli) = most common cause of UTI - Cylindroids have the same significance as Casts (a cast with a tail) o Staphylococcus, Enterococcus - Protein gels more readily under conditions of urine-flow stasis, acidity and o Motility differentiates them from amorphous urates & phosphates presence of Na+ and Ca2+ - A cast structure should have an even & definite outline, parallel sides and two 5. Yeasts rounded ends o True yeast infection = Yeast + WBCs (If yeast only = contamination) - Has uniform diameter (about 7-8x the diameter of RBCs) o Small, refractile oval structure that may or may not bud - Examination is performed along the coverslip edges with subdued light o Branched, mycelial forms are seen in severe infections o Candida albicans = seen in DM and vaginal moniliasis FORMATION OF CASTS 6. Parasites 1. Aggregation of uromodulin into individual protein fibrils attached to RTE cells o Trichomonas vaginalis 2. Interweaving of protein fibrils to form a loose fibrillar network  Most frequently encountered parasite in urine 3. Further protein fibril interweaving to form a solid structure  Pear-shaped flagellate with jerky motility 4. Possible attachment of urinary constituents to the solid matrix  Agent of Ping-Pong disease 5. Detachment of protein fibrils from the epithelial cells  Reported as rare, few, moderate, or many per HPF 6. Excretion of the cast  When not moving, may resemble WBC, T.E.C. or R.T.E. cell o Enterobius vermicularis egg  Most common fecal contaminant o Schistosoma haematobium egg  Blood fluke with terminal spine  Causes hematuria  Associated with bladder cancer o Other parasites include: Trichuris, Strongyloides, Giardia, various amoebae o Various insects or “bugs" (lice, fleas, bedbugs, mites, and ticks) KEEP IN MIND! - Urinary Bladder Cancer Markers (Specific) o NMP = Nuclear Matrix Protein o BTA = Bladder Tumor Antigen 7. Spermatozoa o Oval, slightly tapered head 1. HYALINE CAST o Long, flagella-like tail o Prototype cast (beginning of all types of cast) o After sexual intercourse o Most frequently encountered & the most difficult cast to discover o Colorless and translucent 8. Mucus Threads o Normal value = 0-2/LPF o Has low refractive index o Physiologic Stress = strenuous exercise o Major constituent: Tamm-Horsfall protein (Uromodulin) o Pathologic = Glomerulonephritis, pyelonephritis, CHF, CKD MICROSCOPIC QUANTITATIONS (Strasinger) 2. RBC CAST - Quantitate an average of 10 representative fields. o Most fragile cast - Do not quantitate budding yeast, mycelia elements, Trichomonas, or sperm, o Indicates bleeding within the nephron but do note their presence with the appropriate LIS code o Easily identified by its orange-red color Quantitated None Rare Few Moderate Many o Significance = Glomerulonephritis, strenuous exercise Epithelial cells per LPF 0 0-5 5-10 20-100 >100 o Blood Cast Crystals per HPF 0 0-2 2-5 5-20 >20  Contains hemoglobin from lysed RBCs (normal)  Homogeneous appearance with orange-red color Bacteria per HPF 0 0-10 10-50 50-200 >200  Same significance as RBC cast Mucus threads per LPF 0 0-1 1-3 3-10 >10 Casts per LPF 0 Numerical ranges: 0-2, 2-5, 5-10, >10 3. WBC/LEUKOCYTE/PUS CAST RBCs per HPF 0 Numerical ranges: 0-2, 2-5, 5-10, 10- o Indicates inflammation or infection within the nephron WBCs per HPF 0 25, 25-50, 50-100, >100 o Resembles RTE cast. To distinguish, use phase microscopy and supravital Squamous epithelial cells Rare, few, moderate, or many per LPF stain. Transitional epithelial cells, Rare, few, moderate, or many per HPF o Significance = Pyelonephritis, acute interstitial nephritis yeasts o Pseudoleukocyte Cast Renal tubular epithelial cells Average number per 10 HPFs  Not a true cast (DO NOT report as cast!) Oval fat bodies Average number per HPF  Clump of leukocytes Abnormal crystals, casts Average number per LPF  Seen in lower UTI 4. EPITHELIAL (RTE) CAST Calcium Oxalate (Dihydrate and Monohydrate) o Cells visible on the cast matrix are smaller, round and oval cells - The most frequently observed urinary crystal o Significance = Advanced tubular destruction, tubular damage - Dihydrate (Weddellite) = more common; envelope, bipyramidal, octahedron - Monohydrate (Whewellite) = oval/dumbbell 5. BACTERIAL CAST - ↑ Foods rich in oxalic acid (tomato, asparagus) and ascorbic acid - Ascorbate when metabolized becomes oxalate and combines with calcium to o Identified by performing Gram stain form calcium oxalate o Significance = Pyelonephritis - ↑ Ethylene glycol (anti-freeze agent) or methoxyflurane poisoning (MH) - Soluble in dilute HCl 6. GRANULAR CAST (COARSE AND FINE) o Granules are derived from the lysosomes of RTE cells during normal OTHER NORMAL ACID CRYSTALS metabolism (nonpathologic) o Cells disintegrate when the cast is retained in the tubule before being flushed Calcium Sulfate out - “Cigarette-butt” appearance o Finely granular cast has a sandpaper appearance - Soluble in acetic acid o Significance = Glomerulonephritis, pyelonephritis, stress, strenuous exercise Hippuric Acid - Yellow-brown/colorless elongated prism 7. FATTY CAST - Soluble in water and ether o Fat globules not stained by Sternheimer-Malbin stain (only the cast matrix is Acid Urates (Na+, K+, NH4) - Rare form of uric acid stained) - Brown spheres or clusters o Identification: - Resembles ammonium biurate, leucine & sulfamethoxazole crystals  TAG & neutral fats = Lipid stains - Turns into uric acid after adding acetic acid  Cholesterol = Polarizing microscope (“Maltese cross”) - Soluble in heat and alkali o Significance = Nephrotic syndrome, Toxic tubular necrosis, Diabetes mellitus, Monosodium or Sodium Urates crush injuries - Rare form of uric acid - Tiny, slender, colorless needles; spherulite or beachball (rare) 8. WAXY CAST - ↑ Acute gout (intracellular crystals) and chronic gout (extracellular crystals) o Final degenerative form of all types of casts o Brittle, highly refractile, with jagged ends NORMAL ALKALINE CRYSTALS o Ground glass appearance Amorphous phosphates (Ca2+ & Mg2+) o Significance = Stasis of urine flow, chronic renal failure - Most common cause of turbidity in alkaline urine - Fine, or ‘lacy' white precipitate (macroscopic) 9. BROAD CAST - Granular in appearance (microscopic) o Often referred to as renal failure cast (Strasinger) - Soluble in dilute acetic acid o Indicates destruction (widening) of the tubular walls Ammonium biurate o Any type of cast can be broad (most common are granular & waxy) - Alkaline counterpart of uric acid and amorphous urates - Yellow-brown thorny apples o 2-6x wider than ordinary cast - Seen in old specimens o Significance = Extreme urine stasis, renal failure - ↑ Presence of urea-splitting bacteria (urea ammonia) 10. MISCELLANEOUS CASTS - Turns into uric acid after adding HCl or acetic acid - Soluble in acetic acid with heat o Pigmented Cast Triple Phosphate (Magnesium Ammonium Phosphate, Struvite)  Hyaline matrix with coloration due to pigment incorporation - Colorless, prism-shaped or coffin-lid; fern-leaf (Harr)  Incorporated bilirubin (golden-brown) - Feathery appearance when they disintegrate  Hemoglobin or myoglobin (yellow to red brown) - Presence of urea-splitting bacteria (urea ammonia) o Mixed Cellular Cast - Soluble in dilute acetic acid  Cast containing multiple cell types Magnesium Phosphate  Glomerulonephritis (RBC& WBC casts) - Colorless, elongated rectangular or rhomboid plates  Pyelonephritis (WBC & RTE casts or WBC and bacterial casts) - End or corner may be notched o Crystal Cast - Edges can be irregular or eroded  Casts containing urates, calcium oxalate and sulfonamides are - Soluble in acetic acid occasionally seen Calcium Phosphate (Apatite) - Dibasic calcium phosphate (stellar phosphate)  Deposition of crystals in the tubule or collecting duct o Colorless, flat plates thin prisms in rosette form SEDIMENT CONSTITUENTS – Chemical or Unorganized Sediment (Crystals) o Rosettes may resemble sulfonamide crystals - Monobasic calcium phosphate o Irregular, granular-appearing sheets or plates CRYSTALS o Less common than dibasic form - Other forms: - Excretion is termed crystalluria o Hydroxyapatite (basic calcium phosphate) - The most recognized but the most insignificant part of urine sediment (Turgeon) o Brushite (calcium hydrogen phosphate) - Formed by precipitation of urine solutes (salts, organic compounds, medication) - Soluble in dilute acetic acid - Presence of crystals in fresh urine is most frequently associated w/ concentrated Calcium Carbonate specimen (↑ S.G.) - Small, colorless, dumbbell, tetrads or spherical-shaped - Factors that contribute to crystal formation: - Forms gas (effervescence) after adding acetic acid o pH - Misidentified as bacteria o Solute concentration o Temperature ABNORMAL ACID CRYSTALS - Usually reported as rare, few, moderate or many per HPF Cystine - Abnormal crystals may be averaged and reported per LPF - Colorless, refractile hexagonal plates, often laminated - Mistaken as hexagonal uric acid crystals - Abnormal crystals generally require confirmation before they are reported to the - ↑ in Cystinuria and Cystinosis physician Uric acid Cystine NORMAL ACID CRYSTALS Color Yellow brown Colorless Amorphous Urates (Ca2+, Mg2+, Na+, & K+ urates) Solubility in ammonia Soluble Soluble - Fluffy orange or pink sediment (brick dust) due to uroerythrin Solubility in dilute HCl Insoluble Soluble - Yellow brown granules (microscopic) Birefringence Birefringent Not birefringent - Clumps resemble granular casts (termed "pseudocasts") Cyanide-nitroprusside reaction Negative Positive - Turns into uric acid after adding acetic acid Cholesterol - Turns into ammonium biurate after adding ammonium hydroxide - Rectangular plate with notch in one or more corners (staircase pattern) - ↑ in Gout, Chemotherapy - Resembles crystals of radiographic dye - Soluble in heat and alkali - in Nephrotic syndrome (lipiduria) Uric Acid - Soluble in chloroform - Product of purine metabolism Radiographic dye (Meglumine diatrizoate, Renografin, Hypaque - Most pleomorphic: Rhombic (diamond), 4-sided flat plate (whetstone), lemon- - Flat, four-sided plates often with a notched corner shaped - Other forms include-long8 thin prisms or rectangles - Wedges, barrel, rosettes, irregular plates, laminated forms are also seen - Resembles cholesterol crystals. To differentiate: - Hexagonal forms may be mistaken as cystine crystals o Check patient history - Present at pH 5.7, it is in its ionized form as urate o Correlate with other UA results (Ex: S.G. >1.040 using refractometer) - ↑ in Lesch-Nyhan syndrome, Chemotherapy, Gout - Soluble in 10% NaOH - Soluble in alkali  Tyrosine URINE SCREENING FOR METABOLIC DISORDERS - Fine colorless to yellow needles in clumps or rosettes - ↑ in Liver disease (more commonly found than leucine) 2 CATEGORIES OF AMINOACIDURIA - Soluble in alkali or heat OVERFLOW TYPE RENAL TYPE Leucine - ↑ Amino acid in blood - Normal Amino acid in blood - Yellow-brown oily-looking spheres w/concentric circles & radial striations - ↑ Amino acid in urine - ↑ Amino acid in urine - Precipitated with tyrosine after adding alcohol - Examples: PKU, MSUD, cystinosis - Due to defective tubular - May resemble fat globules reabsorption of amino acids - ↑ in Liver disease - Examples: Cystinuria, Fanconi's - Soluble in hot alkali or alcohol syndrome Bilirubin - Clumped granules or needles with bright yellow color (Strasinger) Inborn Error of Metabolism (EM) - Reddish brown needles that cluster in clumps or spheres (Turgeon) - Failure to inherit a gene that codes for a particular enzyme. - Granules and plates have been observed (Brunzel) - No gene = No enzyme - ↑ in Liver disease - Soluble in acetic acid, HCI, NaOH, acetone and chloroform I. PHENYLALANINE -TYROSINE DISORDERS ABNORMAL ACID CRYSTALS 1. PHENYLKETONURIA Sulfonamide - Fan-shaped needles, sheaves of wheat. rosettes, arrowheads, petals, round- 𝑃ℎ𝑒𝑛𝑦𝑙𝑎𝑙𝑎𝑛𝑖𝑛𝑒 → 𝑃ℎ𝑒𝑛𝑦𝑙𝑎𝑙𝑎𝑛𝑖𝑛𝑒 ℎ𝑦𝑑𝑟𝑜𝑥𝑦𝑙𝑎𝑠𝑒 𝑇𝑦𝑟𝑜𝑠𝑖𝑛𝑒 shaped, whetstones - Mistaken as calcium phosphate crystals. To differentiate: - The most well-known of the aminoacidurias o Calcium phosphate: soluble in acetic acid - (-) gene that codes for phenylalanine hydroxylase o Sulfonamide = (+) Lignin test & Diazo reaction - Other forms are due to lack of tetrahydrobiopterin - Possible tubular damage (may deposit in nephrons - ↑ Phenylpyruvic acid (a ketone) in urine - Currently, they are modified and their solubility is no longer a problem - “Mousy” odor of urine, sweat and breath odor (due to phenylacetic acid) - Soluble in acetone - May lead to severe mental retardation - Forms: - Screening tests: o Sulfamethoxazole = brown rosettes or spheres w/ irregular radial o FeCl3 tube test = (+) blue-green color striations o Phenistix strip = (+) gray to gray-green o Acetylsulfadiazine = yellow-brown sheaves of wheat w/ eccentric binding o Guthrie bacterial inhibition test o Sulfadiazine = yellow-brown sheaves of wheat w/ eccentric binding - Confirmatory test: Ion exchange HPLC Ampicillin - Colorless needles that tend to form bundles following refrigeration GUTHRIE BACTERIAL INHIBITION TEST: - ↑ in massive doses of penicillin - B. subtilis is cultured w/ B2-thienylalanine (TE) - B2-TE Inhibits the growth of B. subtilis. OTHER ABNORMAL CRYSTALS Hemosiderin - Phenylalanine counteracts the action of B2-TE - Coarse, yellow-brown granules - (+) result: growth, (-): no growth - Resembles amorphous urates - (+) Rous test (Prussian blue stain) Acyclovir 2. TYROSYLURIA/TYROSINEMIA - May be seen in alkaline urine - (-) gene that codes for: - Colorless slender needles o Type 1: Fumarylacetoacetate hydrolase (FAH) - Strongly birefringent with polarized light o Type 2: Tyrosine aminotransferase Indinavir Sulfate o Type 3: p-hydroxyphenylpyruvic acid dioxygenase - Slender colorless needles or slender rectangular plates - May also be seen in severe liver disease - Feather-like crystals that aggregate into wing-like bundles - Arranged in fan-shaped or starburst forms, bundles, or sheaves - “Rancid butter” urine odor - Associated with renal blockage & stone formation in HIV-positive individuals - Screening Tests o FeCl3 tube test = (+) transient green o Nitroso-naphthol = (+) orange-red URINARY SEDIMENT ARTIFACTS - Confirmatory Tests o Chromatography - Starch granules o Quantitative serum assay of tyrosine o Spheres with dimpled center o "Maltese cross" formation on polarizing microscope PHENYLALANINE AND TYROSINE METABOLISM  Oval fat bodies  Fatty casts  Fat droplets  Starch granules - Oil droplets = mistaken for RBCs - Air bubbles = mistaken for RBCs - Pollen grains = spheres with cell wall & concentric circles - Hair and fibers = mistaken for casts - Fecal contamination REVIEW!!! 1. Nomarski microscope is a type of what microscopy? a. Phase contrast b. Brightfield c. Polarizing d. Fluorescent 2. Oval fat bodies are seen in: a. Acute glomerulonephritis b. Interstitial nephritis c. Nephritic syndrome d. Nephrotic syndrome 3. Colorless needles that tend to form bundles following refrigeration: a. Uric acid b. Sulfonamide c. Ampicillin d. Leucine 4. What are the forms of triple phosphate crystal? 5. Sequence of cast degeneration starting from the worst Answer key: A, D, C - State laws require that blood be collected 24 hours after birth and before the 2. ARGENTAFFINOMA newborn leaves the hospital - Tumor of argentaffin or enterochromaffin cells - produce serotonin (carried by - "Testing for may substances is now performed using tandem mass platelets) → metabolized into 5-HIAA spectrophotometry (MS/MS). It is capable of screening the infant blood sample for - Screening tests specific substances associated with particular inborn error of metabolism." o FeCl3 tube test = (+) Blue-green o Nitrosonaphthol with nitrous acid = (+) Violet 3. ALKAPTONURIA - Patient must not eat bananas, pineapples, tomatoes, avocados, chocolates, - (-) gene that codes for Homogentisic acid oxidase walnuts, & plums (they ↑ serotonin) - ↑ Homogentisic acid in blood and urine - Urine darkens after becoming alkaline from standing at room temperature IV. CYSTINE DISORDERS - Brown - or black-stained cloth diapers - “Sulfur” urine odor - Reddish-stained disposable (plastic) diapers - Treatment = D-Penicillamine - Homogentisic acid causes black pigmentation in the connective tissues and ears (ochronosis) 1. CYSTINURIA - Screening Tests - Renal type of aminoaciduria o FeCl3 tube test = (+) transient blue - Defective tubular reabsorption of: o Clinitest = (+) yellow precipitate o Cystine (the only one which crystallizes; least soluble) o Alkalinization of fresh urine o Ornithine - Confirmatory Tests o Lysine o Paper/thin-layer chromatography o Arginine o Capillary electrophoresis - Tests for Cystinuria and Cystinosis o Brand's modification of Legal's nitroprusside 4. MELANURIA  Reagent = Cyanide nitroprusside - Caused by melanoma (tumor involving melanocytes)  (+) Red-purple color - Tumors secrete 5,6-dihydroxyindole, which oxidizes to melanogen then to melanin o Thin layer or ion-exchange chromatography - Urine darkens upon air exposure o High-voltage electrophoresis - Deficient production of melanin results in albinism - Screening Tests 2. CYSTINOSIS o FeCl3 tube test = (+) Gray/black ppt - Inborn error of metabolism → Overflow type o Sodium nitroprusside test = (+) Red - (-) gene that codes for an enzyme responsible for cystine metabolism o Ehrlich test = (+) Red - Types = Nephropathic cystinosis, intermediate cystinosis, and ocular cystinosis - Cystine deposits in many areas of the body (BM, cornea, lymph nodes & internal II. BRANCHED-CHAIN AMINO ACID DISORDERS organs) 3. HOMOCYSTINURIA - Defects in the metabolism of methionine (leads to ↑ homocystine) - (-) gene that codes for the enzyme cystathione β-synthase - Detected by the Silver-nitroprusside test = (+) Red-purple color 1. MAPLE SYRUP URINE DISEASE (MSUD) - Most common IEM in the Philippines V. PORPHYRIN DISORDERS (PORPHYRIAS) - (-) Gene that codes for the enzyme complex known as branched-chain α-keto acid dehydrogenase (BCKD) - ↑ Ketoacids of Leucine, Isoleucine and Valine - "Caramelized sugar/Maple syrup/Curry" urine odor - Presence of ketonuria in a newborn is significant - Screening Test o 2,4-dinitrophenylhydrazine (DNPH) = (+) Yellow turbidity/precipitate - Confirmatory Test o Gas or thin-layer chromatography o Nuclear magnetic resonance spectro 2. ORGANIC ACIDEMIAS - Isovaleric acidemia = "sweaty feet" urine odor due to isovalerylglycine o Glutaric acidemia also presents with a sweaty feet urine - Propionic acidemia - Methylmalonic acidemia = detected using p-nitroaniline test = (+) Emerald green color III. TRYPTOPHAN DISORDERS - Disorders of porphyrin metabolism 1. INDICANURIA - Urine color - Indigo blue urine color (upon air exposure) o red, purple, burgundy-red, purplish red, “portwine” - Seen in: o Colorless in = lead poisoning (Harr, Henry) o Hartnup disease (“Blue diaper syndrome”) o Intestinal disorders Consider it porphyria if: - Screening test: Obermayer's test - Red-tinged urine o FeCl3 + Urine + Chloroform = (+) Violet color - Negative for blood reagent strip - Diet and medications ruled out  Colorless Dark Red or Purple, REVIEW!!! Nonfluorescent Intensely Fluorescent 1. Urine darkens after becoming alkaline, what is present? - Porphobilinogen - Uroporphyrin a. Melanin - D-aminolevulinic acid - Coproporphyrin b. Methemoglobin - Uroporphyinogen - Protoporphyrin c. Homogentisic acid d. Methyldopa - Coproporphyrinogen 2. What is the positive result in the Guthrie bacterial inhibition test: - Protoporphyrinogen a. Growth around the disk b. No growth around the disk SCREENING TESTS c. Lysis of colonies Specimen = Urine, Stool, Blood, Bile d. Clearing of the medium Ehrlich's reaction Detects D-ALA, porphobilinogen 3. Cetyltrimethylammonium bromide test is a screening test for: a. Phenylalanine Fluorescence at 550- Tests for uroporphyrin, coproporphyrin, & protoporphyrin b. Tryptophan 600 nm (+) Violet/Pink/Red fluorescence c. Mucopolysaccharides Free erythrocyte CDC- CDC-recommended test for lead poisoning d. Porphyrins recommended test for 4. Positive color in the MPS test? lead protoporphyrin a. Red (FEP) b. Blue c. Yellow Disorder Enzyme Deficient Elevated Clinical d. Green 5. Urine color in lead poisoning: Compound/s Symptoms a. Portwine Acute Uroporphyrinogen ALA Neurologic b. Blue intermittent synthase Porphobilinogen Psychiatric c. Brown/black porphyria d. Colorless Porphyria Uroporphyrinogen Uroporphyrin Photosensitivity cutanea tarda decarboxylase Congenital Uroporphyrinogen Uroporphyrin Photosensitivity Answer key: C, A, C, B, D erythropoietic cosynthase Coproporphyrin porphyria RENAL DISEASE Variegate Protoporphyrinogen Coproporphyrin Photosensitivity porphyria oxidase Neurologic CLASSIFICATIONS OF RENAL DISEASE Erythropoietic Ferrocheletase Protoporphyrin Photosensitivity - Glomerular disorders protoporphyria o Majority are of immune origin (immune complexes, IgG, IgA) Lead poisoning -- ALA Photosensitivity o Common: proteinuria, hematuria, casts Protoporphyrin - Tubular disorders - Port wine urine color is more prevalent in the erythropoietic porphyrias - Interstitial disorders - Lead po1soning inhibits ALA Synthetase and ferrocheletase enzymes GLOMERULAR DISORDERS DISORDER AND ETIOLOGY FINDINGS VI. MUCOPOLYSACCHARIDE [MPS] DISORDERS 1 Acute Post-Streptococcal - Macroscopic hematuria, (MUCOPOLYSACCHARIDOSIS) Glomerulonephritis proteinuria, dysmorphic - Impaired metabolism of mucopolysaccharides or glycosaminoglycans (protein + - Deposition of immune complex, RBCs, RBC casts, polysaccharides, located in the connective tissues) formed in conjunction of Group A granular casts, - Frequently found in urine are dermatan sulfate, keratan sulfate and heparan Streptococcus (S. pyogenes) infection - (+) ASO titer sulfate on the glomerular membranes 2 Rapidly Progressive (Crescentic) 1. HURLER SYNDROME Glomerulonephritis o A.k.a. Gargoylism or MPS Type I - Deposition of immune complexes from - Macroscopic hematuria o MPS accumulate in the cornea of the eye systemic immune disorders (ex: SLE) - Proteinuria o (+) Skeletal abnormalities & mental retardation on the glomerular membrane - RBC casts 2. HUNTER SYNDROME - Cellular proliferation of epithelial cells o Sex-linked recessive, rarely seen in females inside the Bowman's capsule form o (+) Skeletal abnormalities & mental retardation "crescents” 3 Goodpasture Syndrome 3. SANFILIPPO SYNDROME - Deposition of antiglomerular basement - Macroscopic hematuria o Mental retardation is the only abnormality membrane antibody to glomerular and - Proteinuria alveolar basement - RBC casts SCREENING TESTS 4 Wegener's Granulomatosis - Acid albumin test = (+) White turbidity - Anti-neutrophilic cytoplasmic auto- - Macroscopic hematuria - Cetyltrimethylammoniumbromide (CTAB) Test = (+) White turbidity antibody (ANCA) binds to neutrophils - Proteinuria - Mucopolysaccharide (MPS) Paper Test = (+) Blue color in vascular walls producing damage to - RBC casts small vessels in the lungs and VII. PURINE DISORDER glomerulus - Perinuclear ANCA (p-ANCA) forms LESCH-NYHAN DISEASE when neutrophils are fixed in ethanol - (-) gene that codes for the enzyme hypoxanthine guanine - Cytoplasmic ANCA (c-ANCA) forms phosphoribosyltransferase when neutrophils are fixed with - ↑ Uric acid in the blood and urine formalin - “Orange sand” in diapers 5 Henoch Schönlein Purpura - Occurs in children following viral - Macroscopic hematuria VIII. CARBOHYDRATE DISORDERS respiratory infections - Proteinuria - Glucose strip and (+) Copper reduction test - Decrease in platelets disrupts vascular - RBC casts - Melituria = presence of any sugar in urine integrity 6 Membranous Glomerulonephritis (MGN) 1. Galactosemia/Galactosuria - Thickening of glomerular membrane - Microscopic hematuria o Inability to metabolize galactose to glucose following IgG immune complex - Proteinuria o Enzymes absent: deposition associated with systemic  Galactose-1-phosphate uridyl transferase (GALT) disorders  Galactokinase 7 Membranoproliferative  UDP-galactose-4-epimerase Glomerulonephritis (MPGN) o ↑ Galactitol, galactonate and galactose-1-phosphate - Cellular proliferation affecting the - Hematuria o Associated with infant failure to thrive, liver disorders, cataracts and capillary walls or the glomerular - Proteinuria severe mental retardation basement membrane, possibly 2. Glucosuria = Diabetes Mellitus immune-mediated 3. Lactosuria = seen during pregnancy and lactation - Glomeruli have visible lobular 4. Fructosuria = associated with parenteral feeding appearance 5. Pentosuria = associated with ingestion of large amounts of fruit - “Tram track” 8 Chronic Glomerulonephritis INTERSTITIAL DISORDERS - Marked decrease in renal function - Hematuria DISORDER AND ETIOLOGY FINDINGS resulting from glomerular damage - Proteinuria, Glucosuria 1 Cystitis (Lower UTI) precipitated by other renal disorders - Cellular & granular casts - Ascending bacterial infection of - WBCs, Bacteria, NO CAST - Progression to renal failure - Waxy and broad casts the urinary bladder - Microscopic hematuria 9 IgA Nephropathy (Berger's Disease) - Acute onset of urinary - Mild proteinuria, increased pH - Deposition of lgA on the glomerular - Early stages: Hematuria frequency and burning membrane resulting from increased - Late stages: See chronic 2 Acute Pyelonephritis (Upper UTI) levels of IgA GN - Infection of the renal tubules & - WBCs, Bacteria 10 Minimal Change Disease, MCD interstitium related to - WBC casts, bacterial casts, (Nil Disease/Lipoid Nephrosis) interference of urine flow to the - Microscopic hematuria - Little cellular changes in the - Heavy proteinuria bladder, reflux of urine from the glomerulus - Transient hematuria bladder (vesicoureteral reflux) - Glomeruli look normal by light - Fat droplets & untreated cystitis microscopy 3 Chronic Pyelonephritis - Electron microscopy reveals loss of - Recurrent infection of the renal - WBCs, Bacteria, WBC casts, podocyte foot processes tubules & interstitium caused Bacterial casts, granular casts - Disruption of podocytes primarily in by structural abnormalities - Waxy and broad casts children following allergic reactions & affecting the flow of urine - Hematuria, proteinuria immunizations 4 Acute Interstitial Nephritis - Associated with HLA-B12 antigen - Allergic inflammation of the - Hematuria, proteinuria 11 Focal Segmental Glomerulosclerosis renal interstitium in response to - WBCs (↑ eosinophils, >1%) (FSGS) certain medications - WBC casts, NO BACTERIA - Disruption of podocytes in certain - Proteinuria numbers and areas of glomeruli, - Hematuria RENAL FAILURE others remain normal - ↓ Glomerular filtration rate (< 25 mL/min) - IgM and C3 are evident on the sclerotic - Azotemia (↑ BUN & Creatinine) areas (using IF) - Electrolyte imbalance 12 Diabetic Nephropathy (Kimmelstiel- - (-) renal concentrating ability → Isosthenuria Wilson Disease) - Proteinuria & renal glycosuria - Most common cause of ESRD - Microalbuminuria - Telescoped sediment - Deposition of glycosylated proteins on - + Micral test o Variety of casts seen in the same specimen (cellular, coarsely granular, finely the glomerular basement membranes granular, waxy) caused by poorly controlled blood o Simultaneous appearance of the elements of acute, chronic GN, and glucose levels nephrotic syndrome 13 Alport Syndrome o ↑ casts (granular, waxy, broad) - Genetic disorder showing lamellated - See Nephrotic Syndrome and thinning of glomerular basement RENAL CALCULI/RENAL LITHIASIS membrane - May form in the calyces and pelvis of the kidney, ureters, and bladder 14 Nephrotic Syndrome - Lithotripsy uses high-energy shock waves to break kidney stones into pieces - Disruption of the electrical charges that produce the tightly fitting - Conditions Favoring the Formation of Renal Calculi: podocyte barrier resulting in massive loss of proteins & lipids o pH - The rate of proteinuria in nephrotic syndrome is >3.5 g/day o Chemical concentration - Occurs in patients with MCD (in children), MGN (in adults), FSGS and o Urinary stasis MPGN - Primary UA Finding = Microscopic hematuria BLOOD URINE Serum Findings: - ↓ Albumin - ↑ Albumin - Albumin, α1, gamma - globulins Renal Calculi Information/Description - ↓ Lipase - ↑ Lipase - α2-macroglobulin Calcium oxalate - Major constituent of renal calculi - ↑ Lipids - ↑ Lipids - β-globulin (LDL) calculi - Very hard, dark in color with rough surface - ↑ Apo B100 Uric acid & Urate - Associated w/ increased intake of foods w/ high purine - ↑ LDL and Urinalysis Findings: calculi content, and w/ UKD VLDL - Albumin, a1, B, gamma-globulins - Yellowish to brownish red & moderately hard - ↑ Chole & - (-) a2-macroglobulin Cystine calculi - Seen in hereditary disorders of cystine metabolism TAG - Oval fat bodies - Yellow-brown, greasy & resembles an old soap - Fatty casts - Least common calculi - Waxy casts Phosphate calculi - Pale & friable Triple phosphate - Accompanied by urinary infections involving urea- TUBULAR DISORDERS calculi splitting bacteria (Proteus vulgaris) DISORDER AND ETIOLOGY FINDINGS - Branching/staghorn calculi resembling antlers of a 1 Acute Tubular Necrosis deer - Damage to renal tubular cells caused by - Microscopic hematuria, Rare Calculi: ischemia or toxic agents proteinuria - Sulfonamide calculi - Urine odor = “odorless” - RTE cells, RTE casts - Silica calculi = ingestion of silica over a long period of time - Hyaline, granular, waxy - Triamterene calculi = insoluble diuretic; mustard-colored stones and broad casts - Adenine calculi = associated with inherited enzyme deficiency & hyperuricemia 2 Uromodulin-associated Kidney Disease - Xanthine calculi = associated with a genetic disorder w/ an absence of xanthine (UKD) - RTE cells oxidase - Inherited defect in the production of - Hyperuricemia normal uromodulin by the renal tubules and increased uric acid causing gout - Normal uromodulin is replaced by abnormal forms that destroy the RTE cells 3 Fanconi Syndrome - Generalized failure of tubular reabsorption - Glucosuria in the proximal convoluted tubule - Possible cystine crystals (amino acid) 4 Diabetes Insipidus - Neurogenic DI = hypothalamus fails to - Low specific gravity produce ADH - Polyuria (>15 L/day) - Methods for Calculi Analysis - Nephrogenic DI = renal tubules fail to o Optical crystallography respond to ADH o Radiograph diffraction 5 Renal Glucosuria o Infrared spectroscopy - (N) Blood glucose = ↑ Urine glucose - Glucosuria o Electron beam analysis - Defective tubular reabsorption of glucose o Mass spectroscopy  REVIEW!!! AMNIOTIC FLUID 1. Thickening of the glomerular membrane following IgG immune complex - Present in the amnion – a membranous sac that surrounds the fetus deposition? - The placenta is the ultimate source of amniotic fluid water and solutes a. Membranous GN b. Berger’s disease PRIMARY FUNCTIONS OF AMNIOTIC FLUID c. Chronic GN d. Nephrotic syndrome 1. Cushion for the fetus 2. What is the indicator of diabetic nephropathy? 2. Stabilizes temperature a. Lipiduria 3. Allows fetal movement b. Hematuria 4. Proper lung development c. Microalbuminuria d. Glucosuria AMNIOTIC FLUID VOLUME 3. (+) WBC, bacteria, but no cast - From fetal urine and lung fluid a. Cystitis - Normal = 800-1,200 mL (3rd trimester) b. Pyelonephritis - During 1st trimester, 35 mL of amniotic fluid is derived primarily from the maternal c. Interstitial nephritis d. Tubular necrosis circulation 4. Renal calculi: yellow-brown, greasy and resembles an old soap: - Fetal urine = major contributor to the AF volume after the 1st trimester of a. Calcium oxalate pregnancy b. Phosphate c. Uric acid POLYHYDRAMNIOS OLIGOHYDRAMNIOS d. Cystine ↑ amniotic fluid volume (1,200 mL) Causes: Causes: Answer key: A, C, A, D - Decreased fetal swallowing of urine - Increased fetal swallowing of urine - Neural tube defects (ex. Spina - Membrane leakage AMNIOTIC FLUID & hCG bifida) - Urinary tract deformities - Others: fetal structural anomalies, - Others: congenital malformations, HUMAN CHORIONIC GONADOTROPIN (hCG) cardiac arrhythmias, congenital premature amniotic membrane - Produced by the syncytiotrophoblast cells of the placenta infections, chromosomal rupture, umbilical cord - Peaks during 1st trimester of pregnancy (↑ blood, urine, amniotic fluid) abnormalities compression - Composed of 2 subunits: o Alpha = hCG, LH, FSH, TSH (identical subunits) SPECIMEN COLLECTION o Beta = Confers specificity for hCG - Method of collection = Amniocentesis o Up to 30 mL collected in sterile syringe HOME-BASED HCG PREGNANCY TEST KIT o 2nd trimester amniocentesis = Assess genetic defects (Ex: Trisomy 21/Down - Principle: Enzyme-immunoassay syndrome) - Specimen: 1st morning urine o 3rd trimester amniocentesis = Fetal lung maturity (FLM), Fetal hemolytic - Cut-off point: 25 mIU/mL disease (HDN) - Anti-hCG source: Rabbit - Quadruple screening tests prior to performing amniocentesis: o Alpha-fetoprotein CAUSES OF FALSE-POSITIVE AND FALSE-NEGATIVE PREGNANCY TESTS o Human chorionic gonadotropin (hCG) FALSE-POSITIVE FALSE-NEGATIVE o Unconjugated estriol (UE3) - Molar pregnancy, Midcycle LH - Too early (commonly) or too late o Inhibin A surge (rarely) testing - Hematuria or proteinuria - Dilute urine (low S.G) SPECIMEN HANDLING - Malignancies (gynecologic & other) - Adulterated urine - Test for Fetal Lung Maturity = Placed on ice on delivery, kept refrigerated or frozen, - Postpartum & post-abortion (up to - Ectopic pregnancy (rarely negative, Filtration prevents loss of phospholipids 4 weeks) almost always positive) - Test for Cytogenetic Studies = Kept at room temperature or at 37oC - Chinese herbal medications - Impending or missed abortion - Test for HDN = Protected from light - Perimenopausal (LH elevation) (rarely negative, almost always - Premature ovarian failure (LH positive) AMNIOTIC FLUID VS. MATERNAL URINE elevation) ANALYTE AMNIOTIC FLUID MATERNAL URINE Less reliable Protein + - HCG BIOASSAYS (BIOLOGICAL PREGNANCY TESTS) Glucose + - Test Animal Used Mode of Injection Positive Result More reliable Urea (mg/dL) < 30 >300 Hogben Female frog Lymph sac (urine) Oogenesis Creatinine < 3.5 >10 Galli-Mainini Male frog Subcutaneous Spermatogenesis (mg/dL) (urine/serum) Friedmann/ Virgin female Marginal ear vein Corpora lutea & FERN TEST Hoffman rabbit (urine) corpora - Detects ruptured amniotic membranes hemorrhagica - Also used to diagnose early pregnancy (↑ estrogen) Ascheim- Immature Subcutaneous Formation of - Procedure: Specimen (Vaginal Fluid) → Slide (Air Dry) → (+) Fern-like Zondek female mice (urine) hemorrhagic follicles crystals = AMNIOTIC FLUID (Due to presence of sodium chloride and & corpora lutea proteins) Frank- Immature Subcutaneous Ovarian hyperemia Berman female rats (urine) Kupperman Female virgin Intraperitoneal Ovarian hyperemia AMNIOTIC FLUID COLOR rat (urine) COLOR CLINICAL SIGNIFICANCE Kelso Female virgin Subcutaneous Ovarian hyperemia Colorless Normal rat (urine) Blood-streaked Traumatic tap, abdominal trauma, intra-amniotic hemorrhage Yellow HDN (Bilirubin) KEEP IN MIND! Dark-green Meconium = 1st fetal bowel movement (sign of distress) - ELISA tests are very sensitive, giving positive reactions as early as 10 days Dark red-brown Fetal death after conception - Urine specimen for pregnancy testing should have a specific gravity of at least 1. TEST FOR FETAL LUNG MATURITY 1.015 - Respiratory distress syndrome o Most frequent complication of early delivery o 7th most common cause of morbidity and mortality in the premature infant o Caused by insufficiency of lung surfactant (phospholipids) production & fetal lung immaturity - Surfactant normally appears in mature lungs and allows the alveoli to remain open throughout the cycle of inhalation and exhalation - Surfactant keeps the alveoli from collapsing by decreasing surface tension and allows them to inflate with air more easily  TEST INFORMATION REVIEW!!! 1. Lecithin/ - Reference method 1. Specimens for Fetal Lung Maturity (FLM) testing should be stored: Sphingomyelin - Lecithin = for alveolar stability a. At room temperature or at 37oC ratio - Sphingomyelin = serves as a control (due to constant b. Protected from light production) c. Refrigerated or frozen 𝐿𝑒𝑐𝑖𝑡ℎ𝑖𝑛 o L/S ratio is measured using thin layer d. Any of these 𝑆𝑝ℎ𝑖𝑛𝑔𝑜𝑚𝑦𝑒𝑙𝑖𝑛 chromatography (TLC) 2. Cause of dark red-brown amniotic fluid? ↑ Numerator = ↑ Ratio - Ratio of >2.0 = mature fetal lungs a. Meconium o Cannot be done on a specimen contaminated by b. HDN blood or meconium (falsely increases the ratio c. Intra-amniotic hemorrhage [Strasinger]) d. Fetal death 2. Amniostat- - Immunologic test for phosphatidylglycerol (PG) 3. Alpha fetoprotein is decreased in FLM o Not affected by blood or meconium a. Down syndrome o Production of PG is delayed among diabetic b. Spina bifida mothers c. Anencephaly 3. Foam stability - Amniotic fluid + 95% Ethanol → Shake for 15 secs → d. Hepatocellular carcinoma (Foam/Shake Stand for 15 mins 4. Sub-unit unique for HCG? test) - (+) Foam/Bubbles = MATURE FETAL LUNGS a. Alpha (Presence of phospholipids) b. Beta 4. Microviscosity - The presence of phospholipids decreases c. Gamma (Obsolete test) microviscosity d. Delta o Microviscosity is the friction experienced by a 5. Hogben bioassay test for hCG uses what animal? single particle undergoing diffusion because of its a. Male frog interaction with its environment at the micrometer b. Female toad length scale c. Female rabbit - Measured by fluorescence polarization d. Female rat o Surfactant to albumin (S/A) ratio is measured o Dye bound to surfactant had longer fluorescence & low polarization Answer key: C, D, A, B, B o Dye bound to albumin had decreased fluorescence & high polarization SPUTUM AND BRONCHOALVEOLAR LAVAGE 5. Lamellar body - Type ll pneumocytes produce alveolar surfactants count (LBC) stored in the form of lamellar bodies o Lamellar body diameter is similar to that of SPUTUM platelets, therefore, LBC can be obtained with the - Not a sterile body fluid use of platelet channel of hematology analyzers - From upper & lower (sterile) respiratory tract o LBC can be done using impedance and/or optical - Tracheobronchial secretions (mixture of plasma, electrolytes, mucin & water) scatter methods - Added with cellular exfoliations, nasal and salivary gland secretions and normal o >32,000/uL lamellar body count = ADEQUATE oral flora FLM - 95% water and 5% solids 6. OD 650 nm - ↑ Lamellar bodies = ↑ O.D. (Absorbance) - Secretions are viscoelastic - some of the properties of a liquid and some of a solid - O.D. of >0.150 is equivalent to: o L/S ratio of > 2.0 - Sialic acid - most important single component of sputum viscosity o The presence of PG - Acceptable sputum specimen = 25 WBC/LPF SPECIMEN COLLECTION 2. TEST FOR FETAL AGE - 1.5 to 2.0 mg/dL amniotic fluid creatinine = prior to 36 weeks' gestation - 1st morning = most preferred sample (most concentrated; routine) - >2.0 mg/dL amniotic fluid creatinine = 36 weeks (9 months) - 24-hour sputum = for volume measurement - Throat swab = for pediatric patients 3. TEST FOR HDN - Sputum induction = for non-cooperative patients - Tracheal aspiration = for debilitated or unconscious patients - A.k.a Optical Density (Absorbance) 450 - Specimen preservation methods = Refrigeration or 10% formalin - Absorbance of amniotic fluid: o Normal = ↑ at 365nm, ↓ at 550nm MACROSCOPIC EXAMINATION o HDN = ↑ at 450 nm (bilirubin) Volume ↓ Bronchial asthma, acute bronchitis, early pneumonia, stage of - Results are plotted on a Liley graph: healing o Zone I = Non-affected or mildly affected fetus ↑ Bronchiectasis, lung abscess, edema, gangrene, tuberculosis, o Zone II = Moderately affected fetus (requires close pulmonary hemorrhage monitoring) Color Colorless or Made up of mucus only o Zone III = Severely affected fetus (requires translucent intervention) White or yellow ↑ Pus (TB, bronchitis, jaundice, - Interferences= cells, meconium, debris, and hemoglobin (peak absorbance at pneumonia) 410nm) Gray ↑ Pus & epithelial cells - The oldest routinely performed lab test on AF evaluates the severity of fetal Bright green or ↑ Bile; P. aeruginosa infection, lung anemia due to HDN greenish abscess 4. TEST FOR NEURAL TUBE DEFECTS (NTD) Red or bright red Fresh blood or hemorrhage, TB, bronchiectasis - Spina bifida ("'split spine”) is a birth defect where there is incomplete closing of Anchovy sauce or Old blood, pneumonia, gangrene the backbone & membranes around the spinal cord. rusty brown - Anencephaly is the absence of a major portion of the brain, skull, and scalp that Prune juice Pneumonia, chronic lung cancer occurs during embryonic development - Screening test = Alpha-fetoprotein (AFP) Olive green or grass Cancer o ↑ in Neural tube defects green o ↓ in Down syndrome Black Dust or dirt, carbon, charcoal, - Confirmatory test = Acetylcholinesterase anthracosis, smoking - AFP is the major protein produced by the fetal liver during early gestation (prior to Rusty (with pus) Lobar pneumonia 18 weeks) (S. pneumoniae) Rusty (without pus) Congestive heart failure TESTS FOR FETAL WELL-BEING AND MATURITY Currant, jelly-like Klebsiella pneumoniae infection Test Normal Values at Significance Odor Odorless Normal Term Foul or putrid Lung gangrene, advanced necrotizing Bilirubin scan ∆ A450 >.025 Hemolytic disease of the tumors newborn Sweetish Bronchiectasis, tuberculosis Alpha-fetoprotein 2.0 Fetal lung maturity bacterial infection Amniostat-FLM Positive Fetal lung maturity/ Phosphatidylglycerol Consistency Mucoid Asthma, bronchitis Foam Stability Index >47 Fetal lung maturity Serous or frothy Lung edema Microviscosity (FLM-TDx) >55 mg/g Fetal lung maturity Mucopurulent Bronchiectasis, tuberculosis with cavities Optical Density 650 nm >0.150 Fetal lung maturity Lamellar body count >32,000/uL Fetal lung maturity  MACROSCOPIC STRUCTURES CLINICAL SIGNIFICANCE REVIEW!!! Dittrich' s Yellow or gray material, size of a Bronchitis, bronchiectasis 1. What is the most preferred sample for 4. Majority of the plugs pinhead Bronchial asthma routine sputum analysis? granulocyte seen in Produces foul odor when crushed a. Tracheal aspiration bronchoalveolar b. 1st morning sputum lavage is? Pneumoliths Hard concretions in a bronchus Histoplasmosis (most c. 24-hour sputum a. Neutrophil or (lung stones) common) d. Throat swab b. Macrophage Broncholiths Yellow/white calcified TB Chronic tuberculosis c. Lymphocyte structures/foreign material 2. Curschmann’s spirals, Creola bodies, d. Eosinophil Bronchial Branching tree-like casts of the Lobar pneumonia, Charcot-Leyden crystals, and Dittrichs plugs casts bronchi bronchitis, diphtheria are all associated with: 5. Sweat testing is Layer 1st (top) = frothy mucus Bronchiectasis, lung a. Broncholithiasis used to diagnose? formation 2nd (middle) = opaque, water abscess, gangrene b. Bronchiectasis a. Cystic fibrosis c. Bronchitis b. A and D (3 layers) material d. Bronchial asthma c. P. jirovecii 3rd (bottom) = pus, bacteria, pneumonia tissues 3. In the sputum analysis, the only significance d. Mucoviscidosis Foreign Bronchial calculi (calcium Pneumoconiosis of this finding is its resemblance to bodies carbonate & phosphate) Blastomyces? Asbestos bodies, silica particles a. Blue bodies b. Curschmann’s spirals MICROSCOPIC STRUCTURES CLINICAL c. Myelin globules d. Alveolar macrophage SIGNIFICANCE Elastic fibers Slender fibrils w/ double contour & Tuberculosis Answer key: B, D, C, A, B curled ends Charcot- Colorless, hexagonal, double pyramid, Bronchial asthma (3