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LECTURE NOTES 4: CLINICAL MICROSCOPY health problems, depending on where it occurs in the OVERVIEW: body. SAFETY IN THE CLINICAL LABORATORY RENAL FUNCTION TW...

LECTURE NOTES 4: CLINICAL MICROSCOPY health problems, depending on where it occurs in the OVERVIEW: body. SAFETY IN THE CLINICAL LABORATORY RENAL FUNCTION TWO TYPES OF INFECTION URINALYSIS (PHYSICAL, CHEMICAL, MICROSCOPIC) A. COMMUNICABLE INFECTION METABOLIC DISORDERS AND RENAL DISEASES It spread from one person to another through a SPUTUM & BRONCHOALVELAR LAVAGE variety of ways that include: contact with blood and SWEAT bodily fluids; breathing in an airborne virus; or by being AMNIOTIC FLUID bitten by an insect. CEREBROSPINAL FLUID SEMINAL FLUID B. NOSOCOMIAL INFECTION SYNOVIAL FLUID Also known as infections acquired during the SEROUS FLUID process of receiving health care that was not present GASTRIC FLUID during the time of admission. FECALYSIS CHAIN OF INFECTION CLINICAL MICROSCOPY The spread of an infection within a community is described as a “chain,” several interconnected steps that Clinical Microscopy utilizes microscopes to perform the describe how a pathogen moves about. Infection control scientific analysis of non-blood body fluids such as urine, and contact tracing are meant to break the chain, semen, and stool. preventing a pathogen from spreading. The spread of infection can be described as a chain with six links: The chemical and microscopic examination of urine (urinalysis) involves a number of tests to detect and CHAIN OF INFECTION measure compounds that pass through the urine. This INFECTIOUS Include not only bacteria but also provides valuable diagnostic information concerning AGENT viruses, fungi, and parasites. The metabolic dysfunctions of both renal and non-renal origin. virulence of these pathogens depends on: Analysis of stool samples (fecalysis) is used to measure number, things such as stool concentration and composition. potency Information from these tests can provide early detection ability to enter and survive in the of gastro-intestinal bleeding, uncover parasite exposure, body and identify liver and biliary duct disorders. Semen susceptibility of the host analysis is important for evaluating fertility and in post RESERVOIR Any person, animal, arthropod, plant, vasectomy. soil or substance (or combination of these) in which an infectious agent SAFETY IN THE LABORATORY normally lives and multiplies. PATHOGEN The infectious agent depends on the A pathogen is defined as an organism causing disease to reservoir for survival, where it can its host, with the severity of the disease symptoms reproduce itself in such manner that it referred to as virulence. They are taxonomically widely can be transmitted to a susceptible diverse and comprise viruses and bacteria as well as host. unicellular and multicellular eukaryotes ANIMATE RESERVOIRS include people, INFECTION insects, birds, and other animals. The invasion and growth of germs in the body. The germs may be bacteria, viruses, yeast, fungi, or other INANIMATE RESERVOIRS include soil, microorganisms. It can begin anywhere in the body and water, food, feces, intravenous fluid, may spread all through it. It can cause fever and other and equipment. PORTAL OF Means by which a pathogen exits from INFECTION CONTROL EXIT a reservoir. For a human reservoir, the An infection-control program has four main functions: portal of exit can include: 1. To protect patients, employees, and visitors from blood, infection. respiratory secretions 2. To screen employees for infectious diseases and to fluids from gastrointestinal or require immunization when needed. urinary tracts. 3. To provide evaluation and treatment to health workers MODE OF Once a pathogen has exited the who have been exposed to infections while performing TRANSMISSION reservoir, it needs a mode of their duty. transmission to transfer itself into a 4. To monitor employees and patients who are at risk of host. This is accomplished by entering infection and collect data from patients and health the host through a receptive portal of workers who have been exposed to such danger. entry. Transmission can be by: direct contact, UNIVERSAL PRECAUTION indirect contact All blood and bodily fluids should be treated as if they are through the air infectious, regardless of whether the source is known to PORTAL OF Infectious agents get into the body be infected. ENTRY through various portals of entry, including: STANDARD PRECAUTION mucous membranes Contact with urine, saliva, feces, vomit, nasal secretions, non-intact skin sputum, and even breast milk can be contagious and respiratory tract should be handled as such. Gastrointestinal tract genitourinary tract HAND HYGIENE It is an essential part of standard precautions in the Pathogens often enter the body of the healthcare setting. It is effective way to prevent infections host through the same route they that can be transmitted from the patient to healthcare exited the reservoir. personnel during procedures. SUSCEPTIBLE The final link; someone at risk of HOST infection. When a virulent pathogen ROUTINE HAND WASHING enters an immune-compromised Uses plain soap and water person, infection generally follows. Hands are visibly dirty Before eating Whether exposure to a pathogen After using restroom results in infection depends on several After known exposures to Clostridium difficile, factors related to the person exposed Bacillus anthracis; and infectious diarrhea during (the host), the pathogen (the agent), norovirus outbreaks and the environment. HAND ANTISEPSIS Host factors that influence the An antimicrobial soap or alcohol-based hand outcome of an exposure include: sanitizer to remove transient microorganisms presence or absence of natural Alcohol-based hand sanitizer is preferred when barriers hands are not visibly dirty functional state of the immune Put the sanitizer on hands, rub the hands together system for about 15 seconds or until it feels dry. presence or absence of an invasive device. STEPS IN PERFORMING THE ROUTINE HAND-WASHING TECHNIQUE  1. Stand a few inches from the sink to avoid Should be worn over the cuffs of the lab gown contamination. 2. Turn on the faucet and place hands under the PROPER REMOVAL OF GLOVES running water. After one glove is removed, the second glove can be 3. Use soap and work up lather to ensure that hand removed by sliding the index finger of the ungloved surfaces are reached. hand between the glove and the hand and slipping 4. Scrub for at least 15 seconds. Make sure to scrub all the second glove off. This technique prevents surfaces especially between the fingers and knuckles. contamination of the “clean” hand by the “dirty” 5. Apply a little friction rub hands together for at least second glove. 15 seconds. 6. Rinse the hands from wrist to fingers using a EYEWEAR downward motion. including face shields, goggles, and masks, should be 7. Dry hands using a clean paper towel. used when there is potential aerosol mists, splashes, 8. Use the paper towel to close the faucet, except when or spray to mucous membranes (mouth, eyes, nose). the letter is foot or motion activated. SEQUENCE FOR PUTTING ON PPE BY THE CENTERS FOR PERSONAL PROTECTIVE EQUIPMENT DISEASE CONTROL AND PREVENTION (CDC) Personal Protective Equipment is worn to minimize exposure to a variety of hazards. It may may include Donning and doffing is the practice of putting on and items such as gloves, safety glasses and shoes, earplugs or removing personal protective equipment. muffs, hard hats, respirators, or coveralls, vests. DONNING When entering the laboratory, appropriate clothing that Donning refers to putting on personal protective covers you from your shoulders to your ankles, and closed equipment. shoes with socks, are required. In general, the more skin covered the better. Long hair should be pulled back and held securely away from the face. Long nails should be cut to prevent easier contamination. Bulky and hanging jewelry should be removed. OUTER COVERINGS Includes gown, laboratory coats, and sleeve protectors, should be worn when there is a chance of splashing or spilling on work clothing. The outer covering must be made of fluid-resistant material, must be long-sleeved, and must remain buttoned at all times. GLOVES Must be worn when potential for contact with blood or body fluids exists (including when removing and handling bagged bio hazardous material and when decontaminating bench tops) Must be changed after contact with a patient, there is DOFFING visible contamination, and when physical damage Doffing refers to removing personal protective occurs. equipment. Should not be worn when “clean” devices are used Must not be worn again or washed  B. INGESTION - hands are not sanitized before handling food. Frequent hand washing Avoid hand-to-mouth activities Stop placing items in the mouth C. NON-INTACT SKIN - contamination through breaks or cuts in the skin Cover skin with non-permeable bandages D. PERCUTANEOUS - exposure through the skin due to injuries from needle sticks and other sharp objects Use needle safety devices Wear heavy-duty gloves when cleaning broken glass and never with bare hand E. PERMUCOSAL - infection through mucous membranes of the mouth and nose and the conjunctiva of the eyes Proper handling = avoid aerosols and splashes Avoid rubbing and touching the eyes, nose, and mouth PROPER WASTE DISPOSAL Improper waste disposal may severely endanger public health and/or the environment. The handling of hazardous waste must be regulated from the moment of generation until its disposal at its offsite final destination facility. A waste management system must be devised before work begins on any laboratory activity. Color Coding of Waste Containers according to the LABORATORY SAFETY RISKS Department of Health BIOHAZARD These are biological substances that pose a threat to the COLOR CONTAINER WASTE health of living organisms, primarily that of humans. This Red Sharps/Needles can include medical waste or samples of a microorganism, Yellow Infectious viruses, or toxins (from a biological source) that can affect Yellow with Black Band Chemical Wastes human health. Green Non-Infectious Wet Waste Black Infectious Dry Waste BIOSAFETY Orange Radioactive Waste It is a framework that describes the use of specific practices, training, safety equipment, and specially DISINFECTION designed buildings to protect the worker, community, and An appropriate disinfectant solution is household bleach. environment from an accidental exposure or Add 10 mL of bleach to 90 mL of water or 212 unintentional release of infectious agents and toxins. cups o bleach to 1 gallon of water to achieve the recommended concentration of chlorine 55 ppm BIOHAZARD EXPOSURE ROUTES: for sanitation and 600-800 ppm for disinfection A. AIRBORNE - from splashes and aerosols during Made fresh daily centrifuge and aliquot patients with airborne diseases. Labeled property with the name of solution, the Proper handling practice date and time prepared, the date and time of Wearing PPE properly especially masks Use safety shield and guards  expiration (24 hours), and the initials of CRY: Cryogenic (e.g. liquid nitrogen) preparer. POI: Poisonous (e.g. strychnine, alpha-amanitin) Not recommended for aluminum surfaces RA: Radioactive (e.g. plutonium, cobalt-60, carbon-14) FIRE HAZARDS 1. Enforcement of a no-smoking policy. 2. Installation of appropriate fire extinguishers. 3. Placement of adequate fire detection systems (alarms, sprinklers), which should be tested every 3 months. 4. Placement of manual fire alarm boxes near exit door. 5. Written fire prevention and response procedures 6. Fire drills, which should be conducted to that response to a fire situation response. CLASS OF FIRE & TYPE OF EXTINGUISHER TO BE USED WHEN A FIRE IS DISCOVERED: Rescue anyone in immediate danger Alarm the institutional fire alarm system Contain the potentially affected areas Extinguish the fire; exit the are if possible The National Fire Prevention Association (NFPA) is a global, non-profit organization that promotes safety standards, education, training, and advocacy on fire and electrical-related hazards. The NFPA 704 Diamond, commonly referred to as the NFPA Hazard Diamond, provides a system for identifying the specific hazards of a material and the severity of the hazard that would occur during an emergency response. URINARY SYSTEM For Specific Hazard: The urinary system's function is to filter blood and create ACID: Acid or alkaline, to be more specific; urine as a waste by-product. The organs of the urinary BIO: Biological hazard (e.g. flu virus, rabies virus); system include: COR: Corrosive; strong acid or base (e.g. sulfuric Kidneys: remove waste and extra water from the acid, potassium hydroxide); blood (as urine) and help keep chemicals (such as  sodium, potassium, and calcium) balanced in the body; make hormones that help control blood GLOMERULUS pressure and stimulate bone marrow to make red filtering unit of the kidney, is a specialized bundle of blood cells. capillaries that are uniquely situated between two Ureters: a bilateral thin tubular structures with a 3 to resistance vessels 4 mm diameter that connect the kidneys to the capillaries are each contained within the Bowman's urinary bladder. capsule and they are the only capillary beds in the Bladder: a subperitoneal, hollow muscular organ that body that are not surrounded by interstitial tissue acts as a reservoir for urine and is located in the located at the beginning of the nephron lesser pelvis when empty and extends into the resembles a sieve abdominal cavity when full has shield of negativity Urethra: the tube that lets urine leave your bladder non-selective filter of plasma substance with and your body molecular weight of 2000 mL/24hrs YELLOW ◆ Causes: Increase fluid BROWN: intake, Diabetes GREEN Pseudomonas infection mellitus, Diabetes BLUE GREEN oxidized phenol; indican insipidus ◆ RBCs (cloudy/smoked) OLIGURIA ◆ Decreased urine = hematuria volume ◆ Clear Red ◆ 20 disks F ◆ Increased number indicates /HP 0- 50- WBCs (0-8/HPF) Bacteria 0 10-50 >200 infection or inflammation F 10 200 ◆ largest cell w/abundnt, irregular /LP SQUAMOUS Mucus Threads 0 0-1 1-3 3-10 >10 cytoplasm & prominent nucleus F EPITHELIAL ◆ Clue cell: covered with Gardnella /LP Numerical Range: 0-2, 2-5, 5- CELL Casts vaginalis F 10, >10 TRANSITIONAL /HP ◆ spherical, polyhedral, or caudate RBCs EPITHELIAL F Numerical Range: 0-2, 2-5, 5-10, with centrally located nucleus CELL /HP 10-25, 25-50, 50-100, >100 WBCs ◆ most clinically significant F ◆ originated from nephrons Squamous E.C Rare, few, moderate or many/ LPF RENAL ◆ regular, polyhedral, cuboidal or Transitional Rare, few, moderate or many/ LPF TUBULAR columnar w/ eccentric nucleus E.C EPITHELIAL ◆ Oval Fat Bodies - nephrotic Renal tubular Average number per 10 HPF CELL syndrome E.C ◆ Bubble Cells - acute tubular Oval Fat Bodies Average number per HPF necrosis Abnormal Average number per LPF ◆ TRUE UTI = Bacteria + WBCs Crystals ◆ E. Coli = most common cause of BACTERIA UTI CASTS ◆ Bacteria only = contamination/old ◆ Cylinduria - excretion of casts specimen ◆ Unique to kidney ◆ TRUE YEAST INFECTION = Yeast + ◆ Formed in PCT and CD WBCs YEASTS ◆ Candida albicans = Diabetes CAST DESCRIPTIONS mellitus, Vaginal moniliasis HYALINE ◆ Most frequently encountered & PARASITES ◆ Trichomonas vaginalis = most (0-2/LPF) most difficult cast to discover  ◆ Pathologic = glomerulonephritis, pyelonephritis ◆ Most fragile cast RBC CAST ◆ Indicates bleeding with nephron ◆ Muddy brown cast BLOOD CAST ◆ Hemoglobin from lysed RBCs ◆ Pus cast ◆ Indicates inflammation or WBC CAST infection within nephron ◆ Pathologoic = pyelonephritis, acute interstitial nephritis ◆ Cells visible on the cast matrix are RTE CAST small, round, oval cells ◆ Pathologic = tubular damage ◆ TRUE UTI = Bacteria + WBCs ◆ E. Coli = most common cause of BACTERIAL UTI CAST ◆ Bacteria only = contamination/old NORMAL DESCRIPTIONS specimen ALKALINE ◆ Derived from lysosomes of RTE CRYSTALS cells during normal metabolism ◆ Most common cause of turbidity in GRANULAR ◆ Finely granular cast has a AMORPHOUS urine CAST sandpaper appearance PHOSPHATES ◆ Fine, lacy white precipitate ◆ Pathologic = glomerulonephritis, AMMONIUM ◆ “thorny apple” appearance pyelonephritis BIURATE ◆ Seen in old specimen ◆ Pathologic = Nephrotic syndrome, FATTY CAST ◆ Colorless, prism shaped, “coffin- diabetes mellitus TRIPLE lid” appearance ◆ Final degenerative form of all PHOSPHATE ◆ Feather appearance when types of casts disintegrated ◆ Brittle, highly refractile, with WAXY CAST MAGNESIUM jagged ends ◆ Colorless, rhomboid plates PHOSPHATE ◆ Ground glass appearance ◆ Apatite ◆ Pathologic = chronic renal failure ◆ Dibasic calcium phosphate ◆ Renal failure Cast (stellar): colorless, flat plates, thin ◆ Indicates destruction (widening) of CALCIUM BROAD CAST prisms in rosette form the tubular walls PHOSPHATE ◆ Monobasic calcium phosphate: ◆ Pathologic = renal failure irregular, granular appearing sheets or plates CRYSTALS ◆ Small, colorless, dumbbell, tetrads CALCIUM ◆ Crystaluria- excretion of crystals ◆ Forms effervescence after adding CARBONATE ◆ Most recognized but most insignificant sediment acetic acid ◆ Formed from precipitation of urine solutes ◆ Factors for crystal formation: pH, solute NORMAL ACID DESCRIPTIONS concentration, temperature CRYSTALS ◆ Fluffy orange or pink sedeiment AMORPHOUS (brick dust) = uroerythrin URATES ◆ Microscopic = yellow brown  granules ◆ Pathologic = increase in Gout METABOLIC DISORDERS ◆ Most pleomorphic A. PHENYLKETONURIA ◆ Product of purine metabolism Most well known aminoacidurias URIC ACID ◆ Rhombic (diamond), lemon-shaped (-) gene that codes for phenylalanine ◆ Pathologic = Lesch-Nyhan hydroxylase syndrome Mousy urine, sweat and breath odor ◆ Most frequently observed Screening Test: Guthrie bacterial inhibition test ◆ Dihydrate (Weddellite) - envelope, B. Subtilis is cultured with beta-2- CALCIUM bipyramidal, octahedron thienylalanine (TE) and inhibits the growth OXALATE ◆ Monohydrate (Whewellite) - oval/ of B. subtilis. Phenylalanine counteracts the dumbbell action of beta-2-TE. CALCIUM Confirmatory test: Ion exchange HPLC ◆ “cigarette butty appearance” SULFATE B. TYROSYLURIA/TYROSINEMIA HIPPURIC ACID ◆ Colorless elongated prism (-) gene that codes for: MONOSODIUM ◆ Tiny, slender, colorless needles Type 1: fumarylacetoacetate hydrolase URATES ◆ Pathologic = Acute & Chronic gout (FAH) Type 2: tyrosine aminotransferase ABNORMAL DESCRIPTIONS Type 3: p-hydroxyphenylpyruvic acid ACID dioxygenase CRYSTALS Seen in liver disease ◆ Refractile hexagonal plate, often Rancid butter urine odor CYSTINE laminated Screening Test: Nitroso-napthol = (+) orange red ◆ Rectangular plates with notch in Confirmatory Test: Chromatography one or more corners C. ALKAPTONURIA CHOLESTEROL ◆ “staircase pattern” (-) gene that codes for homeogenistic acid ◆ Pathologic - increase in nephroctic oxidase syndrom Urine darkens after becoming alkaline from RADIOGRAPHI ◆ Fla four-sided plates often with standing atroom temperature C DYE notched corners Reddish-stained disposable (plastic) diapers ◆ Fine colorless to yellow needles in Screening test: FeCl3 tube test = (+) transient clumps or rosettes blue TYROSINE D. MELANURIA ◆ Pathologic - increase in liver disease Caused by melanoma ◆ Yellow brown oily looking sphere Tumors secrete 5,6-dihydroxyindole (oxidizes LEUCINE with concentric circles and radial melanogen to melanin) striations Urine darkens upon air exposure ◆ Clumped granules and needls with Screening test: FeCl3 tube test = (+) gray/black bright yellow color precipitate BILIRUBIN E. MAPLES SYRUP URINE DISORDER (MUSD) ◆ Pathologic - increase in liver disease Most common inborn error of metabolism ◆ “sheaves of wheat” appearannce (-) gene that codes for the enzume complex SULFONAMIDE known as branched-chain-alpha-keto acid ◆ Possible tubular damage dehydrogenase (BKCD) ◆ Colorless needles = form following Caramelized sugar/maple syrup/ curry urine refrigeration AMPICILLIN odor ◆ Pathologic - massive dose of Screening Test: 2,4-dinitrophenylhydrazine penicillin (DNPH) = (+) yellow turbidity/precipitate  Confirmatory: GCMS Findings: Macroscopic hematuria, protenuria, F. HARTNUP DISEASE glucosuria, cellular & granular cast, waxy & “Blue-diaper syndrome” broad casts Indigo blue urine color upon air exposure G. NEPHROTIC SYNDROME Screening test: Obermayer’s test Disruption of the electrical charges that produce G. LESCH NYHAN DISEASE the tightly fitting podocyte barrier resulting in (-) gene that codes for enzyme hypoxanthine massive loss of proteins & lipids guanine phosphoribosyltransferase Findings: oval fat bodies, fatty & waxy cast “Orange Sand in diaper” due to increase uric acid in blood and urine INTERSTITIAL DISORDER A. CYSTITIS GLOMERULAR DISORDER Ascending bacterial infection of the urinary A. GOODPASTURE SYNDROME bladder Deposition of antiglomerular basement Acute onset of urinary frequency and burning membrane antibody to glomerular and alveolar Findings: WBCs, Bacteria, Microscopic basement membrane hematuria Findings: Macroscopic hematuria, protenuria, B. ACUTE PYELONEPHRITIS RBC casts Infection of the renal tubules & interstitum B. WEGENER’S GRANULOMATOSIS Findings: WBCs, Bacteria, Microscopic Anti-neutrophilic cytoplasmic auto-antibody hematuria, WBC cast, bacterial cast (ANCA) binds to neutrophils in vascular walls C. CHRONIC PYELONEPHRITIS producing damage to small vessels in the lungs Reccurent infection of the renal tubules & and in the glomerulus interstitium caused by structural abnormalities Findings: Macroscopic hematuria, protenuria, affecting flow of urine RBC casts Findings: WBCs, Bacteria, Microscopic C. HENOCH SCHONLEIN PURPURA hematuria, WBC cast, bacterial cast, granular Occurs in children following viral respiratory cast, waxy & broad cast infections D. ACUTE INTERSTITIAL NEPHRITIS Decrease in platelets disrupts vascular integrity Allergic inflammation of the renal interstitum in Findings: Macroscopic hematuria, protenuria, response to certain medication RBC casts Findings: WBCs, Bacteria, Microscopic D. BERGER’S DISEASE hematuria, Increased eosinophils, WBC cast Deposition of IgA on the glomerular membrane resulting from increased levels of IgA TUBULAR DISORDER Findings: Macroscopic hematuria, protenuria, A. ACUTE TUBULAR NECROSIS glucosuria, cellular & granular cast, waxy & Damage to renal tubular cells caused by broad casts ischemia or toxic agents E. ALPORT SYNDROME Odorless urine Giant platelet syndrome Findings: Macroscopic hematuria, protenuria, Genetic disorder showing lamellated and RTE cells & catss, hyaline, granular, waxy, & thinning of glomerular basement membrane broad casts Findings: Oval fat bodies, fatty & waxy B. FANCONI SYNDROME F. CHRONIC GLOMERULONEPHRITIS Generalized failure of tubular reabsorption in Marked decrease in renal function resulting the proximal convulated tubule from glomerular damage precipitated by other Findings: Glucosuria, cystine crystals renal disorders C. DIABETES INSPIDUS Neurogenic: hypothalamus fails to produce to ADH  Nephrogenic: renal tubules fails to respond to ◆ Measured using thin layer chromatography ADH ◆ Ratio: > 2.0 = Mature Fetal Lungs Findings: low SG, polyuria Test for Hemolytic Disease of the Newborn D. RENAL GLUCOSURIA OPTICAL DENSITY 450 Normal Blood Glucose, Increased Urine glucose ◆ Absorbance of amniotic fluid Defective tubular reabsorption of glucose ◆ Normal: increase at 365 nm and decreased Findings: Glucosuria at 550 nm ◆ HDN: increased at 450 nm AMNIOTIC FLUID ◆ Results plotted on Liley graph Present in amnion - membranous sac that surrounds Zone I: non affected or mildly affected the fetus fetus The placenta is the ultimate source of amniotic fluid Zone II: moderately affected fetus water and solutes Zone III: severely affected fetus Functions: cushion for fetus; stabilizes temperature; Test for Neural Tube Defects allows fetal movement; proper lung development SPINA BIFIDA (“split spine”) - birth defect where Amniotic Fluid Volume - from fetal urine and lung there is incomplete closing of the backbone and fluid membranes of the spinal cord Normal: 800 - 1200 mL (3 rd trimester - fetal urine ANCEPHALY - absence of major portion of the as major contributor) brain, skull, and scalp that occurs during ◆ 1st trimester: 35 mL (from maternal embryonic development circulation) Screening Test: Alpha-feto protein (major protein Specimen collection: AMNIOCENTESIS produced by fetal liver during early gestation) Up to 30 mL collected in sterile syringe ◆ Increased in neural tube defects 2 nd trimester: assessment of genetic defects ◆ Decreased in down syndrome rd 3 trimester: fetal lung maturity Confirmatory test: Acetylcholinesterase Specimen Handling Test for Fetal Lung Maturity: placed on ice on SPUTUM delivery, kept refrigerated or frozen From upper and lower respiratory tract Test Hemolytic Disease of the Newborn: protected Tracheobronchial secretions (mixture of plasma, from light electrolytes, mucin & water) FERN TEST: detects ruptured amniotic membranes Secretion are viscoelastic and also used to diagnose early pregnancy Acceptable specimen = 25 WBC/LPF Specimen: Vaginal Fluid (air dried on slide) SIALIC ACID - important single component of sputum Result: (+) fern-like crystalls = Amniotic Fluid viscosity Amniotic Fluid Color SPECIMEN COLLECTION: First morning specimen Colorless to pale yellow - normal (most preferred routine sample) Blood-streaked - traumatic tap Tracheal aspiration: for debilitated or Yellow - HDN unconscious patients Dark green = meconium (1st fetal bowel Specimen Preservation: Refrigeration or 10% movement) formalin Dark red-brown - fetal death SPECIMEN COLOR Test for Fetal Lung Maturity Red/ Bright Red - Fresh blood or hemmorrhage LECITHIN/SPHINGOMYELIN (L/S) RATIO: Anchovy sauce/ Rusty brown - old blood, ◆ Reference method pneumonia ◆ Lecithin (for alveolar stability) Olive green/Grass green - cancer ◆ Sphingomyelin (serves as control) due to Rusty with pus - Lobar pneumonia constant production Rusty w/o pus - congestive heart failure  Currant, jelly-like - Klebsiella pneumoniae Pia Mater: innermost layer; lines the surface of DITRICH’S PLUGS: yellow or gray material; Bronchial the brain & spinal cord asthma Choroid Plexus: produces CSF by selective filtration BRONCHOLITHS: hard concretion in a bronchus at 20 mL/hr (lung stones); Histoplasmosis Arachnoid Villi: reabsorbs CSF CHARCOT-LEYDEN CRYSTALS - colorless, hexagonal, Blood Brain Barrier: protects brain from chemicals double pyramid, often needle-like = Bronchial circulating in the blood that can harm the brain asthma tissue CURSCHMANN’S SPIRALS - coiled mucus strands = Specimen Collection: Lumbar puncture (between L3- Bronchial asthma L4 for adults or L4- L5 for infants) MYELIN GLOBULES = colorless globules occuring in CSF TUBES variety of sizes = Mistaken as Blastomyces Tube 1: Chemistry/ Serology CREOLA BODIES = clusters of columnar epithelial Tube 2: Microbiology cells = Bronchial asthma Tube 3: Hematology Tube 4: Microbiology/Serology BRONCHOALVEOLAR LAVAGE CSF Total Volume Procedure for collecting the cellular milie of the Adults = 90 - 150 mL alveoli by use of a bronchoscope through which Neonates = 10 - 60 mL saline is instilled CSF Appearance Diagnostic test for Pneumocystits carinii in Crystal Clear = Normal immunocompromised patients Turbid = Increase in WBCs Xanthochromic SWEAT ◆ PINK = slight amt of oxyhemoglobin CYSTIC FIBROSIS - specimen for diagnosis ◆ YELLOW = oxyhemoglobin -> bilirubin Autosomal recessive metabolic disorder affecting ◆ ORANGE = heavy hemolysis the mucous secreting glands of the body Bloody = Increase in RBCs Associated with pancreatic insufficiency, Pellicle = Tubercular meningitis respiratory distress & intestinal obstruction TRAUMATIC TAP GIBSON AND COOKIE PILOCARPINE Uneven blood distribution on three tubes IONTOPHORESIS Clot formation due to plasma fibrinogen Pilocarpine + mild current = induce sweat Clear supernatant production INTRACRANIAL HEMORRHAGE Application of 0.15 mAcurrent for 45 minutes Even blood distribution on three tubes Sweat is tested for sodium and chloride No clot formation >70 mEq/L = diagnostic for CF Xantochromic CELL COUNT CEREBROSPINAL FLUID Performed immediately 3RD major body fluid WBC Count = routinely performed Functions: supply nutrient to the nervous system, ◆ Normal values remove metabolic waste, produce mechanical barrier ◆ Adults = 0-5 WBCs/uL to cushion the brain & spinal cord against trauma ◆ Neonates n= 0 -30 WBCs/uL Meninges: Line the brain and spinal cord RBC Count = done only in traumatic tap Dura Mater: outer layer; lines the skull and ◆ 1 WBC for every 700 RBCs seen vertebral canal Lymphocytes & Monocytes = predominant cells in Arachnoid: spiderweb-like; filamentous inner CSF membrane ◆ 70:30 in adults ◆ Subarachnoid space: below arachnoid; ◆ Up to 80% monocytes in neonates portion where CSF flows  ◆ Indication of viral, tubercular & fungal Methods of collection meningitis ◆ Masturbation Neutrophils = occasinal ◆ Coitus interruptus ◆ Indication of bacterial meningitis ◆ Condom method CSF Protein = 15-45 mg/dL (Adults) & 150 mg/dL Specimen should be delivered to lab within 1 hr (Infants) at room temp Increased in damage to blood brain barrier (most Analysis should be done after liquefaction (30-60 common cause) = meningitis & hemmorhage minutes) Production of immunoglobulins within the CNS = Specimen awaiting analysis should be kept at 37 multiple sclerosis deg C Major CSF Protein = Albumin Seminal Fluid Appearance CSF Glucose = 60-70% or 50-80 mg/dL (normal) Normal = gray white, translucent specimen for blood glucose should be drawn 2 Increased white turbidity = Infection hours prior to spinal tap Red or brown = Increased RBCs Normal in viral meningitis Seminal Fluid Volume Increased when plasma glucose is increased Normal = 2-5 mL Decreased in bacterial, tubercular, and fgungal Increased = Overabstinence meningitis Decreased = Infertility CSF Lactate = 10-22 mg/dL (normal) Seminal Fluid Viscosity Inversely proportionate with glucose Normal = pour in droplets Normal in viral meningitis Increased = decreased motility Increase in bacterial meningitis, tubercular Reporting: 0 = Watery; 4 = gel-Like meningitis Seminal Fluid pH Normal = 7.2 - 8.0 SEMINAL FLUID Increased = infection Seminal fluid analysis is done for fertility testing, Decreased = increased prostatic fluid post-vasectomy semen analysis, forensic analysis Sperm Concentration = > 20 million sperms/mL (alleged rape) (normal) Semen is composed of 5% Spermatozoa, 60-70% Method: Improved Nebauer Counting Chamber Seminal Fluid, 20-30% Prostate Fluid, 5% ◆ Dilution = 1:20 (distilled water as diluent to Bulbuorethral gland immobilize sperm) Spermatogenesis and sperm maturation takes 90 Computation days. ◆ 2 WBC squares Seminiferous Tubules (testes) = site of # sperms counted x 100,000 spermatogenesis ◆ 5 WBC squares ◆ Sertoli cells = nurse cells for developing # sperms counted x 1,000,000 sperm Sperm Count = >40 million per ejaculate Epididymis = site of sperm maturation Computation: Sperm Concentration x Specimen ◆ Stages of Sperm Maturation: Volume Spermatogonium Sperm Motility = >50% motility (within 1 hr), quality 1 Spermatocyte = >2.0 2 Spermatocyte Place drop of semen in slide & cover with Spermatid coverslip. Allow to settle for 1 min then observe Spermatozoon in 20 HPF Specimen Collection Sperm Morphology = >30% normal forms in routine Abstinence = 2 to 3 days but nor > 7 days criteria Collect entire ejaculate Papanicolaou’s stain = for staining  x zHead: oval shaped (normal); poor ovum Presence of crystals = Milky penetration (abnormal) Synovial Fluid Viscosity = 4-6 cm string (normal) Acrosomal cap: 1/2 of the head, 2/3 of the nucleus Mucin Clot Test (Ropes) = Hyaluronic Polymerization Midpiece: contain mitochondria Reagent: 2-5% acetic acid Tail: 45 um in length (includes neck - thickest part CELL COUNT of tail); abnormal (poor motility) WBC Count = frequently performed ( Pernicious anemia Anacidity = failure to produce a pH Pernicous anemia FECALYSIS Around 100 to 200g of stool is passed per day Human feces contains around 75% water & 25% solids Odor of feces = presence of indole and skatole Macroscopic Apperance of Stool Normal: brown Black: Upper gastrointestinal bleeding Red: Lower gastrointestinal bleeding Pale yellow: Bile duct obstruction Frothy: Pancreatic disorder Rice water: Cholera Pea-soup: Typhoid Bristol Stool Chart Type 1: Separate hard lumps like nuts Type 2: Sausage-shaped but lumpy Type 3: Like sausage but with cracks Type 4: Like a sausage or snake, smooth & soft Type 5: Soft blobs with clear cut edges (passed easily) Type 6: Fluffy pieces with ragged edges, a mushy stool Type 7: Watery, no solid pieces, entirely liquid Microscopic Examination Fats = Steatorrhea (increased by >6/day) Muscle Fibers = Creatorrhea (abnormal excretion of muscle fibers in feces Fecal Leukocytes = >3 neutrophils/LPF (invasive condition) ◆ Diarrhea with WBCs = Salmonella, Shigella, Yersinua, E.coli ◆ Diarrhea w/out WBCs = S. aureus, V. cholerae