I. RISK MANAGEMENT AND SAFETY IN THE HISTOPHATHOLOGY LABORATORY

Risk management

– pertains to the process of ensuring

and maintaining personal as well as environmental health

and safety in the laboratory. And it is everyone’s

responsibility to minimize the risk associated with day to day

activity by using safety guards and checking the quality of

the reagent.

Histology (tissue) (study)

Origin: Greek words histos (_____) + logos (_____)

microscopic study of the normal tissues of the body

Histopathology (suffering)

Origin: Greek word pathos

(______)

microscopic study of

tissues affected by

disease

Focus: Microscopic

changes due to

inflammation, infection,

degeneration, tumors, or

cancer.

Goal: Correlation of

microscopic findings with

clinical conditions

Histopathologic

Techniques

the procedures adopted

for the preparation of

material for such studies

Histotechnology

science centering on the

microscopic detection of

tissue abnormalities for

disease diagnosis and the

treatment of diseases

MedTechs assigned in the

histopath section.

Responsible for preparing

specimens from surgery

or biopsy for microscopic

screening.

Produce prepared

mounted slides evaluated

by pathologists (in the

Philippines).

In some countries (e.g.,

US), trained MedTechs can

read slides.

Histotechnologists

promote disease

prevention and health

awareness.

Histotechnologists

prepares surgical

specimens for

microscopic screening by

the surgical pathologist

Diagnostic

Cytology

microscopic examination of cells different body sites ex. fluids in the body

Exfoliative Cytology

microscopic study of cells that have been desquamated from epithelial surfaces ex. cervico-vaginal smears for Pap smear stained with Papanikolau stain

HAZARDS IN THE HISTOPATHOLOGY LABORATORY

● Histopathology is considered one of the

most dangerous areas in the clinical

laboratory.

First step in risk management:

Identify potential hazards and remain aware in the workplace.

CHEMICAL HAZARDS

Cleaning agents and disinfectants, drugs,

anesthetic gases, solvents, paints, and

compressed gases

● Safety Data Sheet (SDS):

○ Contains severity, usage, and

precautions.

○ Required in every laboratory.

(1-9)

DOT labeling includes four-digit hazard number (_____)

LABELING STANDARDS

● DOT labeling includes four-digit hazard number (1–9).

(Health Hazard) (Fire Hazard) (Reactivity) (Specofoc hazards) (no hazard) (severe hazard)

● NFPA (National Fire Protection Association):

○ Blue: (_______)

○ Red: (_________)

○Yellow: (________)

○ White: (___________)

○ Rating: 0 = (________), 4 = (_______)

Permissible Exposure Limit (PEL)

Legal limit set by OSHA

Threshold Limit Values (TLV)

Recommended upper limit set

by ACGIH (guideline).

Occupational Exposure Limits (OEL)

General term for any

regulatory or recommended

limit.

(Time Weighted Average (TWA / TWAEV) (Short Term Exposure Limit (STEL / STEV): (Ceiling Limit (CL / CEV) (Immediate Danger to Life or Health (IDLH):

TYPES OF TLV

	8 hours
	over 15 minutes (always higher than TWA).
	sudden, extreme or maximum chemical exposure at any moment.

	sudden, extreme danger, not routine work

(human carcinogen) (0.75ppm) (2ppm) (0.5 ppm over 8 hours)

EXAMPLE: Formalin

● Classfied as (__________) (nose &

throat cancer).

● TWA: (_______)

● STEL: (________)

● Action level: (__________)

● Control measures:

○ Engineering controls (ventilation)

○ Change work practices

○ Provide respirators or substitutes

(Chemical) (ingredients) (Manufacturer’s) (address) (making) (purchased) (Expiration) (warnings) (procedures)

LABELING

● (________) name and, if a mixture, names of

all (_______);

● (__________) name and (_______) if

purchased commercially, or name of

person (_______) the reagent;

● Date (________) or made

● (______) date, if known;

● Hazard (______) and safety (_________)

Irritants

CHEMICAL CATEGORIES

cause reversible inflammatory effects at the site of contact with living tissue, especially the skin, eyes and respiratory passage Ex. Xylene, Chloroform

Corrosive Chemicals

CHEMICAL CATEGORIES

cause destruction or

irreversible alterations when

exposed to living tissue, or

destroy certain inanimate

surfaces (generally metal)

Ex. Hydrochloric acid,

Glutaraldehyde

Sensitizers

CHEMICAL CATEGORIES

cause allergic reactions in some exposed workers, not just in hypersensitive individuals Ex. Formaldehyde (once sensitized, always sensitive)

(Carcinogens) (Carcinogenic dyes)

CHEMICAL CATEGORIES

(________)

substance that induce tumors, not only in experimental animals but also in humans Ex. Chloroform, Chromic acid, Formaldehyde, Nickel chloride, Potassium dichromate

(___________): Auramine, Basic Fuchsin, Congo Red, Diamino Benzidine

Toxic

Materials

CHEMICAL CATEGORIES

capable of causing death by ingestion, skin contact or inhalation at certain specified concentrations. Ex. Methanol, Chromic acid, Osmium tetroxide, Uranil nitrate

Combustibles

CHEMICAL CATEGORIES

have flash points at or above a specified temperature. Burn only at higher heat. Ex. Silene

Flammable

CHEMICAL CATEGORIES

have flash points below the specified temperature. Ignite easily. Ex. Flammable clearing agents

Explosive

CHEMICAL CATEGORIES

primary example: picric acid,

some silver solutions

Oxidizers

CHEMICAL CATEGORIES

initiate or promote combustion in other materials Ex. Sodium iodate, Mercuric oxide, Chromic acid, Organic peroxides

Apparatus or spills of toxic substances in the fume hood

STORAGE

Pans or trays made of polyethylene or other chemically resistant material

All volatile substances having high chronic toxicity

STORAGE

Ventilated storage area

Dangerous

liquids

STORAGE

Best stored below countertop height Fire-resistant cabinets approved by OSHA.

Dangerous reagents

STORAGE

Stored in plastic or plastic

coated glass bottle

Certain flammable liquids that present unusual fire and explosion risk

STORAGE

must NEVER be stored in

refrigerator or freezer

Flammable liquids

STORAGE

cabinets and safety containers that are approved by the OSHA; should be kept in a fire-resistant cabinet and not in areas exposed to direct sunlight

Concentrated bottles

STORAGE

kept in locked metal cabinets

Caustic and corrosive materials

STORAGE

stored near the floor or close to the ground

Light sensitive chemicals

STORAGE

placed in dark containers in cool and dark storage areas

Acids

STORAGE

stored by themselves in an acid cabinet away from formaldehyde, bases, alcohols and oxidizers

Bleach

STORAGE

should not be stored next to

acids or methanol

Acetone, Xylene, Alcohol

STORAGE

should never be stored inside

a regular laboratory

refrigerator

(Dilution process) (Add acid to water) (chlorine gas release) (irritant and corrosives) (chlorine gas release) (Cryogens such as liquid nitrogen, as well as dry ice) (

STORAGE OF HAZARDOUS CHEMICALS

● (________) should be done under a

chemical fume hood.

● (________), never water to acid.

● Avoid mixing chlorine solutions to prevent

(_________).

● Hypochlorite solutions are classified as

(_____).

● NEVER MIX different chlorine solutions or

store them with cleaning products to

prevent (3) (________).

● (______________________), can be hazardous and should

NEVER be handled with barehands.

● To dispose of dry ice, allow it to sublimate

or evaporate to the atmosphere in a

wellventilated area where CO2 vapor

cannot build up.

(slips and falls) (ay and before leaving) (safe walking surface) (organized) (stacked, blocked, interlocked) (clean) (puncture-proof containers.)

PHYSICAL HAZARDS

● The most obvious physical hazards are (_______) from working in wet locations and the ergonomic hazards of lifting, pushing, pulling and repetitive tasks.

● Clean the work area throughout the (________&_______) the lab for the day.

● Keep all aisles and walkways in the lab clear to provide a (_________) and an unobstructed exit.

● Material storage should be (________).

● Bags, containers, and bundle stored in tiers should be (_____, ______, ____) and limited in height.

● keep workspace (_______)

● Discard sharps in (_______________)

(Electric shocks, arc blasts, electrocution, fires and explosions) (6 feet) (ground-fault circuit interrupters (GFCIs) (checked) (flexible extension cords) (DO NOT TOUCH THEM) (

ELECTRICAL HAZARDS

● (_______________________________).

● Wet locations: Within (________) of water sources.

● First, equip all electrical power outlets in wet locations with (_______________) to prevent accidental electrocutions.

● Before turning equipment on, all power cords must be (________) to be sure that they are in good condition.

● Improper use of (_______) -

● If you see a person being electrocuted, (_______________)!; use non-conductive item (e.g., wooden broom).

(humans) (Specimens, body contaminated solutions, or objects fluids) (Grossing) (Fixed tissues) (hallways) (freezers or in refrigerators) (appropriate)

BIOLOGICAL HAZARDS

● anything that can cause disease in (_______), regardless of their source.

● Sources: (_____, _________, _________),

● High-risk activity: (__________) of specimens.

● Lower risk: (________) (fixative deactivates most infectious agents except prions).

● If biological materials are used in the area, they should not be stored in (________), in unlocked (__________).

● Biohazard signs should be placed in (___________) areas.

PRIONS

● are infectious agents that cause spongiform encephalopathies such as Creutzfeldt-Jakob Disease (CJD), scrapie, and mad cow disease.

● Resistant to standard sterilization and chemical decontamination.

● Special treatment for suspected CJD tissue:

○ Immerse in formalin 48 hours

○ Treat in concentrated formic acid 1 hour

○ Additional formalin fixation 48 hours

(Inhalation) (non-intact skin) (mucous membranes)

TRANSMISSION OF BIOLOGICAL HAZARDS

● (_________) of aerosols

● Contact with (________)

● Contact with (____________)

(neutralizing and containment) (latex or nitrile) (plastic aprons) (biohazards) (dustpan and brush) (sponges) (towels and mops) (adsorbent) (bleach) (baking soda) (vinegar) (neutralizing) (sealable) (heavy)

HANDLING SPILLS

● Spill neutralizing and containment kits

○ good quality (_______) gloves

○ disposable (_________) for chemical spills

○ disposable gowns for (_________)

○ (_________) for powders

○ (_____)

○ (__________)

○ (________) material

○ (______)

○ (______) for acids

○ (______) for alkalis

○ commercial (_______) product

○ (________) plastic bucket

○ (_______) plastic bags

(wipe with gloves; dispose) (evacuate personnel) (trained emergency) (10%) (thoroughly)

HANDLING SPILLS

● Small spill: Few grams/milliliters; (___________) as hazardous waste.

● Significant spill: Large or toxic; (________); only (_________) response team handles cleanup.

● Decontamination: (____) bleach solution; avoid dust creation.

● After cleanup: Wash hands (____)

(Annual) (accessible and regularly refilled) (source ) (fresh air) (bag and mask) (Advanced Cardiovascular Life Support (ACLS) (100%) (clothing and footwear)

FIRST AID MEASURES

● (_______) safety training recommended.

● First aid kits must be (_______________).

● Exposed to chemical, infectious or toxic waste products:

○ Immediately remove the person from the (_____) of contamination and move to (______).

○ If the person is not breathing, do not use mouth to mouth, or mouth to nose ventilation, because of the danger to the rescuer. Instead, use a resuscitation (_______).

○ If pulse is absent, start external cardiac massage and follow standard (_______________) guidelines

■ Give (________) oxygen by mask if available. '

■ Remove all contaminated (______&_____) into a sealable collection bag and launder contaminated clothing thoroughly.

(water for 15-30 minutes.) (eye wash) (soap) (water) (re-use) (medical assistance)

● For accidental skin contact with hazardous chemicals

○ the affected area should be washed with copious amounts of (___________)

○ Emergency showers should be as accessible as (_____) stations. If the hazardous substance is not readily water-soluble, use (________) with the (______) wash.

○ Immediately remove contaminated clothing and launder before (______).

○ Seek (____________) following skin contact.

(15-30 minutes) (100 feet) (15–35°C.) (not recommended)

● For accidental splashing

○ the affected eye should be rinsed for (_________)

○ Eye wash stations: No more than (______) from hazard; temperature (____)

○ Portable bottles (__________) (insufficient liquid, contamination risk).

(Awkward postures:) (Contact stress:) (Duration & Repetition:) (Static postures:)

ERGONOMICS

● (____________) Bending wrists, leaning forward.

● (____________) Resting body part against hard/sharp surface

● (___________) Long exposure increases risk

● (_________) Holding position for long periods causes strain.

(specifications electrical) (use) (shower and first aid kits) (detailed list) (volatile chemicals) (sufficient ventilation) (heat resistant cabinet) (materials safety data sheet) (labeled) (appropriate protective gears)

lab. They can be distracting and can increase the potential for an accident to occur. They can also become contaminated if handled while working with hazardous materials.

● Every instrument used in the laboratory should meet (___________) and have instructions regarding its (_____). safety written

● Eye wash station, safety (______&______) should be standard facilities in a laboratory. Fire extinguishers, emergency shower systems, emergency eye washers, first aid, emergency blankets, and hoods must be checked monthly.

● To avoid the unnecessary purchase of chemical materials, a (________) of chemical materials must be prepared. Only a minimum amount of (______) must be kept in the laboratory.

● Chemical material should be stored and safely secured where there is (_________). Combustible chemical material must be stored in a (________). Acids and bases must be separately stored.

● Every chemical compound used in the laboratory should have a (_____________) on file that specifies the nature, toxicity, and safety precautions to be taken when handling the compound.

● All chemical material must be (_______) with the name, characteristics, danger level, and precautionary measures.

● Laboratories must have available (______________) for all individuals: safety devices, goggles, gloves, lab coats, and face-shields.

(Collect and seal) (formalin) (tissue grinder) (soil, drains and waterways) (kitty litter” or a commercial product) (safety devices) (Fire safety procedures) (ends) (hand protection) (documented and investigated) (medical advice)

● The laboratory must have a method for disposal of hazardous wastes. (________) absorbed material into labelled containers for disposal. Tissues that are collected should be stored in (_______) and may be disposed by incineration or by putting them through a "(_______)" attached to a large sink (similar to a large garbage disposal unit). Used chemicals must not be released into (___________). Use an absorbent such as sand, “(___________) to collect spills and contain spread.

● One must always be cautious when handling electrical appliances and must be aware of the location of (_______) (fire extinguisher, emergency shower system). Extinguishers with water, carbon dioxide, dry chemical powder or foam are all suitable depending on other products involved in a fire. (_______) should be posted. There must not be any obstacle in the vicinity of the laboratory door.

● Avoid handling the sharp (____) of instruments. Use forceps or other tools to remove sharp instruments from baskets and autoclaves. Workers should use appropriate (________) when hands are exposed to hazards such as cuts, lacerations or thermal burns.

● Laboratory accidents must be (__________) with incident reports and industrial accident reports. Obtain (________) (first aid officer, doctor, poisons information center, ambulance) immediately if major exposure occurs.

PATHOLOGIST

USE & CARE MICROSCOPE

● Examines slides to diagnose disease

HISTOTECHNOLOGIST

● Uses the microscope for quality control of slide preparation (fixation, staining, sectioning).

(cellular details) (Magnifies) (Resolves) (visible)

PURPOSE OF THE MICROSCOPE

● Enlarge image to visualize (________) too small for the naked eye.

● Accomplishes 3 things:

○ (_______) the object

○ (_______) details

○ Makes details (______)

(one lens) (bright field microscope) (condenser, objective and ocular.) (Bright field)

COMPOUND / OPTICAL MICROSCOPE

● Is a microscope with more than (______) and its own light source.

● A compound light microscope is also considered a (_____________)

● Consists of the lens system (_____, _____, ____)

● (________): Specimen appears dark on a bright background.

(Monocular) (Binocular) (Trinocular)

	1 eyepiece, cheap & lightweight
	2 eyepieces, common & comfortable
	3rd eyepiece for camera attachment, more expensive

MAGNIFICATION

● The process that increases the size of the structure under examination. It is achieved by the use of the microscope's lens system.

TOTAL MAGNIFICATION

● Is the product of magnifying power of the

objective and eyepiece, with a normal tube

length of 160mm.

BRIGHT FIELD MICROSCOPE

● Is generally used in compound microscopes, where light is passed through, or reflected off, a specimen

DARK FIELD MICROSCOPE

● Is a technique used to observe unstained and transparent samples causing them to be clearly visible and appear brightly lit against a dark, almost purely black background.

PHASE CONTRAST MICROSCOPE

● Optical microscopy illumination technique

in which small phase shifts in the light

passing through a transparent specimen

are converted into amplitude or contrast

changes in the image.

● Visualizes transparent & live cells without

staining.

POLARIZED LIGHT MICROSCOPE

● Is a contrast-enhancing technique that improves the quality of the image obtained with birefringent materials when compared to other techniques ● Uses polarizer & analyzer at right angles.

(FLUORESCENCE MICROSCOPE) (fluorochromes) (

● refers to any microscope that uses fluorescence to generate an image.

● many wavelengths of light, ranging from the ultraviolet to the visible can be used to cause samples to fluoresce and allow viewing by eye with the use of specifically sensitive cameras.

● Uses (__________); lower magnification shows larger area, faster examination; used in microbiology (e.g., TB detection).

(ELECTRON MICROSCOPE) (electrons)

● Is a microscope that uses a beam of

accelerated electrons as a source of

illumination

● Uses (_______) instead of light → much

higher resolution

TRANSMISSION ELECTRON MICROSCOPE

● 2D image, very thin slices, shows internal structure

● Max magnification: ~10 million×, resolution <50 nm

SCANNING ELECTRON MICROSCOPE

● 3D image, shows surface structure, electron beam bounces off surface

● High depth of field, good for surface morphology

TRANSMISSION ELECTRON MICROSCOPE (TEM)	SCANNING ELECTRON MICROSCOPE (SEM)
Works on the same principle as a light microscope but uses electrons instead of light and electromagnets instead of glass lenses	Uses electron illumination and scans the surface of the specimen with a fine electron beam
Provides a two-dimensional (2-D) image	Provides a three-dimensional (3-D) image

Requires very thin slices of the specimen	Uses bulk specimens; no thin slicing required
Electron beam passes through the specimen	Electrons bounce off the surface of the specimen
Has very high magnification and resolution	Has high magnification and resolution

Resolution can be better than 50 nm	Resolution is high but mainly for surface details
Maximum magnification up to 10,000,000×	High magnification with excellent depth of field
Used to study internal structure of the specimen	Used to study surface structure of the specimen

Specimen must be extremely thin	Specimen is coated with gold
Image is mainly used for detailed internal analysis	Images are black and white and show external features