Microbiology Lab Safety and Procedures

Syllabus, Lab Safety, and Policies

• Key points of the syllabus include attendance, grades, makeups, exams and quizzes, and DSS accommodations.
• Reference page 1 in the Lab Manual and syllabus for more information.
• No food or drink is allowed in the lab.
• Cell phones, iPads, laptops, and headphones are not allowed out during lab.
• Close-toed shoes are required, and no stiletto heels are allowed.
• Being in lab with open-toed shoes or holey shoes will result in dismissal from the lab
• Garments worn on the bottom half of the body must be half-calf or longer.
• Long hair has to be pulled back out of the face.

General Procedures

• Do not invert or shake capped tubes, as the caps are not tight and spills will occur.
• Always use a green test tube rack, located in the back of the lab.
• Hold test tubes by the glass and not the caps.
• Caps are intentionally loose to allow bacteria access to oxygen.
• Label glassware, petri dishes with the name, date, and lab section.
• Use masking tape for glassware.
• Write on the dish for petri dishes, and not the cover.
• In the event of spills, immediately get your TA and do not attempt to clean it yourself

Supplies and Disposal Instructions

• Common supplies include loops, needles, spark lighters, wax pencils, and student test tube racks.
• Used slides go in the metal disinfectant pan in the middle of the tables.
• Prepared slides should be returned to the slide tray.
• Do not throw slides away.
• Remove masking tape from test tubes.
• Place test tubes in the discard area on a test tube rack.
• Petri dishes must be disposed of in biohazard cans.

Lab Best Practices

• Sign the roll upon entering the class.
• All bags and cell phones must be placed on a shelf.
• Wash your hands.
• Disinfect the workspace before each exercise.
• Obtain gloves and a lab coat, if needed.
• Discard all materials in their proper place.
• Wash your hands after each class.
• Disinfect your workspace after each class.
• Dispose of gloves and lab coats in the biohazard can.

Biosafety Levels (BSL)

• BSL stands for Biological Safety Level.
• The levels are BSL 1, 2, 3, and 4.
• Each level consists of using a combination of laboratory practices and techniques, safety equipment (primary barriers), and laboratory facilities (secondary barriers).

BSL 1

• Practices: Standard and microbiological practices.
• Safety equipment: None required.
• Facilities: Open benchtop sink.

BSL 2

• Practices: Limited access, biohazard warning signs, "Sharps" precautions, and a safety manual for waste decontamination policies.
• Safety equipment: Lab coat and gloves.
• Facilities: Autoclave.

BSL 3

• Practices: Controlled access, decontamination of clothing before laundering.
• Safety equipment: Protective lab clothing and a requirement to enter and leave the lab through clothing changing and shower rooms.
• Facilities: Self-closing, double-door access.

BSL 4

• Practices: Separate building.
• Safety equipment: Full-body, air-supplied, positive pressure personnel suit.
• Facilities: Separate building and decontamination facility.

5 Steps of Handwashing

• Wet hands with clean, running water (warm or cold), turn off the tap, and apply soap.
• Lather hands by rubbing them together, including the backs of your hands, between your fingers, and under your nails.
• Scrub hands for at least 20 seconds.
• Rinse hands well under clean, running water.
• Dry hands using a clean towel or air dry them.

Fomites

• Fomites are inanimate objects that can carry disease-causing microorganisms.
• Microorganisms prefer moist objects that frequently come in contact with food, dirt, and vegetation.

Nutrient Agar Plates

• Inoculate 3 nutrient agar plates.
• Prevent puncturing, gauging, cutting, or digging into the plate.
• Nutrient media: Peptones, meat extract, sodium chloride, and distilled water.
• Agar: An extract from marine red algae.
• Used to solidify media and can withstand high temperatures of 100 °C

Incubation

• Petri plates are typically incubated at 37°C.
• Plates are incubated inverted, with the agar side up and the lid on the bottom.
• This arrangement prevents condensation on the lid from dripping onto the agar and potentially causing contamination or improper dilution of the bacteria.

Handwashing Exercise Part 1

• Obtain a nutrient agar plate and use a wax pencil to divide it into four equal sections, numbering them 1-4.
• In Section 1, touch the agar with your index finger without washing your hands.
• After inoculating Section 1, wash your hands with soap and water, and then use the same index finger to touch Section 2.
• After inoculating Section 2, touch a fomite and use the same index finger to inoculate Section 3.
• After inoculating Section 3, rewash your hands with soap and water and use the same index finger to inoculate Section 4.
• Label the plate with your name, date, lab section, and a handwashing label, then invert the plate (agar side up, lid on the bottom) in the class incubator on the tray.

Handwashing Exercise Part 2

• Go to the back of the tables where the NA plates are located and add a small amount of Glo-Germ.
• The Glo-Germ is bright orange, but it dries clear and will not stain.
• Rub the Glo-Germ all over your hands as if it was lotion.
• Use the black light at your table, turn it on, and shine it toward your hands.
• Areas that fluoresce or turn bright orange indicate areas that are not clean.
• Wash your hands with soap and water after looking at them in the black light.
• Disinfect the black light handle with an alcohol wipe.
• Areas on the hands, like under the nails or between the fingers are areas that are not properly cleaned.

Fomites Materials

• 2 NA plates
• 2 sterile cotton swabs
• 1 tube saline
• Wax pencil
• Label the plates with the name, date, lab section, and fomite used.

Review of microbiology lab syllabus, safety rules, and general procedures. Key points include attendance, grades, lab attire, and proper handling of materials. Learn about labeling glassware and using disposal containers.