Introduction to theMicrobiology Lab

Exercises 1-2, 2-1, 3-1, and 3-3

LABORATORY SAFETY

Bio Safety Levels

Student Conduct

• Do NOT smoke, eat, drink, or bring food or drinks into the laboratory room

• Do NOT apply cosmetics or handle contact lenses• Wash your hands thoroughly with soap and water

after handling microbes AND before leaving the laboratory room each day

• Do NOT remove organisms or chemicals from the laboratory

• Work carefully and methodically• COME PREPARED TO LAB FOR THAT DAY’S WORK

Basic Laboratory Safety

• Wear your lab coat and gloves while working in the laboratory– Remove them before leaving– Discard gloves appropriately (not in standard trash)

• NO OPEN-TOED SHOES OR SANDALS• Turn off Bunsen burners when not in use• Secure loose hair and clothing when working with an

open flame• Never pipette by mouth

Safety Equipment

• Know the location of the First Aid Kit• Know the location and proper way to use the

Fire Blanket, Shower, and Fire Extinguisher• Know the location and proper way to use the

Eye Wash Basin

• NOTIFY YOUR INSTRUCTOR OF ANY EVENT THAT NECESSITATES THE USE OF THESE

Reduce Risk of Contamination

• Clean bench with disinfectant before and after lab– 70% alcohol can also be used as a disinfectant– Allow to air dry rather than wipe up

• Never lay culture tubes on bench top• To clean spills: cover with paper towels, saturate with

disinfectant and leave for 20 minutes– Discard in appropriate container

• Place non-essential books, etc. out of the way of your experiments

Disposing of Contaminated Materials

• Broken glassware, needles, wooden cotton swabs, etc. MUST be placed in the “sharps” container

• Culture tubes are to have any labels removed and placed upright in racks on the “kill” cart

• Petri dishes can be discarded into solid waste on the “kill” cart WITHOUT removing the labels

• Gloves are to be discarded in the appropriate biohazard containers

Care of Microscopes

• Microscopes are located at each lab station• Following their use, microscopes:

1. Should have all oil removed from the lenses using LENS PAPER and cleaning solution• YOUR INSTRUCTOR WILL INSPECT THEM BEFORE YOU ARE

PERMITTED TO LEAVE

2. Scanning (4X) lens should be in place3. Stage should be at lowest setting4. Rheostat should be adjusted to its lowest setting5. Cord should be wrapped carefully around the base and

not “flopping” around

Care of the Lab

• Place seats back under benches before you leave

• Wipe down the sink-tops with paper towel if you are among the last students in the lab– Remove any paper towels left behind by less

conscientious students

THE LAB SHOULD LOOK AS NICE AFTER YOU ARE DONE AS IT DID BEFORE YOU STARTED

SCAVENGER HUNT

Essential Equipment• Security Phone• Disinfectant• Antiseptic Hand Solution• Fire Extinguisher• Emergency Shower

– With Eye Wash

• Fire Blanket• MSDS Folder• Biohazard Waste Container• Biohazard Sharps Container• Broken Glass Container• “Kill” Cart

• Incubators• Refrigerated Cold Room• Biological Safety Cabinet• Vortex Mixers• Microscopes• First Aid Kit• Chemical Spill Kit• Microbiology Lab Tools• Autoclave• Fire Alarm• Escape Routes• Gas Shut-off

New Experiments

EXERCISE 1-2GLO-Germ Hand Wash Sanitation System

Glo Germ Hand Wash System

• Effective hand washing minimizes transmission of pathogens by health-care workers and food handlers

• How Effective Is Your Hand Washing Technique?

• Follow the directions outlined in the lab manual…– Work in groups of 2 with one person applying the Glo

Germ gel and the other person holding the UV lamp– Do NOT hold UV lamp if you also have Glo Germ applied– TURN UV LAMP OFF WHEN EXERCISE IS COMPLETE

EXERCISE 2-1The Ubiquity of Microorganisms

Ubiquity of Microorganisms

• Many microbes can easily be isolated from a variety of environments – Soil, water, plants, and animals

• Just how ubiquitous are microbes?

• Follow the directions outlined in the lab manual…– You will work in groups of 4 – Incubate plates 1, 2, and 8 at 25oC.– Incubate plates 3, 4, 5, 6, and 7 at 37oC.

EXERCISE 3-1Introduction to the Light Microscope

Principles of Light Microscopy

• Illumination– Stains add contrast to specimens

• Magnification– Compound microscope has paired ocular lenses and an

objective lens

• Resolution (Resolving Power)– Clarity of an object viewed under the microscope– Distance that two points must be apart to appear as two

distinct points– Resolution of typical light microscope is 0.2μm

Parts of the Light Microscope

Involved in Illumination• Base• On/Off Switch• Lamp (Bulb)• Light Intensity Knob• Sub-stage Condenser• Iris Diaphragm

Involved in Magnification/Focus

• Stage• Stage Mechanisms• Course/Fine Adjustment• Revolving Nosepiece• Objective Lenses

– 4X, 10X, 40X– 100X (Oil Immersion)

• Ocular Lenses

EXERCISE 3-3Microscopic Examination of Eukaryotic Microbes

Examination of Eukaryotic Microbes

• Follow the directions outlined in the lab manual…– You will work individually with your own microscope, but feel

free to divide the specimens up equally for purposes of seeing everything:

Oscillatoria sp. AmoebaSaccharomyces cerevisiae Giardia lambliaAspergillus sp. Spyrogyra sp.Rhizopus sp. DiatomsEuglena sp. Volvox sp.Paramecium caudatum Dog tapewormHuman louse Plasmodium

sp. (in RBCs)