Lab Exercise 1 Introduction to the Microbiology Lab

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					                       Introduction to the Microbiology Lab

Welcome to Microbiology at Miramar College! We look forward to a rewarding and
enlightening experience with you all. Because our labs and protocols involve not only living
organisms, but also some that are potential pathogens, you must adhere to strict safety
guidelines and rules, in order to ensure safety for yourself, your colleagues, and your family
and friends at home. The safety rules and guidelines are set forth below:

   1.   Lab coats are encouraged but not required and must be kept in personal boxes.
   2.   Closed-toed shoes are required for protection from spills and accidents.
   3.   Long hair must be tied back during lab.
   4.   Wash hands before you begin work and before you leave. In addition, it is a good
        idea to wear gloves for most lab exercises.
   5.   Wash down lab bench with disinfectant before and after work.
   6.   No extraneous books, folders, purses, or book bags on lab bench top.
   7.   Report all spills and accidents. Be prepared to identify the microbe as the
        instructor requests. (See Hazardous Waste Protocols)
   8.   No cultures in boxes at any time, and no removal of any culture, plate, or test tube
        from the lab.
   9.   Test tubes will be transported in a test tube rack at all times.

These rules will ensure a good working environment for everyone. Additional protocols and
equipment lists are outlined below.

All materials that have come in contact with a microbial organism are considered hazardous
waste and must be treated by the following guidelines.

Hazardous waste disposal (The Morgue)
  - Test tubes: Always place test tubes in the appropriate test tube rack in the morgue.
      There are 2 types of tubes used (13mm and 16mm) and separate size racks are
      provided for each. Completely remove all media labels before placing in the rack.
 - Glass: Glass items such as slides and coverslips need to be placed in the yellow sharps
      container in the morgue.
 - Additional sharps: Other sharp objects, such as pipets and pipet tips, toothpicks, and
      cotton swabs, need to be discarded in the white tub.
 - Non-sharps material: All other contaminated material, including plates, gloves, and
      paper towels are discarded in the large, plastic Bio-Bag in the morgue.
 - Stains: All stains must be collected for disposal and NOT poured down the sink.
      Staining trays can be emptied into the large white carboy labeled “Stains Collection”
      in the morgue. Trays should then be rinsed liberally with water before being stored.
Non-hazardous waste: Material that has not come into contact with a microbial organism
      can be disposed of in the regular trash or broken glass receptacle.

If you spill any living organism on yourself or any other surface, you must treat it as a

         - Inform instructor or lab technician of spill
         - Saturate surface with disinfectant
         - Let sit for 5 minutes
         - Wipe up and dispose of towels in Bio-Bag in the morgue
         - Thoroughly wash hands and/or affected area with large quantity of warm water
           and soap

Chemical stains and reagents are kept in a common area and can be refilled and used by
students on an ongoing basis.

Stain kits
- When stain kits are running low, please be courteous to other students and take a few
    seconds to refill any bottles in the staining kits that may need it.
 - When refilling staining kits, be sure to use the sinks and not the counters. After
    refilling, please wash any spilled stains down the sink with plenty of running water.
  - If refill bottles are empty, please ask your instructor or lab staff so we can fill them or
    get new bottles.

Chemical reagents
 - In the reagents cabinet, each row and individual canister is labeled but be sure to double
   check that you are taking the correct reagent.
 - When you’re finished with each reagent, please promptly return it to the correct cabinet
   and row. If one is empty, give it to the instructor or lab staff so we can replenish t
       the stock.

Please follow the following guidelines with respect to common areas.

Instructor’s Bench
  - Keep all your lab materials (tubes, plates, loops, needles, etc.) off front bench at all
  - If you have questions regarding your lab work, ask the instructor or lab staff to come to
    your seat to help you.
Back Bench
 - Necessary media, reagents, and other lab equipment will be place here at the beginning
       of each lab. This area needs to be kept clean and clear so that everyone can get the
   supplies they need.
 - Take your supplies back to your seat and continue your work there. Do not do any
   inoculations or other lab procedures on the back bench unless otherwise instructed.

Incubator Room
  - Incubators: Each section will have a designated 25° C and 37° C incubator.
  - Incubator racks: In each section’s incubator, there is a designated rack for each lab
       table. Be sure that you are placing your tests into the correct rack inside the
       correct incubator. • Always double-check that you are using the correct incubator.
       (lab section and temperature)
  - Culture media removal: When you are done incubating your tests, promptly move them
       into the 4°C for storage or properly discard of them. Do not let old media pile up!
  - NEVER move another student’s media or change the incubator temperature.

You must properly label each and every culture you work with in the lab.

Test tubes: Using provided paper label, write your name, date, and lab #/ organism and
       secure to the test tube using a rubber band. DO NOT write on test tubes or
glassware with markers or pens.

Petri plates: Using a marker, label the outer edge of the bottom side of the Petri plate
with your name, date, and lab#/organism.

You will be using various equipment and tools in the lab. You need to know what they are
called and to properly use them.
          - Microscopes – compound and stereo light scopes are
                used in our labs
          - Stains and staining pans – used to make specimens
                easier to see
         -Bunsen burner – used to heat transfer tools to sterilize

         - Inoculating loop: used to transfer bacteria from one source
               to another
         - Inoculation needle used to transfer bacteria from a
               solid media to another media

       Media forms:
       - broths – liquid growth media
       - agar – used to solidify broth media
       - slants – solid media cooled in a tube at a slant
       -   agar plate – solid media poured into a plate
       -   Deep – solid media in a tube (not slanted)
       -   Pour – larger tube with agar media for pouring into a Petri plate

       Incubators: used to keep temperature constant for bacterial growth
       Refrigerators: used to store bacteria for extended time
       Autoclave: used to kill bacteria no longer needed

You must properly label each and every culture you work with in the lab.

Test tubes: Using provided paper label, write your name, date, and lab #/ organism and
       secure to the test tube using a rubber band. DO NOT write on test tubes or
glassware with markers or pens.

Petri plates: Using a marker, label the outer edge of the bottom side of the Petri plate
with your name, date, and lab#/organism.

The ABCs (and DEFs…) of microscope care. Every day when finished with the microscope
you must:

A.     Move microscope stage to lowest position
B.     Move 4x objective into place
C.     Wipe immersion oil off lens/scope with lens paper only
D.     Turn off light
F.     Turn light intensity dial all the way down
G.     Wrap cord around base
H.     Return to correct location

       IMPORTANT: Please report any problems. You are responsible for checking the
       status of the microscope at the beginning of lab each day. As you will be sharing
       microscopes, this is VERY important. If you notice a problem, inform your
       instructor immediately and the appropriately guilty student will lose points.

Below is a list of items you will find in your shared, community box, a list of items you need
to keep in your own personal box, and a microscope status list. Beginning next lab session,
you must get these check-off sheets approved by an instructor or tech before leaving class.

Please note: You are sharing the community box, so DO NOT put items from the
community box anywhere except in the community box. You are also sharing
microscopes, so you must properly store and clean the microscope at the end of each
class period. Points will be deducted if items are lost or missing and if the
microscope is not properly stored and clean.

NAME __________________________________

Lab Day
  1     2     3     4               Item
____ ____   ____ ____       Steaming can/beaker
____ ____   ____ ____       Tube rack
____ ____   ____ ____       Striker
____ ____   ____ ____       Staining pin
____ ____   ____ ____       Bunsen burner
____ ____   ____ ____       Gas hose
____ ____   ____ ____       Wash bottle
____ ____   ____ ____       Immersion oil
____ ____   ____ ____       Gram stain kit
____ ____   ____ ____       Acid fast kit
____ ____   ____ ____       Staining tray

____ ____         ____      ____

Lab Day
 1      2           3       4                  Item
____ ____         ____    ____                 Inoculating loop
____ ____         ____    ____                 Inoculating needle
____ ____         ____    ____                 Lens paper
____ ____         ____    ____                 Bibulous paper
____ ____         ____    ____                 Slides
____ ____         ____    ____                 Marker

____ ____         ____    ____
Lab Day
1       2     3      4             Status
____ ____    ____   ____   Light off
____ ____    ____   ____   Light power turned down
____ ____    ____   ____   Cord wrapped
____ ____    ____   ____   Stage in lowest position
____ ____    ____   ____   4x objective in place
____ ____    ____   ____   No slides on stage
____ ____    ____   ____   No oil on lenses

____ ____    ____   ____