250 LE1 Study Guide 251 (1).docx

### General Lab and Safety Procedures **What things do we do in the lab when we enter and leave?** Upon arrival- disinfect your bench top with lab disinfectant get out required materials (lab manual and pencil) wash your hands with soap and water (20 seconds) Upon exiting- All lab materials r...

### General Lab and Safety Procedures **What things do we do in the lab when we enter and leave?** Upon arrival- disinfect your bench top with lab disinfectant get out required materials (lab manual and pencil) wash your hands with soap and water (20 seconds) Upon exiting- All lab materials returned to or as directed by instructor Disenfect bench top, dry with paper towels Push in chair under bench top Wash hands with soap andwater (20 secs) **What safety rules do we follow?** Wear gloves and goggles at all times when using chemicals Hair should be tied up No food or drink No long nails No shoes that are open Be prepared for EACH LAB **Where do different types of trash go?** Contaminated throw away items must go in biohazard container Non-contaminated items go in regular trash bin Contaminated glassware goes in sterilization area **What is the purpose of these things?** The purpose is to demonstrate safe handling of microorganisms while working in lab. **Aseptic Culturing Techniques** Purpose of aseptic techniques **-** prevent environmental microbes from contaminating working cultures and to prevent them from contaminating the environment or ourselves agar: jelly made from seaweed, turns solid at room temperature broth: growth media "soup", used to grow bacteria to study or produce more for experiments converted to Petri plate inoculate: microbes from one culture to another inoculating loop: used to spread bacteria against surfaces or between liquid media inoculating needle: stabbing into solid media **Specific steps when making inoculations:** Broth to Broth: Bunsen burner, sterilize loop, cool, pick culture tube with one hand, pass through burner 3 times, insert loop, withdraw bead of culture, flame tube again Broth to agar slant: Sterilize loop, culture tube in one hand, pass through burner 3 times, insert loop, pass again thru tube and sterilize\ Broth to agar deep: sterilize needle,culture tube through burner, insert needle, and withdraw, Labeling and incubation of media, especially agar plates. Use tape **Identify purpose of a streak plate**, the purpose is to spread a tiny amoun tof culture in Petri plate to have single bacterial cells isolate. **what a pure culture is**, one type of microbe, **how you get a pure culture**. single cell -\> single cell , growth, isolated colony **Introduction to the Microscope** Identify all parts of the microscope on a picture. **Identify functions of the different parts.** **Calculate total magnification.** wide span 4x -\> 40x low power 10-\> 100x high dry 40x-\> 400x oil inmersion 100x -\> 1000x **functions of a microscope** magnification- it enlarges tiny objects (cell or bacteria) see them clearly resolution- see the details, like differents parts of cell parfocalism- microscope stay in focus when changing between different magnification lenses **How to focus and find specimens in the microscope** use lowest magnification lense (4x or 10x) place slide on stage turn coarse focus to bring specimen into view adjust stage to center specimen and have good lighting fine focus to make things clear switch lens if needed and use fine focus again **Cleaning and storage of the microscope** lower stage all way rotate lenses to 4X objective use lens cleanser and q tip to clean all objective and ocular lenses use lens cleaner and Kimwipe to remove oil from stage and knobs turn off light and unplug n wrap around scope instructor check **Staining Labs** **Identify the purpose of each staining procedure we did (name of stain anc chemical used)** simple stain: basic dyes to see shape and arrangement of bacteria negative stain- color background of slide while cells are clear, see shape and structure capsule stain: helps to identify bacteria with capsules gram stain: used to differentiate bacteria (gram positive-purple, gram negative - Pink) acid fast stain- identify bacteria that has a waxy cell wall (red) **Contrast basic and acidic stains.** Basic - color cells directly Acid- repel and color background, leaving cells clear **For differential stains, identify the 4 steps and identify the chemicals (or heat) used for EACH step of EACH of the differential stains. (got that)** gram stain- crystal violet, iodine, ethanol, safranin, blot dry\ negative stain- nigrosine acid fast- carbon fuchsin, heat, iodine, acid alcohol wash, methylene blue endospore- malachite green, heat, rinse water, safranin basic stain- crystal violet, water capsule stain- crystal violet, copper sulfate **Be able to identify each stain from pictures through the microscope. (HINT: think about each color or combination of colors you will see from each staining procedure, think about if the cells or the background is stained. Also use context clues from the questions on the exam to help you narrow down to the correct procedure).** **Be able to identify the results of staining procedure from pictures through the microscope (Ex. Gram-positive, acid-fast, endospore-former).** **Be able to identify shape and arrangement through pictures and know the terminology related to shape and arrangement.** ### Cocci (Spherical) - - - - - - ### Bacilli (Rod-shaped) - - - - - - - -