Lab 1

• Ways to adjust light in a bright field compound binocular microscope

  • Rheostat
  • Condenser diaphragm
  • Light Source diaphragm
• Using the microscope

  1. Start on RED lens (course then fine)

  2. After focusing, use fine adjustment only

  3. Once in oil, do not go back to Blue

  4. When done with oil, move to RED lens, then bring stage down and clean
• Oil allows the objective to collect more light to form an image due to its refractive index
• Electromagnetic Spectrum: Energy/light from the sun
• Human blood smear (wright motility methods: wet mount / hanging)
• Eukaryotes are bigger and absorb more light
• Flagella in Eukaryotes move back & forth in a smooth whiplike motion
• Cilia and Pseudopods are present in Eukaryotes
• Flagella in Prokaryotes rotate clockwise or counter-clockwise in an erratic motion
• Brownian motion: vibrating false movements due to H2O molecules
• Live organisms are difficult to observe
• Difference between Eukaryotic and Prokaryotic movement
• Simple stain increases refractive index and helps identify morphology
• Principles of stains

  • Simple, Basic, Differential, Structural
• Basic Stain

  • Chromophore has a positive charge
• Steps for prepping a smear (Agar)
• Fixing a smear
• Stains are salts
• Basic Stain CHROMOPHORE

  • Has a positive charge and is a large molecule that gives color
• Prepping a smear

  Sterilize loop, Transfer loop of water to slide, Heat fix, Pass stain
• Heat fixing


  Kills bacteria, Adheres to slide, "Opens" bacteria wall for stain to enter
• Gram Stain

  Differential Stain based on properties of cell wall, Gram positive and Gram negative differentiation
• Gram Stain Procedure


  Includes steps to know how stain gets in and how each step works
• Simple Stain


  To see shape, Initial Stain with Crystal Violet, Mordant, Iodine, Decolorizer, Counter stain with Safranin
• Gram Positive bacteria
• Gram Negative bacteria
• Gram Stain is a way to identify the type of infectious bacteria; Treatment differs based on the outcome
• Aseptic Technique


  Properties of Agar, Media types like Broth and Agar Plate, Isolation for purity
• Aseptic Technique is performed without contamination
• Fermentation


  Little to no oxygen, converts NADH into NAD+ to recycle and breakdown glucose, purpose is to keep glycolysis cycle running with no energy produced, removes CO2 and produces NAD+ to breakdown more glucose
• Pathogens such as viruses, mold, parasites are missing
• 99% of organisms are not grown in the lab
• CFU's stand for Colony Forming Units
• TNTC stands for Too Numerous to Count
• RODAC stands for Replicate Organism Detection and Counting
• RODAC swabs need moisture
• Viable organisms are counted
• Anaerobic bacteria like C-difficile are mentioned
• Glycolysis is a process where glucose is converted to pyruvate
• Oxygen Requirements: Obligate aerobe needs O2, obligate anaerobe can't have O2, facultative anaerobe wants O2 but can survive without it due to metabolism