EXPERIMENT SUPPLIES

BROWNIAN MOTION OBSERVATIONS
1. Microscope 250X or more
2. Microscope slides and coverslips
3. India ink: must be fresh. Black India ink consists of a suspension of carbon particles.
DIFFUSION
1. To make starch-agar plates
  1. Ingredients
    1. Agar powder (not "nutrient agar" but just plain agar with no other adjectives!)
    2. Sodium Azide (NaN₃) a powerful biocide. (Irreversibly stops cytochromes, hemoglobin and RNA-polymerase)
    3. One petri plate per group (need not be sterile)
    4. Hot Plate
    5. Burner to burst bubbles on surface of freshly poured agar
    6. 10 liters of 0.05% soluble starch solution (most is for the osmosis expt coming next) (this is enough for about 10 groups)
      1. In a liter flask, add 250 ml of COLD water.
      2. Add 5 gm of soluble starch powder; form a smooth, non-lumpy slurry
      3. With constant swirling, bring the slurry to a boil (the appearance should go from white turbidity to clear and somewhat viscous)
      4. Dump the hot solution into 10 liters of cold water and stir to mix.
  2. Preparation (makes 10 plates)
    1. To a liter flask add in this order:
    2. 50 ml cool tap water
    3. 2 gm pure "plain" agar; swirl to make a homogeneous slurry (no lumps)
    4. 250 ml of the above 0.05% starch solution
    5. about 10 granules of sodium azide (will prevent mold growth) Do not lick fingers unless you would like to die as if being strangled or drowned!
    6. With constant swirling, heat to near boiling. The granules of agar will disappear at almost boiling.
    7. Distribute this among 10 petri plates (they don't need to be sterile)
    8. Waft a flame over the surfaces of the molten agar to burst any bubbles. (Do not melt the rims of plastic petri plates - or you will be sorry!)
    9. Do not replace the covers; instead allow the plates to cool and solidify open. Keeping them open while solidifying prevents condensation from forming.
    10. Once solid, replace the covers and distribute to the students.
2. Filter paper and a hole puncher
3. Pencils
4. Small bottle of RED food coloring
5. Tweezers
6. Iodine solution (about 500 ml)
  Add medicinal tincture of iodine to 500 ml of water until the liquid is yellow-orange like this ••• And then guesstimate: add again that amount of tincture of iodine.
7. Put the solution in a cheap, hand-pumped plastic sprayer, which can be purchased at hardware and gardening supply stores.
8. Marking pens
9. Alpha-AMYLASE - I: Since most school systems no longer allow the use of human saliva in lab exercises, a similarly highly active substitute has been sought. There are likely to be many other sources, but the first one found by this author to have very high amylase was a water extract of ginger root. (Potato, sweet potato were of much lower activity. Ginger was higher than even saliva.) Help hint for the nose: Banana will substitute for ginger root as a positive contol BUT only on the starched paper-towel blots.!
  1. Paper towel blot test for Amylase activity:
    1. Spray a Bounty paper towel with spray starch and allow to dry for 20 to 30 minutes. (If you have a reserve of previously made starched towels, test a little piece of one for a positive iodine stain.)
    2. Draw a two inch diameter circle on the starched towel.
    3. Place about a bit of raw ginger root (about half the size of your little fingernail) in the center of the circle.
    4. Smash the ginger with a light bang of a hammer, or crush it using the flat head of a very large nail. DO NOT use items of glass or plastic as crushers: a hand impaled upon a broken one is a mess to clean up!
    5. Just outside of the circle place a drop of your own saliva as a control
    6. Wait five minutes for the enzymes to digest the starch.
    7. Spray the starched towel with the iodine solution mentioned in step 11, above.
    8. Amylase activity is indicated by a white clearing in a blue-black background.
OSMOSIS and DIALYSIS
1. 1 liter graduated cylinder
2. 18 inches of dialysis tubing (ribbon that is 1.75 inches wide)
3. 24 inches of common string
4. Beaker of boiling water (one beaker is enough for whole class)
  All the lengths of dialysis tubing are boiled for a short time to help in opening them.
5. 800 ml of 22% sucrose solution (use table sugar)
6. starch solution: the great bulk of left overs from the diffusion exercise, above.
7. aquarium bubbler that gives large bubbles - a pipette works fine
8. aquarium air pump. You might need to use some "Y" joints from the pumps to supply air to all the graduate cylinders being used. Gang valves work well for this as they have built in valves to adjust the air flow to each of the cylinders.
9. yellow food color
10. linear x linear graph paper
11. Alpha-Amylase - II: Make some ginger root extract (Must be used for the osmosis experiment as this filters much better than does a water extract of mushy bananas.)
  1. Take a piece of ginger root that is about twice the volume of your thumb
  2. Add it to a blender with about 20 ml of tap water and an ice cube (isolated enzymes like it cold!)
  3. Blend until no ice chunks exist.
  4. Filter the mash and liquid through a paper towel in a funnel, that is, in turn, set in a flask. (Don't be surprised if your room begins to smell as if you had just baked ginger snaps!)
  5. Voila! Ginger root extract for use by the class in both this diffusion lab, and in the next procedure - osmosis and dialysis.
  6. Test a drop of this on a bit of starched paper towel; wait 5 minutes; spray with iodine.
  7. If strong activity (white spot on a dark background), the extract is ready for use in the dialysis experiment.
OSMOTIC PRESSURE
1. 12 inches of dialysis tubing (ribbon that is 1.75 inches wide); bring to boil in water for easier opening.
2. 10 gm polyethylene glycol ("PEG")
3. ruler in millimeters
4. Bucket
TRANSPIRATION
1. 12 inches of dialysis tubing (ribbon that is 1.75 inches wide); bring to boil in water for easier opening.
2. As this would be an extra-credit project for willing students, they should improvise.