Modeling the Mitochondria (Part 2/2)

Using a student laboratory kit from Flinn Scientific, students will observe live, functioning mitochondria.  This lab uses a specialized stain that allows students to vividly see mitochondria breaking down sugar. Color change in cells is easily viewed as the dye-sugar mixture is oxidized by mitochondria. Next, students revise the model that they completed yesterday to have a more complete understanding of the relationship between photosynthesis and cellular respiration. This is the second day of a two day lesson.  Here is an overview of what students will learn today.

Using the Determining Specimen Size powerpoint, students should review how to calculate the size of the field of view at high power.  Students should include the calculations in their lab notebooks.

Possible student answers:

• On low power, approximately 10 organisms will fit across the field of view.  
• The letter e is one millimeter long.  
• One should also remember that specimens are inverted and reversed.  
• The conversion factor between 100X and 400X is 0.25.  
• The size of the cell is approximately 375 um.  
• The size of the nucleus is approximately 38 um.
• Field of view diameters

4X: 100X/4X=25       25*1500=37,500 um

20X:  100X/20X=5    5*1500=7500 um

1000X:  100X/1000X=0.1 0.1*1500=150 um 

(Note:  I use this graphic from Sea Urchin Embryology.  I print them off on acetate sheets that are used for the older style overhead projectors. I provide all of my students these rulers.)

Students will use a kit from Flinn Scientific to observe live mitochondria from celery stalks.  (Note: The kit can be found at this link and has supplies for a class of 30.)

Materials needed for this lab: 

• Fresh celery
• Sucrose solution, 0.1M
• Janus Green B stain, 0.01%
• Sodium bisulfite solution, 0.5M
• Microscope slides
• Coverslips
• Microtome or Razor blades (single edge)
• Kimwipe or paper towel
• Forceps

Instructions:

Students should obtain a clean microscope slide.  They should add several drops of 0.1 M sucrose solution to the microscope slide. Next, they should remove the veins from the celery stalk.  Using the microtome, they should carefully cut a very thin horizontal section from fleshy portion of the celery.  With forceps, they should dip the celery piece into sucrose solution.  Then they should place it on the microscope slide making sure not to touch the celery with your hands.  Students should cover the specimen with a coverslip. Next they should place the slide onto the microscope stage with the 10X objective in position.  Students should sketch of what they see. They should estimate the size of the specimen. Remind students to  show their work. Next, they change the objective to 40X and focus the slide making sure to only use the fine focus.  They should examine the specimen very closely for cytoplasmic streaming. Students should identify any structures you can size.   Students should make another sketch of what they see. They should estimate the size of the specimen. Remind students to show their work at every step.

They should locate a single cell near the edge of the coverslip and focus on it. Next, they should hold a small piece of Kimwipe on the left side of the coverslip on the microscope slide. Students should place 3 drops of Janus Green B stain on the microscope slide to the right of the coverslip. They should  allow the Kimwipe to draw the stain through the celery specimen. 

Students should make a sketch of what you see. They should continue to make observations every minute for 5 minutes and record other changes on the worksheet or their lab notebook. Finally, students should add two drops sodium bisulfite solution and wick it through the slide.  They should take observations for 5 more minutes.

Students should dispose of microscope slides in the broken glassware bucket and the celery pieces in the chemical disposal bucket.