Introduction:
      Mitosis  (normal cell division) is an important process in biology because of its roles in asexual reproduction, growth, and development.  As a result, many biology students observe cells in the different stages of mitosis by preparing onion root tip slides.  Students usually enjoy these labs because they get to prepare the slides themselves.  They can see and touch the materials that they are using and go through the process of preparing the slides.
      The process of meiosis is an essential part of sexual reproduction in higher plants and animals. Unfortunately, hands-on meiosis labs are not common.  Activities involving drawings, paper cut-outs or plastic "chromosomes" are used in place of a true hands-on lab involving real plants and animals.  If you have a student or group of students that wish to work beyond the typical dry meiosis labs and try a wet meiosis lab where they make their own slides and look for meiotic divisions, the following lab activities may be useful.

Purpose: These lab activities were designed to:

1. encourage biology teachers to do hands-on biology labs,
2. allow students to identify and collect readily available sources of plant and animal materials,
3. provide an inquiry based lab procedure that advanced biology students can use with practice and patience.

• Biology students
• Grade Level: For High School Students taking an Advanced Biology or Genetics course .  This investigation activity could also be used for an independent project.

Notes to the Teacher: to top

• Requires patient students that have used compound microscopes before.
• Preparation time needed - The plant buds and animal testes need to be collected in the late spring and early summer, when meiosis is most to be occurring.  The buds and testes can be stored in fixative for at least a year.
• All the material from the plants and animals needs to put in fixative (3:1 fixative consisting of 3 parts 100% ethanol and 1 part glacial acetic acid) for several days to bleach out any pigment present.
• Class time needed - In a 45 - 60 minute period, 7 - 10 slides can be made.  The observation time needed to check the slides for meiosis can take another 30 -60 min.
• If petroleum jelly is put around the edges of the cover slips, the slides can be kept for several days.
• Hazards / Precautions - The stain used can stain skin and clothes.  The fixative solution is flammable. Avoid contact with and breathing of any of the chemicals used to kill the insects.

Materials needed for lab:

• Buds from a variety of pollen producing flowering plants (such as beans, spiderworts and lilies)
• Microscope slides and overspills
• A probe and scalpel or razor blade
• Forceps
• A compound microscope capable of at least 400X magnification
• A Bunsen burner or hot plate
• Acetocarmine stain.  Most biological supply companies carry prepared acetocarmine that is ready to use.
• 3:1 fixative consisting of 3 parts 100% ethanol and 1 part glacial acetic acid. (600 ml ethanol and 200 ml glacial acetic acid).  100% methanol can be used in place of the ethanol.
• Testes from grasshoppers, crickets and other insects

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Collection of animals testes for the meiosis lab:
     Collect the insects you plan on using in the late spring and early summer when the male insects are producing sperm.  Grasshoppers are common insects that are easy to catch, have testes that are easy to locate and breed in the late spring while school is still in progress.  House crickets may be a good source of specimens, too.  Consult a zoology or entomology text so you can distinguish between male and female insects.  Put the insects in a killing jar and add a chemical such as dichlorobenzene or ethyl acetate to kill the insect.  Finger nail polish remover or moth balls will work too.  Once the male insects are dead, remove the testes from the insects and store them in the fixative solution  (3:1 ethanol / acetic acid).  Directions for excising and preserving the testes are included in most invertebrate and microtechniques lab books.  I have included a frontal section diagram of the posterior of a male grasshopper in case, an appropriate text can not be located.  The shape of the internal organs will vary between species of grasshoppers.  They can be stored for over a year in fixative.

Directions  for Preparation of Plant and Animal Meiosis Slides:

For Plants:   
     Remove the bud from the fixative and place it in a open flat container (petri plates work well).  Use an eye dropper to add some of the fixative to the bud so the bud doesn't dry out.  The container should be open so the you can use pointed forceps to dissect out the anthers.  Use a dissecting microscope if you need to.  Good lighting and manual dexterity are helpful.  Check a diagram in any good botany book for the location and shape of the anthers in a typical plant bud.  Once the anthers have been isolated, transfer them to a slide and cover the anthers with acetocarmine stain.  Use a probe or razor blade to mash up the anthers in the acetocarmine stain.  Use a fine tipped forceps to remove debris such as anther walls.  Make sure that you have removed the outside of the anthers.  Let the remainder of the anthers set in the stain for several minutes.  Add more acetocarmine stain to the slide if dries out.   Cover the anther remainders on the slide with a cover slip.  Check the heating directions and squash technique that is used for both plant and animal slides below.
 
For Animals:
     Remove the testis from the fixative and place it in slide.  Use forceps or a razor blade to smash and smear the contents of the testis over the central part of the slide.  Remove any large pieces of debris with fine pointed forceps.  Use an eye dropper to cover the material with acetocarmine stain. Let the material set in the stain for at least several minutes.  Add more acetocarmine stain to the slide if dries out.   Cover the testis material on the slide with a cover slip.

Heating to set the stain and the squash technique:
     The next step is heating the slide gently.  If a Bunsen burner or alcohol lamp is used, hold the end of the slide and move it back and forth through the flame.  You have to be careful not to heat the slide so the stain boils or burn your fingers.  Gentle heating will improve the staining process considerably.  Take your time.  If you heat the slide on a hot plate, it has to be set at 80 degrees C.  Allow the slide to set on the hot plate for 3 - 5 minutes.  Make sure the stain on the slide does not boil.  If the material starts to dry out add more stain to the edge of the oversleep.  Capillary action will pull it in.  After heating the slide,  let it cool for a few seconds.  Place the slide between layers of paper towels or napkins.  Press straight down firmly.   You need to be careful not to break the slide.  This squashing process flattens the cell's nuclei and spreads out the chromosomes.  The excess stain that leaks out will absorbed by the towels.

Examination of the Slides:
     The slides are now ready to be examined using a compound microscope. With hard work, you might find meiotic cells.  Remember that you may have to look through quite a few slides to locate cells in different stages of meiosis.  Having diagrams or photos of meiotic cells as reference points is quite useful in the process of searching the slides.  Remember, that you may search quite a few slides before you get lucky.  If you find a slide you want to keep for several days, adding petroleum jelly to the edge of the cover slip will prevent it from drying out.
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 Trouble Shooting Suggestions:

• 1.  Allow the slides to set in the acetocarmine stain for at least 2 - 3 minutes or longer if the cells are not stained so you can see the nuclei.
• 2.  The slides must be heated properly to set the stain.  Do not let them boil or dry out.
• 3.  The specimens must be squashed to flatten the material so nuclei can be seen.
• 4.  This technique requires patience and practice. Making 7 to 10 slides each time increases your chances of locating cells undergoing meiosis.
• 5.  Have backup meiosis slides from a biological supply house so students can see what they are looking for.
• 6. Have the students study diagrams of cells under going both mitosis and meiosis, so they can tell the difference.
• 7. The anthers and testes have to be collected at the right time of the year in your area to ensure that meiosis is occurring.
• 8. Different plants and animals could be used to prepare meiosis slides.  Lilies, spiderworts and beans are good choices for plant slides due to their availability, anther size and large chromosomes.  Grasshoppers and crickets are easily found and make good choices for animal slides.

• This is an older text, but a great resource for preparing slides.