Materials Included In Kit

Additional Materials Required

Prelab Preparation

1. Prepare a 0.01% methylene blue solution. Dilute 1 mL of the 1% methylene blue solution to 100 mL with deionized water. Distribute several milliliters to each group in 50-mL beakers or dropper bottles.
2. Label the bulb of each graduated pipet that will be used to retrieve protists from the culture jars.
3. Dispense 1 mL of methyl cellulose solution into 50-mL beakers or dropper bottles.

Safety Precautions

Methylene blue will stain skin and clothing. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Remind students to wash their hands thoroughly with soap and water before leaving the laboratory. Please review current Safety Data Sheets for additional safety, handling and disposal information.

Disposal

Please consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding. Sterilize protist cultures by adding an equal amount of a 10% bleach solution. Allow to rest for 24 hours then flush down the drain with copious amounts of water. Methyl cellulose solution may be flushed down the drain with large amounts of water. Methylene blue solution may be disposed of according to Flinn Suggested Disposal Method #10, neutralizing with an acid.

Lab Hints

• The prelaboratory assignment may be completed before coming to lab, and the Post-Lab Questions may be completed the day after the lab.
• The lab can be conducted by viewing preserved specimen and live specimen simultaneously engaging 30 students working in groups of 2. Performing the lab in this manner can reasonably be completed in two 50-minute class periods.
• Another option is to complete the preserved specimen portion utilizing a digital microscope camera to identify key structures as a whole class activity. This procedure can be completed within a 50-minute class period. Students, working in groups of two, would view live specimen during the second 50-minute class period.
• If possible, allow the protists to become hungry and then have the students add a drop of food to their depression slides so that they can observe the organisms eating. Paramecia will eat yeast cells. Amoeba will eat algae.
• Some protists, especially the ciliated form, move very quickly which can make locating them a challenge. Included in the kit is methyl cellulose solution designed to slow down protists. Flinn offers a freebie publication with detailed explanations regarding this product. Contact Flinn and request a free PDF publication of Protozoa Exceed the Speed Limit!.
• Many other prepared microscope slides of protists are available from Flinn Scientific Inc. Recommended extensions include—Physarum (slime mold), Catalog No. ML1053; Didinium nasutum, Catalog No. ML1041; Plasmodium, Catalog No. ML1042; Chlamydomonas, Catalog No. ML1045; Mixed green algae, Catalog No. ML1055; Fucus, Catalog No. ML1071; Polysiphonia, Catalog No. ML1072.
• Many other live cultures of protists are available from Flinn Scientific, Inc. Please reference the current Flinn Scientific Catalog/Reference Manual for our current selection.

Further Extensions

• Extend the activity by having student groups research each protist to be observed in the activity.
• Extend the activity by having student groups develop a research project involving the protists. For example, study the phototaxic behavior of euglena or the rate at which the contractile vacuole in paramecium contracts when in slightly salty water versus freshwater.

Answers to Prelab Questions

1. What are the different methods of locomotion used by protists?

   Protists move using cilia, flagella or pseudopods.

2. What is the evolutionary advantage of mixotrophs?

   Mixotrophs are able to continue to grow and reproduce even when one type of food acquisition is not available due to the environmental conditions at that particular time.

Sample Data

Observations

Answers to Questions

1. What features were observed on the prepared slides that were difficult to observe in the living specimens? Why was this so?

   Living specimens are constantly moving and the internal structures are not stained, making internal observations difficult in a living specimen. For example, organelles are visible as dark dots on the prepared slides.

2. Describing the shape of amoeboid species is very difficult. Based upon your experience with amoeba in this activity, explain why this is a problem.

   Amoeba do not have a definite shape because the pseudopods are constantly changing.

3. Unlike the higher plants, plant-like protists do not have roots, stems or leaves. Explain why they do not require these structures.

   Plant-like protists conduct all of the functions of life within the confines of a single cell. Roots, stems, and leaves are all specialized to ensure every cell of a multicellular plant receives all of the nutrients, water, and food it needs to survive.

4. Why do protists live in moist or aqueous environments?

   An aqueous environment provides a source of water for the cytoplasm within the protist cell. Water also provides a media in which nutrients and food, if appropriate, is dispersed.

References

Campbell, N. and Reece, J. 2005. Protists in Biology, 7th ed. 549–567. San Francisco: Pearson Education.

Introduction

Protist is a convenient grouping formed from several kingdoms of life. Members are placed into the protist group because they are eukaryotes that do not belong to the kingdom Animalia, Plantae or Fungi. Until recently these organisms were classified into the kingdom Protista. Recent advances in classification, such as DNA sequencing, have lead to the abandoning of Protista in favor of multiple kingdoms. Protists include the fungi-like slime molds, the plant-like algae and the animal-like protozoa.

Concepts

• Protist
• Locomotion
• Autotroph
• heterotroph
• mixotroph

Background

The diversity of protists is exemplified in the variety of methods the different species use to acquire food. Protists include autotrophs, heterotrophs, decomposers, parasites, and even mixotrophs. Mixotrophs can be either an autotroph or a heterotroph depending upon environmental conditions. Protists may be motile or immobile with some species motile during one stage of life and immobile in another stage. Protists may be unicellular, colonial or multicellular. Some organisms reproduce asexually or sexually and many organisms have both asexual and sexual reproduction depending upon environmental conditions. Some protists are anaerobic but most are aerobic. Protists live in water, in moist places on land, as symbionts, or as internal parasites.

Euglenozoans comprise a diverse group of protists. All Euglenozoans have flagella containing a crystalline structure whose purpose is unknown at this time. The often-studied Euglenas are part of this group. Euglenas have an eyespot and a light detector. These organelles function as sensors so that the euglenas move toward light of appropriate intensity but away from too much light. Most species of euglena are primarily autotrophic but they can engulf prey by way of phagocytosis and thus become heterotrophic if sunlight is not available, thus euglenas are an example of a mixotroph.

The kingdom Alveolata has membrane-bound sacs just under the plasma membrane. Dinoflagellates, apicomplexans, and ciliates are all part of the Alveolata group. Dinoflagellates are an important part of marine and freshwater plankton. Dinoflagellates have two flagella which make them spin as they move through the water. Dinoflagellates are another example of a mixotroph. A coastal phenomenon called “red tide” occurs when there is a sudden increase in the dinoflagellate population. The apicomplexan species are animal parasites. The most well-known apicomplexan, Plasmodium, causes malaria in humans. Plasmodium has a complex life cycle that includes life stages within mosquitoes that bite and then infect the human host. Ciliates are one of the most common types of protist. Ciliates have cilia protruding from the cell membrane plus two types of nuclei—a large macronucleus and a tiny micronucleus. The macronucleus contains multiple copies of the ciliate’s genome, specifically the genes responsible for feeding, waste removal, and water balance functions. The micronucleus is exchanged during sexual reproduction. Paramecia and Stentor are two types of ciliates that are readily visible in pond water.

The kingdom Stramenopila derives its name from the Latin stramen meaning straw and pilos meaning hair. Stramenopiles have flagella with numerous fine, hair-like projections. Included within Stramenopila are the water molds, diatoms, golden algae, and brown algae. Diatoms are unicellular algae that have a cell wall composed of silica. The complex silica wall contains tiny holes through which the diatom interacts with the environment. These silica “shells” do not decompose when the diatom dies. We use these shells in filters for pools and other applications.

Cercozoa and radiolarians both use thread-like pseudopods to move; Amoebozoa move using lobe-shaped pseudopods. Pseudopods are extensions created as bulges from the cell surface. An amoeba moves by cytoplasmic streaming. Under a light microscope the organelles can be seen streaming into the pseudopod. Slime molds are now grouped as part of Amoebozoa because of their method of movement. There are two kinds of slime molds—cellular slime molds and plasmodial slime molds. Cellular slime molds live a majority of their lives separately as single-cell amoeboid protists. Plasmodial slime molds also begin life as single cells but these cells quickly merge together to form a large, multinucleated plasmodial cell. When slime molds sense a detrimental environmental change, they form fruiting bodies similar to those found in fungi. These fruiting bodies release a new generation of amoeboid cells. Slime molds are not static creatures; they creep along the ground or up trees “hunting” for bacteria, other protists or decomposing organic material to ingest. Some slime molds can move up to 2 cm per hour. Because of their large size and unique characteristics slime molds are used by scientists to study details of the cell cycle.

The final two protist groups are red algae and green algae. Algae are said to be plant-like due to their ability to photosynthesize sunlight. The accessory pigments of the red algaes absorb blue and green light. Blue and green wavelengths of light penetrate deeper into the ocean than red and yellow wavelengths. This allows red algae species to survive at deeper depths. Green algae likely gave rise to the land plants. Species of green algae range from the mobile, unicellular Chlamydomonas to the rolling, colonial Volvox to the immobile, multicellular Ulva or sea lettuce.

Experiment Overview

The purpose of this laboratory is to explore the many diverse characteristics of the protists. Both live and preserved specimens will be observed during this activity.

Materials

Prelab Questions

1. What are the different methods of locomotion used by protists?
2. What is the evolutionary advantage of mixotrophs?

Safety Precautions

Methylene blue will stain skin and clothing. Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Wash hands thoroughly with soap and water before leaving the laboratory. Please follow all laboratory safety guidelines.

Procedure

1. Observe the prepared microscope slides with a compound microscope. Begin each observation using the low power objective switching to higher power to observe the internal and external features of each protist. Sketch each type of protist on the Characteristics of Protists Worksheet. Label the following items in each sketch—nucleus, cytoplasm, and method of movement, if appropriate (pseudopods, flagella, cilia). Record the magnification used.
2. Observe and sketch each type of living specimen using a compound microscope. Place one drop of the protist culture in the well section of a clean depression slide. Note: Be careful to use the correct graduated pipet for each culture. Some of the protists are prey for others. Protists can be difficult to locate in a culture jar. It may take several attempts to locate specimens. The use of a stereoscope will make this easier. See specific notes below for assistance in locating each type of protist.
   1. Amoeba are found in and around the bits of debris in the culture. They often encyst (form a ball shape) and become immobile during shipping. Amoebas move very slowly; do not use methyl cellulose. Locate a colorless bit of debris and observe it closely with minimal light transmitting through the microscope. Many times these bits of debris are actually amoeba which will begin to move away from the light.
   2. Euglena are attracted to light and are usually green in color. They can be found swimming throughout the culture and are visible as green specks using a stereomicroscope. Use methyl cellulose solution to slow the euglena down.
   3. Paramecia are light tan to colorless. Paramecia do not like light. The light transmitting through the preparation should be kept to a minimum. Paramecia can be found swimming throughout the culture. Use methyl cellulose solution to slow the paramecia down. Paramecia cultures are shipped with their prey which are also protozoa. These prey called Chlamydomonas are round with two flagella. Paramecia have a large oral groove into which food is guided. Once enough food is present, a vacuole forms so that the food may be digested. Paramecia also have large contractile vesicles which regulate the amount of water located within the organism. With careful observation the contractile vesicle can be seen filling and emptying. When paramecia die they release trichocysts—long thread-like projections originating from specialized cells just below their cell membrane.
   4. Volvox are large, green colonial protists. They are visible without using a stereomicroscope as green balls throughout the culture. Do not use methyl cellulose solution.
   5. Diatoms have a green or golden color. They can be found swimming throughout the culture. Do not use methyl cellulose solution.
3. Carefully observe the movement of each protist.
4. Add a drop of methyl cellulose solution, if needed to Euglena and Paramecium only, in order to make careful observations of the internal organelles and the external cilia or flagella. Use a clean toothpick to stir the methyl cellulose into the drop of protist culture. Label the following items on each sketch—nucleus or nuclei, cytoplasm and method of movement, if appropriate (pseudopods, flagella, cilia).
5. Some organelles become more visible when stained. Carefully add one drop of methylene blue stain to the depression slide. Carefully mix the stain using a clean toothpick. Locate an organism and carefully observe the internal structures. Add any new features to the original sketch using a blue pencil.
6. Rinse the depression slide with water and dry with lens paper between each use.

Student Worksheet PDF

10904_Student1.pdf