Chapter 1. Mirror Experiment Activity 5.8

Mirror Experiment Activity 5.8

The experiment described below explored the same concepts as the one described in Figure 5.8 in the textbook. Read the description of the experiment and answer the questions below the description to practice interpreting data and understanding experimental design.

Mirror Experiment activities practice skills described in the brief Experiment and Data Analysis Primers, which can be found by clicking on the “Resources” button on the upper right of your LaunchPad homepage. Certain questions in this activity draw on concepts described in the Experimental Design primer. Click on the “Key Terms” buttons to see definitions of terms used in the question, and click on the “Primer Section” button to pull up a relevant section from the primer.

Experiment

Background

During the mid-1900s, biologists debated the nature of the cell plasma membrane. Was it a “fluid” structure, whose components could move freely? Or were the lipids and proteins within the plasma membrane firmly anchored and relatively immobile? The advent of antibody staining techniques provided researchers with a new tool to track the movement of proteins and lipids within cell membranes. Specific proteins or molecules could now be tagged with fluorescent markers and subsequently visualized. This technique could be used to help determine the true nature of the plasma membrane.

Hypothesis

Cells derived from different species express unique surface antigens, or molecules located on the cell surface. These surface molecules associate with lipids or proteins within the plasma membrane. If a mouse cell and a human cell could be successfully fused and cultured, the expression of species-specific surface antigens could be assessed.

Investigators could then evaluate which of two alternative hypotheses is correct. If the cell membrane were a fluid structure, then proteins and lipids from both species would mix within the membrane, forming a “mosaic” structure of interspersed mouse and human surface antigens. If the cell membrane were rigid, then the membrane on one side of the cell would contain only mouse-specific surface antigens, and the membrane on the other side of the cell would contain only human-specific surface antigens.

Experiment

In the 1960s, L. D. Frye and M. Edidin successfully fused mouse and human cells, forming heterokaryotic cells that contained two distinct nuclei - one mouse and one human - and a mixture of mouse and human cytoplasm. These cells were cultured for between 5 and 40 minutes and subsequently stained for mouse and human surface antigens. Fused mouse/human heterokaryotic cells were placed into three different categories based on the distribution of mouse and human antigens within the plasma membrane (Figure 1):

non-homogenous membranes: mouse surface antigens are on one side of the cell and human surface antigens are on the other side.
intermediate membranes: either mouse or human surface antigens are restricted to one side of the cell, while the other antigen type is dispersed on both sides.
homogenous or “mosaic” membranes: both mouse and human cell surface antigens are evenly dispersed on both sides of the cell.

Figure 1

Results

After 40 minutes in culture, most heterokaryotic cells had “mosaic” membranes, containing a homogenous distribution of mouse and human surface antigens. The plasma membrane thus appeared to be a fluid structure, in which proteins and lipids could move.

Source

Frye, L. D., Edidin, M., 1970. “The rapid intermixing of cell surface antigens after formation of mouse-human heterokaryons.” J Cell Sci. 7: 319–35.

Question 1 of 8

Question

During the course of their experiments, Frye and Edidin cultured heterokaryotic mouse/human cells at different temperatures. They observed the results shown in Figure 2, below. What conclusions could they have drawn from this data?

The majority of cells cultured at room temperature (26°C) have non-homogeneous plasma membranes.

The majority of heterokaryotic cells cultured at low temperatures (0°C or 15°C) have intermediate plasma membranes.

The majority of heterokaryotic cells cultured at low temperatures (0°C or 15°C) have non-homogeneous plasma membranes.

The majority of heterokaryotic cells cultured at low temperatures (0°C or 15°C) have mosaic plasma membranes.

Temperature has no effect on the membrane composition of heterokaryotic cells; at any temperature, cells have an equal chance of having non-homogeneous, intermediate, or mosaic plasma membranes.

Figure 2

Correct.

Incorrect.

Question 5 of 8

Question

To assess the expression of mouse and human surface antigens within cell membranes, Frye and Edidin employed antibody staining. During this process, cells are exposed first to a primary antibody that binds a specific target protein or molecule. Cells are then exposed to a fluorescently labeled secondary antibody, which recognizes and binds to the primary antibody. What did Edidin and Frye likely use as controls during their antibody staining experiments?

Cells exposed only to primary antibodies.

Cells exposed only to secondary antibodies.

Cells exposed to neither primary nor secondary antibodies.

Cells exposed to primary and secondary antibodies for molecules known to be present in mouse or human cells.

All of the answer options could be used as controls.

control

Operations or observations that are set up in such a way that the researcher knows in advance what result should be expected if everything in the study is working properly.

Table

Experimental Design

Testing Hypotheses: Controls

Hypotheses can be tested in various ways. One way is through additional observations. There are a large number of endemic species on the Galápagos Islands. We might ask why and hypothesize that it has something to do with the location of the islands relative to the mainland. To test our hypothesis, we might make additional observations. We could count the number of endemic species on many different islands, calculate the size of each of these islands, and measure the distance from the nearest mainland. From these observations, we can understand the conditions that lead to endemic species on islands.

Hypotheses can also be tested through controlled experiments. In a controlled experiment, several different groups are tested simultaneously, keeping as many variables the same among them. In one group, a single variable is changed, allowing the researcher to see if that variable has an effect on the results of the experiment. This is called the test group. In another group, the variable is not changed and no effect is expected. This group is called the negative control. Finally, in a third group, a variable is introduced that has a known effect to be sure that the experiment is working properly. This group is called the positive control.

Controls such as negative and positive control groups are operations or observations that are set up in such a way that the researcher knows in advance what result should be expected if everything in the study is working properly. Controls are performed at the same time and under the same conditions as an experiment to verify the reliability of the components of the experiment, the methods, and analysis.

For example, going back to our example of a new medicine that might be effective against headaches, you could design an experiment in which there are three groups of patients—one group receives the medicine (the test group), one group receives no medicine (the negative control group), and one group receives a medicine that is already known to be effective against headaches (the positive control group). All of the other variables, such as age, gender, and socioeconomic background, would be similar among the three groups.

These three groups help the researchers to make sense of the data. Imagine for a moment that there was just the test group with no control groups, and the headaches went away after treatment. You might conclude that the medicine alleviates headaches. But perhaps the headaches just went away on their own. The negative control group helps you to see what would happen without the medicine so you can determine which effects in the test group are due solely to the medicine.

In some cases, researchers control not just for the medicine (one group receives medicine and one does not), but also for the act of giving a medicine. In this case, one negative control involves giving no medicine, and another involves giving a placebo, which is a sugar pill with no physiological effect. In this way, the researchers control for the potential variable of taking medication. In general, for a controlled experiment, it is important to be sure that there is only one difference between the test and control groups.

Correct.

Incorrect.