Columbia University Summer Research Program for Science Teachers

How do we operate the spectrophotometer?

Jose Merced, John Dewey H.S., Brooklyn, NY

Mentor: Alan Stall, Department of Microbiology, College of Physicians and Surgeons

Columbia University, New York, NY

What ideas do you have for developing an Action Plan in this area?
I would like to share with my students three things that I feel would enrich my lessons:

a) The first is electrophoretic methods.

Electrophoresis is the movement of charged particles in an electric field. It has been applied to the movement of cells in suspension and of colloidal particles. It is useful in clinical chemistry for the identification of proteins.[Content Standard E- Understandings about technology] The rate of movement depends not only on the intensity of the electric field but also on the size, shape, and electric charge of the colloidal particle or molecule. Therefore, in making identification it may be helpful to perform electrophoresis at more than one pH.

Free and zone electrophoresis differ from each other in that the former a solution of the charged particle is placed directly in the electric field and analyzed. In the latter the solution being analyzed is supported on filter paper, a layer of starch granules, an agar gel, or some other solid or semisolid medium. This latter technique is simpler in decreasing the interference from diffusion and in providing stability to the separated fractions, since they are in a solid medium rather than in a liquid.

b) The second is the use of the hemacytometer.

The hemacytometer is a heavy glass slide manufactured to precise specifications of the National Bureau of Standards (NBS). It is used routinely in performing laboratory procedures such as red blood cell counts, white blood cell counts, and platelet counts but may also be used to count cells in other body fluids.

c) Use of the spectrophotometer to perform quantitative analysis of antibody and antigen precipitation reactions.

Students would benefit in the understanding of how standard curves are used to determine unknown concentrations of various solutions.

Do you anticipate implementation problems, and what ideas do you have about overcoming them?
The main problem that I anticipate is the inability to utilize viable murine cells for analysis. The method that I plan to use to overcome this obstacle is to order synthetic blood cells or to use yeast cells as a substitute for leukocytes. I also spent time during this past summer using the fluorescence microscope with attached camera to prepare projection slides that can be used in my classroom.


Aim: How do we operate the spectrophotometer?
I.O.: The student should be able to:
* Explain the principle of the spectrophotometer
* List the parts of the spectrophotometer
* State Beer's law
* Use the spectrophotometer

Write the following on the chalkboard:

1. Working in small groups, allow the students a few minutes to experiment with a ziplock bag containing the following items: small flashlight, piece of black cloth, tiny mirror, and a small piece of clear plastic.
2. Have them determine the properties of light using these items.
3. Elicit that light can be transmitted and absorbed. [Content Standard B- Energy/Matter Interactions]
4. Show the students a spectrophotometer.
5. Ask them to explain how it would work, by allowing them an opportunity to carefully inspect it.
6. List the parts and explain the function of each part. [Content Standard Unifying Concepts- Form and function]
7. Demonstrate how to use the spectrophotometer

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