Incorporating Biotechnology into the Classroom - VIRUS EPIDEMIC LESSON PLAN

Detection and Spread of a Viral Epidemic in Arizona

INTRODUCTION

This is a diagrammatic representation of the Hepatitis B virion and the surface antigen components. Hepatitis B virus causes both acute and chronic liver infections in man.

A recent outbreak of the hepatitis B virus has just been reported in the Phoenix area. An extraordinarily large number of patients have been diagnosed with the virus. The virus can be spread by the exchange of bodily fluids, including kissing. Several cases have been reported at this school, at least one of which is present in this class.
You have shared fluids with at least one person in this class. After obtaining a sample of your body fluids, it will be assayed for the presence of the hepatitis B virus proteins by a Western blot. The spread of the virus will be assessed and the route of progression the possible epidemic determined.

NOTE : This lesson plan deals with discussions on sexually transmitted diseases and benefits of monogamy. If desired, the lesson plan can be altered to deal with the influenza virus which can be transmitted by being in close proximity to infected individuals.

OBJECTIVES

1. Learn about viruses.
2. Learn how to perform a Western blot.
3. Learn about proteins, antigens, and antibodies.
4. Learn about epidemiology and disease progression and spread.

PROCEDURES

DAY 1

Part I (Fluid mixing)

1. Obtain a 1ml body fluid sample (one per student)
2. Obtain a piece of nitrocellulose (approx. 1/2" x 3). Pick up the nitrocellulose with tweezers. Never touch the nitrocellulose with your fingers.
3. Label the nitrocellulose on the bottom edge with a number corresponding to each person in a group, again being sure not to touch the nitrocellulose with your fingers (Use a ball-point pen)
4. Pipet 2 ul of each sample onto the nitrocellulose on the left side of initials.
5. Students should be separated into pairs (and record who they are paired with)
6. Each pair of students will mix their fluid samples together.
7. Students then pair off with a different person (and record who they are paired with)
8. Each pair of students now discuss whether or not they wish to mix their fluid sample. If they choose to mix, then they mix their samples together (students record whether they mixed or not).
9. Students repeat steps 4 and 5 two more times (being sure to record who they are paired with).

Part II (Western blot)

1. Pipet 2 ul of each sample onto the nitrocellulose to the right side of initials.
2. Allow the drops on the nitrocellulose to dry 1 minute.
3. Place the nitrocellulose in a tray and add 10 ml Milk solution
4. Incubate overnight in a refrigerator

DAY 2:

1. Add 5 ul secondary antibodies and mix (Keep antibodies on ice while using)
2. Incubate 15 minutes with occassional shaking
3. Pour off the antibody/milk mixture
4. Rinse the nitrocellulose three times with 10 ml Rinse solution per wash. (1minute per wash)
5. Add 10 ml Rinse solution to the Developer Chemicals and mix well (this is the Developer solution).
6. Pour off the last rinse and add 10 ml Developer solution
7. Incubate until color can be seen (approx 2 minutes) with occassional shaking
8. Wash nitrocellulose in water and record results:
   Positive = purple color
   Negative = no color

RECIPES

Rinse solution: (store at room temperature)
0.2 M Tris pH 7.5
1 mM MgOAc
(Autoclave to sterilize)

Milk solution: (Stored at -20^oC)
3 g carnation dry milk / 100 ml
20 mM Tris, pH 7.5
180 mM NaCl
0.02% Na-Azide

Developer chemicals: (store at -20^oC in 15 ml conical tubes)
5 mg Fast Blue BB salt (Sigma #F3378) plus 3 mg Naphthol AS Phophate (Sigma #N5625)

DATA:

Student pairs Mixing (yes/no)



RESULTS:


Students with a positive Western blot result have been infected with the virus. These students will record their names on the classroom board. Positive students:





Plot the spread of the virus in order to determine who was the original infected person.