Part III.  Laboratory for Detecting Sickle Cell Anemia.
(To the teacher: this lab is best done AFTER students do the part II genetics activity, at the very least, and ideally do Parts I and II.)

You are a laboratory technician in the newborn screening section of the BBD Pathology Lab, a branch of the state health department.  You and your coworkers are going to test today’s samples of newborn blood to see if there are any cases of sickle cell anemia in the newborns. 

You already know that individuals with sickle cell anemia have abnormal hemoglobin in their red blood cells.  They make hemoglobin S instead of the normal hemoglobin A.  Carriers of the disease make hemoglobin A and S.  In the laboratory, you can use electrophoresis to detect abnormal hemoglobin protein because it is known that hemoglobin S moves slower than normal hemoglobin during electrophoresis. 

Materials:
Agarose gel 1%
Electrophoresis chamber
Electrophoresis buffer TBE
Power supply
Sample of normal hemoglobin A, labeled N for normal.
Newborn samples for screening, numbered P1, P2, etc. for Patient.
Micropipettors

Procedure:

    • Obtain two 0.5mL tubes, one containing the Normal (N), and one containing your Patient sample (P).  Record the number of your particular patient sample______.
    • Place a gel into the electrophoresis chamber.  Fill the reservoirs with buffer, and then add just enough buffer to cover the gel.
    • Load 10-15μL of each sample into a separate well with a micropipette.   Label the wells you used in the sketch below.
    • Note how your gel is oriented with respect to the positive and negative poles.  (Remember- Run to red.)
    • Place the cover on the chamber and plug the electrodes into to the power supply, black to black, red to red.
    • Turn on power supply; look for bubbles rising from the wires at each end to ensure the electrophoresis box is working correctly.
    • Run your gel at 100-125 volts until the dye is about ¾ of the way down the gel.  This should take about 10-15 minutes.
    • Turn off the power supply and disconnect the electrodes.

    Complete your laboratory report and submit to your laboratory supervisor.

    Test Your Understanding:

    1. If a person was carrier (heterozygous) for sickle cell hemoglobin, how many bands would you see?___________________

     

    1. If a person had sickle cell anemia, how many bands would you see?__________
    1. If a person was normal for hemoglobin, how many bands would you see?______

     

    1. How does the gel distinguish normal from sickle cell hemoglobin?

     

    1. Why do you need a Normal sample for each gel you run?

     

     

    6.Would it be valid to use a known sickle cell sample instead of a known Normal sample for each gel you run?
    Explain your answer:

     

    1. When you report your class results for these newborns, which samples were positive for sickle cell anemia:_________________________________
    1. Did you detect any carriers and if so, list:

     

    Credits: Megan Waugh and DeAnn Campbell of OCCC.

     

     

     

     

     

    		 Professional Development