EXERCISE 11

 

Immunology

 

Supplies required:

 

7 10 x 75 mm test tubes, 1 ml of 0.l mg/ml solution of bovine serum albumin (BSA), 5 ml of borate buffer, 0.5 ml of rabbit anti-BSA serum (0.5 ml), 7 capillary tubes (recommended size - 1.5-1.8 x 100 mm),tissues,

Critoseal support

 

1.         An Eppendorf tube with 1 ml of anti-BSA serum will be supplied. Label this tube #1.

2.         Obtain six empty Eppendorf tubes and label them #2-#7. Pipette 0.5 ml of pH 8 borate buffer into each of tubes #2-#7.

3.         Prepare 1:2 dilutions of anti-BSA in these tubes as follows:

a. Pipette 0.5 ml of anti-BSA from tube #1 into tube #2. Mix well.

            b. With a clean pipette, transfer 0.5 ml of the diluted anti-BSA serum from tube #2 to tube #3.

            Mix well.

            c. Repeat this dilution procedure through tubes #3-#6, using a clean pipette for each transfer.

4.         Tube #6 should now contain 1.0 ml of diluted anti-BSA serum.

5.         Tube #7 should contain only buffer and will serve as a negative control.

6.         Draw the antiserum from tube #1 into a capillary tube so that the tube is about one-half full.

            Wipe the tube carefully with tissue to remove excess serum and, taking care to avoid an air bubble, draw BSA (antigen) into the tube until the tube is filled to within 2 cm of the top.

            Withdraw the tube from the BSA solution and wipe the tube carefully with tissue.

            Allow an air bubble of about 1 cm to fill the bottom of the tube. Insert the bottom of the tube into the Critoseal support.

            Repeat with tubes #2 - #7.

7.         Incubate the tubes at room temperature for 30 minutes, looking for precipitation in each tube at ten minute intervals. Record your results in the table below as follows:

 

 

                        -        no precipitation

                        +/-      trace amount of precipitate (0-0.5mm)

                        +       small precipitate (0.5mm)

                        ++     precipitate (1-2mm)

                        +++ strong precipitate (3-4mm)

                        ++++ very strong precipitate (4mm or greater)

 

Record the titer (reciprocal of the highest dilution which shows precipitate).

II.        Human Pregnancy Tests: Practical Applications for Immunology

 

This will be done as a demonstration. 

 

Supplies required: Clearview ^TMHCG II pregnancy test; urine specimen

 

1.        Ensue that the test packet and the urine specimen are at room temperature.

2.        Put on gloves.

3.        Tear open the foil wrapper and remove the test. Place it on a clean, dry, flat surface.

4.         Insert the tip of the provided pipette into the urine sample and squeeze the top bulb fully.

5.         Release the top bulb to draw up the urine.

6.         Dispense the sample into the sample window by gently squeezing the top bulb.

            Do not squeeze the bottom bulb when using the pipette.

7.         Wait 5 minutes. a blue color will be seen moving across the windows as the test begins to work. A line should appear in the control window to indicate that the test is complete.

 

Results:

A blue line in the result window = positive for pregnancy.

No blue line in the result window = negative for pregnancy.

No blue line in the control window = failed test, inaccurate results.