Influenza Haemagglutination: Page 3/23

Perform the experiment as follows:

	To the plastic microtitre plate, add 50µl of phosphate buffered saline (PBS) to each of the wells in the first two rows of the plate (A1-A12 and B1-B12). Use the 12th well of these two rows (A12 and B12) for the negative controls (no allantoic fluid). Why are these negative controls necessary? Add 50µl of undiluted influenza-containing allantoic fluid (AF) to A1 and mix gently. With a fresh pipette tip, transfer 50µl to A2 and mix gently. Repeat this procedure for the whole row, discarding 50µl from the last well in the series (A1-A11).
Click here to see how to set up the assay...

To the plastic microtitre plate, add 50µl of phosphate buffered saline (PBS) to each of the wells in the first two rows of the plate (A1-A12 and B1-B12). Use the 12th well of these two rows (A12 and B12) for the negative controls (no allantoic fluid). Why are these negative controls necessary?
Add 50µl of undiluted influenza-containing allantoic fluid (AF) to A1 and mix gently. With a fresh pipette tip, transfer 50µl to A2 and mix gently. Repeat this procedure for the whole row, discarding 50µl from the last well in the series (A1-A11).

Use a 100-fold dilution of the allantoic fluid in PBS to set up the second row (B1-B11) in the same way.
Using a clean pipette tip, add 50µl of red blood cells (RBC's) to each well, including the negative controls (resuspend the cells before using). Work from right to left (starting with the negative controls and and finishing with the lowest virus dilution).
Mix the cell suspensions by tapping the plate gently.