This study investigates the immune response of Drosophila against pathogenic microbes by introducing bacteria into the hemolymph. The interaction between the bacteria and the immune cells is examined to understand the mechanisms of bacterial degradation and immune activation.
To investigate the immune response of Drosophila against pathogenic microbes, take a suspension of viable bacterial pathogens. Dip a sterile needle into the suspension, collecting a small amount.
Place an anesthetized fly under a dissection microscope. With the infected needle, prick the fly at the sternopleural plate of the thorax. The prick introduces bacteria into the hemolymph — circulatory fluid in the body cavity.
Place the infected flies inside a vial containing fly food, and maintain them for an adequate period.
The bacteria utilize the hemolymph as a rich source of nutrients and multiply — resulting in a systemic infection. Hemolymph contains hemocytes, circulating immune effector cells.
The hemolymph is also associated with the fat body, a specialized tissue for energy storage. The pattern recognition receptors on the fat body cells interact with the bacteria. This interaction induces the cells to secrete antimicrobial peptides, mediating bacterial killing.
The circulating hemocytes interact with the bacteria via phagocytic receptors, causing their engulfment into phagosomes and resulting in bacterial degradation.
A subset of the pathogens evades the host's immune response, leading to the death of the flies.
Monitor the fly mortality and perform downstream assays to assess the immune response against the pathogen.
To prepare the infecting pen, melt the end of a 200-microliter micropipette tip and insert a 0.15-millimeter insect minutien pin into the molten plastic. Allow the plastic to solidify so that the pin is held in place with 0.5 centimeters of pin extending from the plastic.
Anesthetize flies with carbon dioxide, and prick the flies in the sternal pleural plate of the thorax with the needle avoiding the attachment sites of the wings and the legs. Remove the flies from the minutien pin using soft forceps. Make sure to dip the pin into the bacterial suspension between pricking each fly.
Place the pricked flies into a fresh file with new fly medium laying the vial on its side until all of the flies have recovered from the anesthesia.
繁體中文
