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Fig. 6. TM9SF4 mutant larval haemocytes have defective phagocytosis and encapsulation. (A) Circulating plasmatocytes were isolated from third instar larvae and incubated for 15 minutes with fluorescent latex beads. The internalisation of FITC-latex beads was observed following addition of quenching Trypan Blue solution. (B) Using the same procedure as in A, the internalisation rate of FITC-labelled beads or E. coli or S. aureus, was calculated as the number of internalised particles per haemocyte from 300-500 haemocytes. A phagocytic rate of 100% was attributed to control Rev45 cells in each experiment. The results are the mean ± s.d. of three independent experiments. A significant difference (Student's t-test, P<0.03) was found in phagocytic rate for latex beads and E. coli, but not S.aureus between Rev45 and TM9SF4 mutant cells (left panel). Directed expression of TM9SF4 mainly in the haemocyte lineage through the srpGal4 driver line (srpGal4/Y; TM9SF4¹; UASTM9SF4/+ larvae) partially rescued the phagocytic properties of circulating plasmatocytes (right panel) (P<0.01, Student's t-test). (In this experiment, larvae were raised at 18°C.) (C) Encapsulation rate of control (w¹¹¹⁸, Rev45) and mutant (TM9SF4) larvae following wasp parasitisation. The total number of parasitised larvae examined is indicated on the top of each histogram, the number in parenthesis indicates the number of larvae presenting a dark capsule. Experiments were performed at 24°C and 29°C as indicated.

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