Mouse anti myc monoclonal antibody 9b11

Thin blood smears from cultured parasites were prepared and fixed as previously described (29 (link)). Blocking was done with 10% normal goat serum (Invitrogen)–PBS at 37°C for 30 min. To confirm the expression of Cas9, blood smears were immunostained with mouse anti-FLAG monoclonal antibody (M20008; Abmart, Shanghai, China) at 1:100 dilutions in PBS with 0.05% Tween 20 (T-PBS) and incubated at 37°C for 60 min. To evaluate myc epitope tagging of SBP3, the slides were immunostained with mouse anti-myc monoclonal antibody (9B11; Cell Signaling Technology, Danvers, MA, USA) at a 1:500 dilution. After 3 washes with T-PBS, the blood smears were incubated with Alexa Fluor 488-conjugated secondary goat anti-mouse IgG antibody (Invitrogen) (1:500 dilutions) at 37°C for 30 min. The smears were washed again 3 times with T-PBS and incubated with 1 μg/ml of Hoechst 33342 solution (Dojindo, Kumamoto, Japan) at 37°C for 20 min. After 3 washes with T-PBS, the smears were examined under a Nikon A1R confocal laser scanning microscope (Nikon, Tokyo, Japan).