Fig. 5.

Confirmation of antibody specificity in regards to HsMUC13’s localization within P. berghei during liver-stage infection. a Confirmation of both MUC13 knockdown, in cells expressing a pool of four anti-mucin13 shRNAs which have been integrated into the hepatocyte genome, and MUC13 knockout, via CRISPR/Cas9, during infection via western blot. Values indicated are relative protein expression as determined by densitometry. b Effect of MUC13 knockdown and knockout in the indicated cells infected with P. berghei expressing either luciferase (measured via total luminescence) or GFP (measured by FACS). Data is presented as mean ± s.e.m with n = 3 with individual biological replicates overlaid. c Confocal microscopy images of HC04 liver cells, either unmodified (WT), expressing an shRNA pool for MUC13 (KD) or with MUC13 knocked out (KO clone B10), infected with P. berghei 48 hpi. Cells were labeled with a rabbit polyclonal antibody (dilution 1:500, 1 mg/ml stock) against the intracellular domain of HsMUC13 (MUC13 antibody #1 AbCam #Ab65109). P. berghei was detected using a UIS4 antibody (1:500 dilution, 1 mg/ml stock LS-204260, LifeSpan Biosciences). Primary antibodies were detected with a goat anti-rabbit (Alexa Fluor 488, green) and a goat anti-mouse (Alexa Fluor 647, red). Cell membranes and nuclei were stained with Hoechst 33342 (blue) and CellMask deep red (magenta), respectively. Merged images between HsMUC13, P. spp HSP70, and Hoechst shown. Scale bars 10 μm; 63× oil objective