FIG 3.

Transcription and expression of C. parvum INS1 in vitro. (A) Relative transcription level of the INS1 gene (cgd1_1680) at specified times postinfection, as determined by reverse transcription-quantitative PCR. HCT-8 cells were infected with C. parvum oocysts and cultured for specific time points, and RNA was collected from three wells per time point. Gene expression profiles are from two separate experiments with different time points. Data from the Cryptosporidium 18S rRNA gene were used in data normalization. Values are plotted as the means ± standard deviations (SD). (B) Immunofluorescence staining of transgenic INS1-3HA parasites. HCT-8 cells were infected with INS1-3HA oocysts. After 48 hpi, coverslips were fixed and stained with rat anti-HA followed by goat anti-rat IgG Alexa Fluor 488, rabbit pan-Cp followed by goat anti-rabbit IgG Alexa Fluor 568, and Hoechst for nuclear staining. Scale bars, 2 μm. (C) Immunofluorescence staining of INS1-GFP parasites. HCT-8 cells were infected with INS1-GFP oocysts. After 48 hpi, coverslips were fixed and stained with rabbit anti-GFP followed by goat anti-rabbit IgG Alexa Fluor 488, rat pan-Cp followed by goat anti-rat IgG Alexa Fluor 568, and Hoechst for nuclear staining. Scale bars, 2 μm.