QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

Help viewing high resolution images Return to article

Fig. 3. (A) The polyclonal antibody Isa-1 recognises the 90 kDa TbNRKC protein, which is expressed in the procyclic (Pro) and bloodstream forms (BS) of T. brucei (2.5x10⁶ cells/well loaded) on western blot. Controls: pre-immune on procyclic (lane 1) and bloodstream (lane 2) forms, no primary antibody (lanes 5 and 6). (B) The Isa-1 polyclonal antibody specificity to the NRKC protein was tested on western blot of the purified recombinant TbNRKC protein (lanes 1 and 2) and whole cells (lanes 3 and 4) without (-) or with (+) pre-incubation of Isa-1 with the recombinant protein (saturation assay). (C) TbNRKC is a basal body protein. T. brucei cytoskeletons (a-d) were prepared for immunofluorescence. A basal body and weak FAZ signals were observed using Isa-1 (c). Isa-1 specificity was assessed by saturation assays (e-h). A tetracycline induced (48 hours) NRKC-GFP expressing cell (i-l) illustrates direct GFP fluorescence on the basal bodies. Cytoskeletons of NRKC-GFP-expressing cells (m-p) probed with anti-GFP antibody (n) and YL-1/2 (o) confirms the basal body location of TbNRKC. a,e,i, phase-contrast images; m, phase-contrast image plus DAPI staining. Bar, 5 µm. (D) Flagella probed with Isa-1 visualised by electron microscopy. The majority of the Isa-1 labelling is on the proximal end of the mature basal body (arrowhead), some label is observed on the transition zone. The Isa-1 labelling of the immature basal body (arrow) cannot be clearly defined as proximal or distal because the immature basal body in vivo is orthogonal to the mature basal body. Some labelling can be observed between the mature and the immature basal bodies. Bar, 250 nm.

Return to article