Biochemical and immunological characterization of the microfibrillar ecto‐endoplasmic boundary in the ciliate Isotricha prostoma

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Abstract
The cytoplasm of the ciliated protozoan I. prostoma is compartmented by a continuos fibrillar system made up of a double layer of 4 nm-diameter filaments: the microfibrillar ectoendoplasmic boundary (EEB). Isolation of this structure after treatment of the cells in a buffer of low ionic strength in the presence of the detergent Triton X-100 evidenced connections linking the 2 filamentous layers. One-dimensional electrophoresis on SDS[sodium dodecyl sulfate]-polyacrylamide gel of EEB fractions revealed several major proteins with apparent MW between 11 and 23 K [kilodaltons]. Of these, 2 neighboring bands of MW22 and 23 K were removed from gels and used as antigens to obtain rabbit antibodies. The antiserum obtained reacted specifically with injected proteins as shown by the technique of immunological detection on nitrocellulose sheets using the peroxidase reaction product. EM localization of the antigens with anti-IgG coupled with colloidal Au showed significant labeling of the EEB within the cortex of Isotricha permeabilized with Triton X-100. Apparently, the 22-23 K antiserum will prove to be a useful tool for the comparative study of other nonactin filament systems in Protozoa.