Figure 2.

Generation of SC-specific antisera and detection of the SC protein in whole-saccule lysate. (A) Schematic of the predicted domain organization of the SC and the location of the immunizing peptides. The location of the amino and carboxy noncollagenous domains (N-NC and C-NC, respectively) are indicated. The SC consists of a single fibrillar, collagenous domain (indicated by the black bar) separating amino- and carboxyl-terminal noncollagenous domains (indicated by the black zig-zag pattern). The number of amino acids in each domain is indicated below that domain. The approximate location of the sequences of the SC-specific peptides are indicated by the boxed portions of the SC protein. C-NC₁ and C-NC₂, SC carboxyl-terminal noncollagenous domain peptides; N-NC, SC amino-terminal noncollagenous domain peptide. (B) The reactivity of sera to 1 μg of the indicated immunizing peptides and lack of reactivity to irrelevant peptides was confirmed by examining the ability of 10⁻² and 10⁻³ dilutions of immune and preimmune sera to bind peptides immobilized on nitrocellulose. (C) Western blot analysis of whole sunfish saccule lysate with SC-specific antisera. In all panels, the position of the protein molecular weight markers (in kDa) is indicated with lines and the arrow(s) points at the detected protein(s). Each of the immune sera and the preimmune sera were diluted 1:500 for the analysis shown. Sera: anti-N-NC, SC amino-terminal noncollagenous domain peptide specific sera; anti-C-NC₁ or anti C-NC₂, SC carboxyl-terminal noncollagenous domain peptide-specific sera; or PI, preimmune sera.