C cell (parafollicular cell) —Immunoreactive thyroglobulin: Purification, identification and immunological characterization

Summary

In relation to our earlier finding that the thyroglobulin-like material responsible for the cytochemical immunoreaction of C cells was obtained in the peak I fraction of Bio-Gel A-5 m, which included faster sedimenting components of thyroglobulin, the present study has identified the positive reacting component and clarified its immunochemical and immunohistochemical properties.

• 1. The peak I fraction of dog and hog thyroglobulin was chromatographed on a Bio-Gel A-50 m column. Antiserum to the faster eluted peak I ^′₁ only immunoreacted with C cells. The peak I ^′₁ was then refiltered on Bio-Gel A-150 m column. Antiserum to peak I ^″₁ fraction of both species which was eluted in the first part had high immune specificity for C cells.

• 2. When 4–30% and 2–16% continuous gradient gels of polyacrylamide were employed, peak I ^″₁ represented a single electrophoretic band corresponding to the component with the largest molecular weight in thyroglobulin. The protein was named C-thyroglobulin. The molecular weight was approximately 2,600,000, four times as large as 19 S, as calculated by relative mobility on the 2–16% gradient gel.

• 3. In double diffusion tests, anti-peak I ^″₁ antiserum produced two immunoprecipitin lines with its own antigen. The reaction was different from that of anti-19 S antiserum which formed a single line.

• 4. On immunoperoxidase staining, anti-peak I ^″₁ antiserum reacted to C cells in exactly the same way as anti-calcitonin antiserum.

• 5. When anti-peak I ^″₁ antiserum was absorbed with calcitonin, the subsequent reaction of the C cells was greatly decreased. The absorption of anticalcitonin antiserum with increased amounts of peak I ^″₁ abolished the C cell reaction. On the basis of these observations, the possibility that C-thyroglobulin is a biosynthetic precursor of calcitonin exists.