Figure 1.

Expression of chimeric sIgA in Sp2/0 cells. (A) Schematic representation of the human SC expression vector containing genes for histidinol and ampicillin resistance as well as a 3.2-kb EcoRV-BamHI DNA fragment including a 1.82-kb human SC coding sequence with a stop codon introduced at amino acid 590, the position of normal SC processing, fused to a 1.42-kb Ig 3′ region with a poly(A) addition site (Ig 3′ untranslated region). (B) Assembly and secretion of SC. Transfectants secreting sIgA were pulsed for 5 min with a mixture of ³⁵S-labeled methionine and cysteine and chased with a 100-fold excess of unlabeled methionine and cysteine. At the specified times, cells were rapidly cooled to 0°C and lysed by boiling in buffer containing SDS. SC and molecules covalently bound to SC were precipitated as described. The immune complexes were analyzed by SDS/PAGE in 6% Tris⋅glycine gels under nonreducing conditions. Molecular mass standards are indicated on the left. The positions of sIgA and SC are marked on the right. (C) Analysis of immunoprecipitates in 12.5% Tris⋅glycine gels under reducing conditions. The prestained molecular mass protein standards (Amersham) are indicated on the left; the positions of SC, α, κ, and J chain, on the right.