Pre-screening of crude peptides in a serological bead-based suspension array.

Most serological assays detect antibody responses in biological samples through affinity of serum antibodies for antigens provided in the assay. Certain antigens, however, may be difficult to produce and/or may contain unwanted epitopes. In these cases, a practical alternative may be the use of peptides as representatives for specific epitopes. Peptides can be obtained after purification in large quantities for a modest price, but screening of a large set of peptides during development may be relatively expensive. To cut costs of screening peptides for a new serological assay, the concept was investigated of using cheap non-purified (crude) peptides instead of purified peptides. Peptides were selected that represent three well-described linear epitopes of viral proteins: VP2 of canine parvovirus (CPV), gp41 of human immunodeficiency virus (HIV) and E2 of classical swine fever virus (CSFV). Crude and purified biotinylated peptides with either a short or long spacer between the biotin and the epitope were used to test their capability to bind antibodies in a bead-based suspension array. The results show that, in a bead-based suspension array, crude peptides can function as antigen for specific monoclonal antibodies, and that the acquired signals are less than with purified peptides. CSFV-derived crude peptides were also able to detect specific antibodies in swine serum, indicating the applicability of crude peptides for pre-screening large numbers of different peptides during the development of serological peptide-based assays.

Synthesis of water-soluble scaffolds for peptide cyclization, labeling, and ligation.

The synthesis and applications of water-soluble scaffolds that conformationally constrain side chain unprotected linear peptides containing two cysteines are described. These scaffolds contain a functionality with orthogonal reactivity to be used for labeling and ligation. This is illustrated by the chemical ligation of two dissimilar constrained peptides via oxime ligation or strain-promoted azide-alkyne cycloaddition in aqueous media.