Isolation of intact Sm/RNP antigens from rabbit thymus.
A comparison of the immunologically reactive components of the highly conserved Sm and RNP autoantigens from various mammalian tissue sources suggested the complete absence of a major 26K to 27K Sm-specific polypeptide in rabbit thymus extracts prepared by conventional procedures. A simple modification of the solubilization protocol, achieved by sonicating a suspension of commercial rabbit thymus acetone powder in 0.35 M NaCI, gave an extract containing the full complement of immunologically reactive Sm and RNP proteins detectable in other mammalian species. Without further manipulation, extracts were immediately passed through an immunoaffinity column constructed from human SLE IgG with both anti-Sm and anti-RNP reactivities. The proteins of the purified Sm/RNP were recovered in sufficient quantities for direct analysis by protein staining or immunoblot assays. The antigenic polypeptides were recovered intact and consisted of a single 73K RNP-specific species together with Sm-specific proteins of 26K to 27K (a doublet) and 13K. These proteins were easily visible by protein stain as were nonantigenic components of 35K, 32K, 11K, and less than 10K. The same polypeptides were present in affinity-purified Sm/RNP from HeLa cells, although the RNP protein was slightly smaller. The resolution and integrity of the complexes isolated by this simple two-step procedure, requiring less than 4 hr for completion, is remarkable, and the protein composition of the product compares quite favorably with antigens isolated from other sources by considerably more lengthy and laborious procedures.
D000818:Animals; D000941:Antigens / Q000302:isolation & purification*; D001324:Autoantigens; D002846:Chromatography, Affinity; D008970:Molecular Weight; D010455:Peptides; D011817:Rabbits; D017411:Ribonucleoproteins, Small Nuclear; D012995:Solubility; D013010:Sonication; D013950:Thymus Gland / Q000276:immunology*; D055517:snRNP Core Proteins
D000941:Antigens; D001324:Autoantigens; D010455:Peptides; D017411:Ribonucleoproteins, Small Nuclear; D055517:snRNP Core Proteins