Establishment of MHC class I-GAD65 FvB transgenic mice. (A) Southern blot of tail genomic DNA. Lanes 1, 2, 4, and 5 were tg(+); lane 3 was non-tg(+). Lanes 1 and 2 are from line 27; lanes 4 and 5 from line 9. (B) Reverse transcription–PCR analysis of tg(+) (+) and non-tg(+) (−) tissues for transgene-specific RNA expression. M, 100-bp marker (GIBCO/BRL); B, brain; K, kidney; L, liver; S, spleen; T, thymus; I, pancreatic islets of Langerhans; C, water. (C) Immunoprecipitation/Western blot analysis of thymus lysates of 2-day-old non-tg(+) (lane 1), line 9 tg(+) (lane 2), and line 27 tg(+) (lane 3) mice. Immunoprecipitation was by mN65, Western by 7673 (see text). (D) GAD65 immunoprecipitation/Western blot analysis of lysates from adult tg(+) (+) or non-tg(+) (−) mice. Immunoprecipitation was by Stiff Man Syndrome patient serum, Western by mN65 (see text). B, brain; K, kidney; L, liver; S, spleen; T, thymus.