October 10, 2002 cultured cell TIG-3 Human, homo sapiens Japanese male 10-week embryo no lung TIG-3 no no 2 mg/ml 8.5 M urea, 2% Triton X-100, 0.2% SDS, 0.4% Ampholine sonication 100,000 x g, 15 min no Immobilen drystrip, 18 cm, pH 4-7 8.5 M urea, 2% Triton X-100, 10 mM acetic acid, 0.01% Orange G, 0.4% Ampholine 15 micro-l on filter paper 300 V-2 h, 500V-0.5 h, 700 V-0.5 h, 1000 V-0.5 h, 1500 V-0.5 h, 2000 V-0.5h, 2500 V-0.5 h, 3000 V-0.5 h, 3500 V-6 h iodoacetamide 7.5% homogeneous gel plate, 19 x 18 cm Cathode: Tris-Tricine, Anode: 0.1 M Tris-Tricine 30 mA constant Silver staining Kit Wako PDQuest, UNIX version manual Trypsin, 30% ACN condition no MALDI-TOF MS, micromass M@LDI ssp5005 Ubiquitin C-terminal hydrolase L1 (UCH-L1) 23.8 kDa 24824.34 Da 5.8 5.33 15 PDL 1112 ppm 40 PDL 867 ppm 65 PDL 806 ppm 74 PDL 686 ppm 76 PDL 631 ppm 78 PDL 573 ppm trypsin 886.51, 981.51, 1064.59, 1815.95, 1842.94 No data No data any site No modification was detected P09936 X04741 Alzheimer's disease brain Carbonylation Free radic Biol Med 2002, 33(4):562, Castegna, et al. ssp5007 stathmin 16.3 kDa 17,171 Da 5.8 5.77 15 PDL 56 ppm 40 PDL 38 ppm 65 PDL 117 ppm 74 PDL 292 ppm 76 PDL 173 ppm 78 PDL 50 ppm trypsin 889.49, 962.38, 1074.60, 1165.58, 13.26.73, 1548.72 No data 1165.55 AIEENNNFSK Ala1 Acetylated Ser15 and Ser24 Phosphorylated SWISS-PROT P16949 GenBank J04991 ssp7001 stathmin 17.3 kDa 17,171 Da 6.0 5.77 15 PDL 116 ppm 40 PDL 120 ppm 65 PDL 382 ppm 74 PDL 476 ppm 76 PDL 265 ppm 78 PDL 137 ppm trypsin 889.49, 962.38, 1074.60, 1165.59, 1326.73, 1388.78 No data 1165.55 AIEENNNFSK Ala1 Acetylated any site No phosphorylation SWISS-PROT P16949 GenBank J04991