Radioiodination and 125I-labeled peptide mapping of proteins on nitrocellulose membranes.
A rapid procedure for generating dozens of 125I-labeled peptide maps from a protein band excised from a single lane of a sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel has been developed. Proteins, which can be rapidly purified by 2 X SDS-PAGE separation, are electroblotted onto nitrocellulose paper (NCP) and located by aqueous naphthol blue-black staining. All subsequent steps of radioiodination, and enzyme or chemical cleavage, are carried out on the NCP making it possible to test a variety of cleavage reagents on the same protein sample. The resultant peptidic residues, which can be separated by thin-layer electrophoresis-thin-layer chromatography (2D TLE-TLC), SDS-PAGE, or HPLC, can be used in comparative studies or they can be recovered for further structural and immunological analyses.
D001426:Bacterial Proteins / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D003101:Collodion; D004591:Electrophoresis, Polyacrylamide Gel; D007457:Iodine Radioisotopes; D009344:Neisseria gonorrhoeae / Q000032:analysis; D010446:Peptide Fragments / Q000302:isolation & purification*; D012967:Sodium Dodecyl Sulfate
D001426:Bacterial Proteins; D007457:Iodine Radioisotopes; D010446:Peptide Fragments; D012967:Sodium Dodecyl Sulfate; D003101:Collodion