A method for application of samples to matrix-assisted laser desorption ionization time-of-flight targets that enhances peptide detection.
Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry has become a fundamental tool for the identification and analysis of peptides and proteins. MALDI-TOF is well suited for the analysis of complex biological mixtures because samples are crystallized onto a solid support that can be washed to remove contaminants and salts prior to laser desorption. A number of approaches for immobilizing samples onto MALDI targets have been put forth. These include the use of different chemical matrices and the immobilization of samples onto different solid supports. In large part though, the preparation of MALDI targets has been an empirical exercise that often requires a unique series of conditions for every sample. Here, a simple method for the application of peptide mixtures onto MALDI targets is put forth. This method differs because peptides are added directly to a sample of nitrocellulose dissolved in acetone, allowing them to interact in solution-phase organic solvent. This solution-phase mixture is then spotted to the MALDI target and evaporated, forming a homogenous solid surface for laser desorption. This procedure is robust, highly sensitive, tolerant to detergents, and easily learned. In our hands, the method provides as much as a 10-fold enhancement to the detection of tryptic peptide fragments derived from in-gel digests.
Landry
F
F
The Program on Cell Adhesion, The Burnham Institute, La Jolla, California, 92037, USA.
Lombardo
C R
CR
Smith
J W
JW
D000818:Animals; D002417:Cattle; D003902:Detergents; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis / Q000302:isolation & purification; D010446:Peptide Fragments / Q000032:analysis / Q000302:isolation & purification; D010455:Peptides / Q000032:analysis* / Q000302:isolation & purification; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D003902:Detergents; D008565:Membrane Proteins; D010446:Peptide Fragments; D010455:Peptides