Importance of matrix:analyte ratio for buffer tolerance using 2,5-dihydroxybenzoic acid as a matrix in matrix-assisted laser desorption/ionization-Fourier transform mass spectrometry and matrix-assisted laser desorption/ionization-time of flight.

Many biological samples destined for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) contain buffers. The presence of these buffers often inhibits the ability to obtain spectra. Here, the results of a study of the effects of six different buffers on spectra of three representative small proteins are reported utilizing 2,5-dihydroxybenzoic acid as matrix. These proteins, bovine insulin, cytochrome c, and bovine albumin have masses from approximately 5000 to 66,000 Da. Three different sample preparation techniques were investigated: aerospray, dried-drop, and acetone redeposition. Both MALDI Fourier transform and time-of-flight mass spectrometry results show that buffer tolerance of MALDI-MS samples depends upon several factors, including the relative amount of the buffer in the MALDI matrix, as well as the identity of the specific buffer. Furthermore, the rate at which buffer tolerance decreases as buffer concentration is increased varies from buffer to buffer. The current results reveal that, at very high matrix:analyte ratios, buffer tolerance of MALDI is dramatically greater than concluded in previous literature reports.