Compared to continuous extraction, pulsed extraction (PE) of ions formed by matrix-assisted laser desorption/ionization (MALDI) in time-of-flight (TOF) mass spectrometers significantly improves mass resolution. Parameters such as extraction voltage, delay time, and correction pulse must be varied, however, to achieve optimum mass resolution over a broad mass range because the PE method is mass dependent. We previously reported a novel method, mass correlated acceleration (MCA), which we have now combined with a reflectron MALDI TOF mass spectrometer to further enhance mass resolution over a broader mass range. Unlike the PE method, MCA is not mass dependent and high resolution mass spectra can be achieved with a single tuning of instrument parameters. The ions may be brought into focus simultaneously, i.e., the multi-channel recording advantage can be more fully realized. The MCA dual-stage ion source design includes an extraction pulse region and an acceleration region that contains a time-dependent waveform correlated with mass. We demonstrate the validity of this novel technique with applications in peptide mixture analysis and protein digests of lysozyme and bovine serum albumin. [1,2] is effective in the ionization of large biomolecules (peptides, proteins, carbohydrates, and oligonucleotides) and other polymers in mass spectrometry. The development of MALDI sources has sparked renewed interest in the time-offlight (TOF) mass analyzer, an excellent counterpart to pulsed laser sources in MALDI. In addition, TOF analyzers offer the multichannel recording advantage capable of recording the entire mass range simultaneously (i.e., no scanning of the mass analyzer is required).Historically, the TOF analyzer has had the reputation of providing low mass resolution and, subsequently, poor mass accuracy. Uncertainties in time of ion formation, initial position, and kinetic energy contribute to the actual flight time. Hence, peak broadening (mass resolution) is affected by these conditions. To compensate for the initial temporal, spatial, and velocity ion distributions, Wiley and McLaren [3] introduced timelag focusing, a dual-stage ion source which provided first-order space focusing (with the detector positioned at an optimum distance) and permitted higher mass resolution measurements. Similar to what we now know as pulsed extraction (PE) or delayed extraction (DE), time-lag focusing used a time delay between the ionization pulse and the extraction pulse. Its more recent counterpart greatly improves the quality of MALDI mass spectra [4 -8]. A disadvantage to PE focusing, however, is its mass dependency: each mass is focused at a different time delay and/or extraction pulse amplitude. Therefore, one or both of these parameters need to be varied in order to obtain mass spectra containing peaks with optimum resolution across the entire mass range. There have been considerable improvements in mass accuracy and mass resolution over a broad mass range [9 -14]; however, our approach differs in that it utilizes a simple e...