About Preview™

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Preview™ quickly samples your data to measure mass errors, digestion specificity, and modifications for subsequent full search. Preview automatically generates a suggested parameters file for a subsequent Byonic search.   Click here to request a free trial copy of Preview.
Please see our publication on Preview from Analytical Chemistry 2011: Preview: a program for surveying shotgun proteomics tandem mass spectrometry data.


Database-search programs for peptide identification by tandem mass spectrometry ask their users to set various parameters, including precursor and fragment mass tolerances, digestion specificity, and allowed types of modifications. Even proteomics experts with detailed knowledge of their samples might find it difficult to choose optimal parameter settings. To address this problem, Preview runs fast initial searches to inform these choices, and to recalibrate precursor and fragment masses based on confident identifications. Preview samples the data set rather than performing an exhaustive search, so it is not a substitute for a full database-search program such as Byonic.

Mass tolerances.

The user sets tolerances for precursor and fragment masses that reflect the type of MS/MS instrument, quality of mass calibration, and data acquisition strategy. For example, on a Thermo Orbitrap instrument with Orbitrap MS and ion-trap MS/MS, the user might configure the program to consider peptides with mass within 10 ppm of the measured precursor mass and to score fragment ions with mass-over-charge (m/z) within 0.5 of the measured m/z of a peak. These generic settings may not work or be optimal for all data sets. The user must also decide if the precursor charge assignments are reliable, and if precursor masses reliably represent the monoisotopic masses or if they include off-by-one errors, in which the first 13C isotope peak is misidentified as the monoisotopic mass.

Digestion specificity.

The user can set the program to consider only peptides with digestion-specific cleavages at both termini (for trypsin, on the C-terminal side of arginine or lysine), or can choose a broader search, allowing nonspecific cleavage at one or both termini.  Nonspecific digestion can vary from negligible to ubiquitous depending upon endogenous peptidases, and missed cleavages vary widely depending upon sample processing conditions.


Perhaps the most difficult choices for the user are which peptide modifications to allow. Some in vitro modifications are ubiquitous, occurring to some extent in almost all shotgun proteomics samples, but others depend upon both the sample and its preparation and can vary unpredictably. Posttranslational modifications (PTMs) also vary from sample to sample and from protein to protein within a sample.

Parameter setting is crucial to obtaining the best results from a proteomics search engine, yet until now there has been no tool specifically designed to help users with this task. Some researchers search their data multiple times with various settings, but this “bracketing” strategy can be prohibitively slow for large data sets and still miss important modifications.

Preview Solution

We developed a fast search program, called Preview, to guide parameter setting for subsequent full searches. Preview has only two required inputs: a set of MS/MS spectra in mzML, mzXML, Thermo RAW or MGF format, and a protein database in FASTA format. The program then computes appropriate settings for precursor and fragment tolerances, estimates the amount and type of nonspecific digestion, measures the prevalence of recognized modifications, and can report unrecognized/unusual mass shifts. Preview operates in a fraction of the time of a standard search program; for example, a search of 10,000 spectra against a database containing 100,000 protein sequences takes about 1 minute on a common desktop PC.

In order to achieve this speed, we made a number of simplifying assumptions in the design of Preview. The foremost assumption is that the 100 most detectable proteins sufficiently represent the entire sample for the full menu of search parameters. Another simplification is that Preview, with some exceptions, searches for unrelated modification types one at a time, thereby avoiding the combinatorial explosion and performance degradation of multiple modification searches. Finally, Preview’s peptide identification algorithm takes shortcuts: it represents both predicted and observed peaks by integer masses, in order to allow a faster algorithm for candidate scoring.

Preview optionally recalibrates m/z measurements and outputs a new spectrum file (in .mgf format) that can be used as input to Byonic. The user can choose to recalibrate precursor measurements, fragment measurements, or both.

Preview can benefit a proteomics laboratory in several ways. Most obviously, it can help the user choose parameter settings for subsequent full search by Byonic or any other search engine. Secondly, even if the user already knows the best parameter settings, m/z recalibration can improve sensitivity and specificity by increasing the scores of true matches relative to those of false matches. Finally, timely feedback on sample preparation artifacts and m/z errors can help improve laboratory practices and experimental reproducibility.


Request a demo copy of Preview by registering here.

Tutorial and Help

Learn more about how Preview works on the Tutorial page and the Help page.