Spectronaut™ Pulsar X

More versatility in proteomics research

Spectronaut™ has delivered highest performance in protein analysis for years, specifically for discovery proteomics applications. Spectronaut™ Pulsar X offers seamless combination and integration of some of the previously separate workflows, extending the applications of the Spectronaut™ software suite.

  • Hybrid library generation: By combining core proteome libraries (project or resource libraries) with sample specific libraries (directDIA), the number of quantifiable proteins can be increased
  • Spike-in workflow: In combination with Biognosys’ new PQ500 reference peptide kit, targeted proteins can be label-free quantified using a comprehensive reference calibration curve spanning five orders of concentration
  • Host-cell proteins: Calibration carry-over allows for higher precision in identification and quantification of low abundance host-cell proteins

 

Spectronaut™ Pulsar X integrates DDA and DIA data for Hybrid Library generation, which yielded excellent results in our hands and opened up exciting possibilities for novel data acquisition strategies.”

Florian Meier
Max Planck Institute of Biochemistry, Martinsried, Germany
(Matthias Mann Group)

 

Hybrid Libraries

A data independent acquisition (DIA) experiment usually delivers a highly convoluted data set that requires a spectral library to extract the fragments of individual peptides hidden in the data. Libraries can be made directly from the samples that are analyzed (project specific library), from data repositories (resource library) or with directDIA - a library-free method that generates libraries on-the-fly from the DIA data.

 

All three library methods come with trade-offs

The main criteria for a good library are its depth and precision. The depth is limited to the peptides that are contained in the library. Only these peptides can be identified in DIA data and in practice larger libraries allow for the identification of more peptides, which translates to more proteins being identified. Library precision refers to how well do the retention times and fragment intensities match the actual data from a specific sample.

Project specific libraries are very precise but they are often limited in depth because the measurement time to generate a deep library may not be feasible. A resource library may be deeper but is usually less precise, because it may have been generated with different samples or a different instrument type compared to the experimental samples. DirectDIA method provides precise libraries from each single sample. However, the depth is lower compared to project specific libraries, which are usually generated from multiple fractionated samples.

 

The solution: Hybrid Libraries lead to best results because they are deep and precise

Spectronaut™ Pulsar X supports a new method of library generation that combines the strengths of all library types and avoids their limitations. The underlying concept is to cover the core proteome with a resource library (or project specific library) and complement the library with directDIA identifications that cover the sample specific proteome.

Spectronaut™ Pulsar X achieves this result through correct statistical treatment of both libraries independently. We call this method Hybrid Libraries. In developing the Hybrid Library technology, our focus was to maximize the number of quantifiable proteins. However, it also comes with a reduction in instrument time overhead, if a resource library is used in combination with directDIA.

To see a showcase of the capabilities of a Hybrid Library workflow, where we used a project specific library, a resource library, and a Hybrid Library to analyze a set of lung cancer samples in DIA mode, please download the Spectronaut™ Pulsar X brochure.

 

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