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Analytical and preparative electrophoresis separation techniques have been essential tools in protein biochemistry and the biological sciences in general. The combination of high resolution electrophoresis techniques with high performance analytical procedures has dramatically enhanced analytical protein biochemistry. In this report we describe the combination of electrophoretic separation techniques with electrospray ionization (ESI) tandem mass spectrometry (MS/MS). A series of different techniques, consisting of automated high performance liquid chromatography (HPLC)-MS/MS, capillary-HPLC-MS/MS, and solid phase extraction (SPE)-capillary zone electrophoresis (CZE)-MS/MS, are described in the context of the identification of high pmol to the low fmol amounts of proteins. Application of these powerful new tools for the analysis of proteins on a large proteome-wide scale is presented. Furthermore, the combination of orthogonal separation techniques, such as immobilized metal affinity chromatography (IMAC) with SPE-CZE-MS/MS, and IMAC followed by HPLC, and by SPE-CZE-MS/MS, are presented for the detailed investigation of post-translational modifications of specific proteins.

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Sensing nonradiation‐induced singlet oxygen (1O2) in whole‐animal is deemed as one of the most challenging tasks in noninvasive techniques due to the µs level lifetime of 1O2 and quenching by numerous reductants in tissues. Here a distinct chemiluminescent (CL) nanosensor (NTPE‐PH) that boasts ultrahigh concentrated CL units in one nanoparticle is reported. Taking advantage of the intramolecular energy transfer mechanism that promises high energy transfer efficiency and the aggregation‐induced emission behavior that guarantees high CL amplification, the NTPE‐PH sensor is sensitive to a nm level 1O2. Experiments demonstrate that the NTPE‐PH yields a highly selective CL response toward 1O2 among common reactive oxygen species. With proved low cytotoxicity and good animal compatibility, real‐time mapping of ultratrace 1O2 in whole‐animal during acute and chronic inflammations is first achieved. It is anticipated that the NTPE‐PH sensor can be a useful tool for monitoring 1O2 variation during immune response and pathological processes corresponding to different stimuli, even with drug treatment included.