Fungal Species Identification by MALDI-ToF Mass Spectrometry.
Methods Mol Biol. 2017;1508:323-337
Authors: Bader O
MALDI-TOF MS has become the standard method for routine identification of most microbial organisms in clinical laboratories and has largely replaced biochemical assays. Classification relies on extensive well curated databases, ideally covering the full spectrum of microorganisms encountered in the specimens at hands. The protocols for harvesting cells and procuring material suitable for downstream MALDI-TOF MS analyses vary in specific details between the different groups of organisms, e.g., gram-positive or -negative bacteria, mycobacteria, or fungi. With respect to fungi, methods further vary between yeasts and moulds; and even among different mould genera if they do not lyse in a similar fashion. Purification of microbial materials from clinical specimen allows the direct identification of bacteria; however this is not yet fully adapted to fungi. In this chapter, I look into the differences between the underlying methods for yeast and moulds, and for production of samples suitable for MALDI-TOF MS species identification from cultures and different clinical materials.
PMID: 27837514 [PubMed - indexed for MEDLINE]
Evaluation of chromatographic conditions in reversed phase liquid chromatography-mass spectrometry systems for fingerprinting of polar and amphiphilic plant metabolites.
Anal Bioanal Chem. 2016 Aug;408(21):5855-65
Authors: Nielsen NJ, Tomasi G, Christensen JH
Metabolic fingerprinting is a relatively young scientific discipline requiring robust, yet flexible and fit-for-purpose analytical methods. Here, we introduce a simple approach to select reversed phase LC systems with electrospray MS detection for fingerprinting of polar and amphiphilic plant metabolites. The approach does not rely on isotopic labeling or biological origin of sample constituent and can also be used for non-biological matrices (e.g., oil or sewage sludge) or for other optimization purposes (e.g., mass spectrometric source parameterization). The LC systems varied in column chemistry and temperature, mobile phase pH/additive, gradient steepness/eluotropic strength, and electrospray mode of operation. The systems were evaluated based on the number of features detected using the matchedFilter algorithm from XCMS and the repeatability of this detection across analytical replicates. For negative ion mode detection, the best performances were obtained with an HSS T3 column operated at low pH, which produced a 3-fold increase in the number of reliable features extracted compared with the worst system. The best system for positive ion mode (i.e., the BEH C18 column operated at intermediate pH) only produced a 50 % increase in the number of reliable features. The data also indicate that baseline removal is unavoidable for reliable intensity estimations using peak areas, and that peak heights may be a more robust measure of intensity when baselines cannot be completely removed or in case of coelution, fronting or tailing.
PMID: 27344456 [PubMed - indexed for MEDLINE]