Experimental factors affecting the quality and reproducibility of MALDI TOF mass spectra obtained from whole bacteria cells

Abstract

Numerous experimental factors are shown to significantly influence the spectra obtained when bacteria are analyzed by MALDI TOF/MS. Detailed investigation of the instrument parameters and sample preparation are all shown to influence the spectra. Of these, the preanalysis sample preparation steps incorporate the most important elements influencing the quality and reproducibility of the spectra. Some of the most important sample preparation factors include the method employed for sterilization, the type of matrix, the matrix solvent and concentration of cells in the matrix, as well as the type and concentration of acid added to the matrix. The effects of these parameters, as well as other aspects of sample preparation and the effects of several instrumental parameters on spectra are presented. Optimization and control of all experimental variables leads to a stable protocol for analysis of bacteria. The protocol employs a Nd:Yag laser and describes both sample handling and instrument conditions which consistently yield reproducible MALDI TOF mass spectra with greater than 25 peaks from both gram-positive and gram-negative bacteria.

Liquid chromatography/mass spectrometry characterization of Escherichia coli and Shigella species

“…The complexity of the lysate is another issue that may lead to ion suppression and/or detector saturation. Other factors such as the matrix and the acid content of the matrix solution can also play a role in the observation of higher mass ions [14], and finding optimal matrix conditions over a wide m/z range can be challenging. Although the LC step causes decreased throughput, this step is likely part of the reason more proteins are observed.…”

Applications of MALDI‐TOF Mass Spectrometry in Clinical Diagnostic Microbiology

“…The most frequently used matrices are 2,5-dihydroxybenzoic acid (DHB), a-cyano-4-hydroxycinnamic acid (CHCA), sinapinic acid (SA), ferulic acid (FA), and 2,4-hydroxy-phenyl benzoic acid. FA, SA, and CHCA have been shown to be effective for the detection of proteins biomarkers (Fenselau & Demirev, 2001;Vaidyanathan et al, 2002;Williams et al, 2003), whereas DHB appears to be the best choice for the detection of glycopeptides and glycoproteins (Giebel et al, 2010). The size and the intensities of the peaks of the detected molecules are dependent on the matrix selected for the experiment.…”

“…The size and the intensities of the peaks of the detected molecules are dependent on the matrix selected for the experiment. DHB and CHCA are usually optimal for the detection of lower mass ions (Hathout et al, 2000;Williams et al, 2003;Ruelle et al, 2004) with a detection up to 10 kDa when the proper solvent is used. Both SA and FA have been shown to be better for the detection of higher mass ions (above 15 kDa) (Madonna et al, 2000;Conway et al, 2001;Ruelle et al, 2004;Vargha et al, 2006) but provide a lower sensitivity than CHCA (Wang et al, 1998;Ruelle et al, 2004).…”

An Evaluation Of Direct Identification Of Pathogens From Blood Cultures By Maldi-Tof Mass Spectrometry

“…Lina Savickaitė, Jelena Kopeykinienė Klaipeda University Hospital, Lithuania sample preparation, matrix solutions and organic solvents, affect the quality and reproducibility of bacterial MALDI-TOF mass spectrometry fingerprints (11)(12)(13)(14)(15)(16). With a special set of MALDI Sepsityper is possible to identify bacteria directly from the positive blood culture (17)(18)(19)(20)(21)(22)(23)(24)(25).…”