Similar to the methods described earlier [39 (link),48 (link)], a single colony of the dry cultures was removed from the agar plates using a toothpick, after which it was deposited on an MSP 384 target polished steel BC plate (Bruker Daltonics, Bremen, Germany) in 8 replicates and overlaid with 1 µL of α-cyano-4-hydroxycinnamic acid matrix solution (Bruker Daltonics, Bremen, Germany).
From the biphasic cultures, 500–700 µL was placed in 1.5 mL Eppendorf tubes and centrifugated at 2400× g for 5 min. Supernatants were decanted, and pellets were washed twice with 500 µL phosphate-buffered saline (PBS) and centrifugated for 5 min at 2400× g. Pellets were then resuspended in 20–100 µL PBS, after which 1 µL was spotted onto an MSP 384 target polished steel BC plate (Bruker Daltonics) in eight replicates, air dried and overlaid with 1 µL of α-cyano-4-hydroxycinnamic acid matrix solution (Bruker Daltonics).
No prior extraction protocol was performed as earlier tests revealed lower spectra quality (i.e., more variance and more dispersion of the peaks) compared to direct spotting.
From the biphasic cultures, 500–700 µL was placed in 1.5 mL Eppendorf tubes and centrifugated at 2400× g for 5 min. Supernatants were decanted, and pellets were washed twice with 500 µL phosphate-buffered saline (PBS) and centrifugated for 5 min at 2400× g. Pellets were then resuspended in 20–100 µL PBS, after which 1 µL was spotted onto an MSP 384 target polished steel BC plate (Bruker Daltonics) in eight replicates, air dried and overlaid with 1 µL of α-cyano-4-hydroxycinnamic acid matrix solution (Bruker Daltonics).
No prior extraction protocol was performed as earlier tests revealed lower spectra quality (i.e., more variance and more dispersion of the peaks) compared to direct spotting.