Diagnosis of Acanthamoebae infection using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
Description
The data was collected as part of the intra-ssay and inter-assay variation test. In each analytical run, six samples were prepared using the centrifugal membrane filters. Each sample was spotted onto six MALDI target spots, leading to six mass spectra per sample. For the intra-assay, 36 mass spectra were evaluated (n=36). The inter-assay encompasses n=108 mass spectra recorded on three consecutive days (36 mass spectra per day). The notation (e.g., A1) refers only to the MALDI target spot used. If there are not six mass spectra per sample per day, these remaining mass spectra were flatline spectra.
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Steps to reproduce
For centrifugal membrane filtration, filter columns with a 5 µm polyvinylidene fluoride membrane and 500 µL sample volume were applied from Sigma Aldrich Millipore® (Darmstadt, Germany). A centrifugation at 500 x g was performed. The sample preparation method was as follows: Amoeba saline (recipe by Page, 1988) with 0.1 % w/v Triton X-100 was used to “harvest” the Acanthamoebae. They were transferred into the filter column, centrifuged, and washed with ethanol (96 %). The retained Acanthamoebae were recovered from the filter by putting the column upside down into a tube overlaid with amoeba saline, followed by centrifugation. The resulting amoeba sediment was resuspended in distilled water, vortexed, transferred into a 2 mL tube, and mixed with ethanol (96 %). After centrifugation, the supernatant was discarded, and the amoeba sediment air-dried. It was then treated with formic acid (70 %) and acetonitrile and centrifuged again. The MBT™ Biotyper® smartfleX from Bruker Daltonik GmbH and Co. KG (Bremen, Germany) was used in our study. 1 µL of the supernatant was spotted onto the single-use MALDI target plate, air-dried, and overlaid with 1 µL of matrix α-cyano-4-hydroxycinnamic acid (HCCA) before final drying.
Institutions
- Hochschule Bonn-Rhein-Sieg Fachbereich Angewandte Naturwissenschaften
- Bundeswehrzentralkrankenhaus Koblenz