title: A versatile cost-effective method for the analysis of fresh frozen tissue sections via matrix-assisted laser desorption/ionisation imaging mass spectrometry.
abstract: BACKGROUND
  There are currently multiple methods available for the preparation of fresh frozen tissue samples for analysis via matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) imaging mass spectrometry (IMS). Although these methods report excellent results, many are expensive automated approaches. With no published attempt to standardise less expensive manual processes, our work aims to provide a robust and repeatable method of sample preparation for MALDI-TOF-IMS that is applicable to a variety of tissue types, well explained, simple and cost effective.
  
  
  METHODS
  Fresh frozen tissue was sectioned at 12 µm and mounted onto liquid nitrocellulose coated slides, washed in a graded alcohol series and then mounted into a modified sublimation apparatus. Matrix is deposited onto the slide to achieve a desired coating of 0.2 mg/cm(2). Once coated, the slide is mounted into a custom-built vapor chamber and recrystallised with 50% acetonitrile (ACN), 0.1% trifluoroacetic acid (TFA) for 1 h at 37°C. The slide is then analysed using MALDI-IMS.
  
  
  RESULTS
  We have successfully implemented this method for a host of tissue samples, including brain, liver, kidney and heart, with no variation in relative spectra or processing method required. When the protocol is followed correctly, sublimations and recrystallisations are highly predictable with limited variation between samples and a very low failure rate. Additional apparatuses can be easily constructed by following the included instructions, that perform as per specifications with no variation.
  
  
  CONCLUSIONS
  We believe that we have described a complete protocol for MALDI-IMS that is easy to use and highly reproducible. The lack of expensive commercially available equipment makes this process very cheap with a relatively low initial outlay and our hope is that more laboratories will begin IMS-based avenues of research based on the work we have performed.
authors.affiliation: Proteomics Core Facility, University of Technology Sydney, Cnr Harris and Thomas St, Ultimo, NSW, 2007, Australia.
authors.forename: Matthew B
authors.initials: MB
authors.lastname: O'Rourke
authors.affiliation: The iThree Institute, University of Technology Sydney, Cnr Harris and Thomas St, Ultimo, NSW, 2007, Australia.
authors.forename: Benjamin B A
authors.initials: BB
authors.lastname: Raymond
authors.affiliation: The iThree Institute, University of Technology Sydney, Cnr Harris and Thomas St, Ultimo, NSW, 2007, Australia.
authors.forename: Steven P
authors.initials: SP
authors.lastname: Djordjevic
authors.affiliation: Proteomics Core Facility, University of Technology Sydney, Cnr Harris and Thomas St, Ultimo, NSW, 2007, Australia.
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Padula
mesh_terms: D000818:Animals; D001923:Brain Chemistry; D003460:Crystallization; D005629:Frozen Sections / Q000191:economics / Q000379:methods*; D008962:Models, Theoretical; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*