title: A method for accurate spatial registration of PET images and histopathology slices.
abstract: BACKGROUND
  Accurate alignment between histopathology slices and positron emission tomography (PET) images is important for radiopharmaceutical validation studies. Limited data is available on the registration accuracy that can be achieved between PET and histopathology slices acquired under routine pathology conditions where slices may be non-parallel, non-contiguously cut and of standard block size. The purpose of this study was to demonstrate a method for aligning PET images and histopathology slices acquired from patients with laryngeal cancer and to assess the registration accuracy obtained under these conditions.
  
  
  METHODS
  Six subjects with laryngeal cancer underwent a (64)Cu-copper-II-diacetyl-bis(N4-methylthiosemicarbazone) ((64)Cu-ATSM) PET computed tomography (CT) scan prior to total laryngectomy. Sea urchin spines were inserted into the pathology specimen to act as fiducial markers. The specimen was fixed in formalin, as per standard histopathology operating procedures, and was then CT scanned and cut into millimetre-thick tissue slices. A subset of the tissue slices that included both tumour and fiducial markers was taken and embedded in paraffin blocks. Subsequently, microtome sectioning and haematoxylin and eosin staining were performed to produce 5-μm-thick tissue sections for microscopic digitisation. A series of rigid registration procedures was performed between the different imaging modalities (PET; in vivo CT-i.e. the CT component of the PET-CT; ex vivo CT; histology slices) with the ex vivo CT serving as the reference image. In vivo and ex vivo CTs were registered using landmark-based registration. Histopathology and ex vivo CT images were aligned using the sea urchin spines with additional anatomical landmarks where available. Registration errors were estimated using a leave-one-out strategy for in vivo to ex vivo CT and were estimated from the RMS landmark accuracy for histopathology to ex vivo CT.
  
  
  RESULTS
  The mean ± SD accuracy for registration of the in vivo to ex vivo CT images was 2.66 ± 0.66 mm, and the accuracy for registration of histopathology to ex vivo CT was 0.86 ± 0.41 mm. Estimating the PET to in vivo CT registration accuracy to equal the PET-CT alignment accuracy of 1 mm resulted in an overall average registration error between PET and histopathology slices of 3.0 ± 0.7 mm.
  
  
  CONCLUSIONS
  We have developed a registration method to align PET images and histopathology slices with an accuracy comparable to the spatial resolution of the PET images.
authors.affiliation: Division of Imaging Sciences and Biomedical Engineering, King's College London, London, UK. tanujpuri82@gmail.com.|Present address: Department of Oncology, University of Oxford, Oxford, UK. tanujpuri82@gmail.com.
authors.forename: Tanuj
authors.initials: T
authors.lastname: Puri
authors.affiliation: Division of Imaging Sciences and Biomedical Engineering, King's College London, London, UK. anastasia.chalkidou@kcl.ac.uk.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Chalkidou
authors.affiliation: Oral Pathology Department, King's College London, London, UK. Rhonda.Henley-Smith@gstt.nhs.uk.
authors.forename: Rhonda
authors.initials: R
authors.lastname: Henley-Smith
authors.affiliation: Division of Imaging Sciences and Biomedical Engineering, King's College London, London, UK. roy.arunabha@gmail.com.
authors.forename: Arunabha
authors.initials: A
authors.lastname: Roy
authors.affiliation: Department of Oncology, Cancer Research UK and Medical Research Council Oxford Institute for Radiation Oncology, University of Oxford, Oxford, UK. paul.barber@oncology.ox.ac.uk.|Institute for Mathematical and Molecular Biomedicine, King's College London, London, UK. paul.barber@oncology.ox.ac.uk.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Barber
authors.affiliation: Department of Clinical Oncology, Guy's & St Thomas' NHS Foundation Trust, London, UK. Teresa.GuerreroUrbano@gstt.nhs.uk.
authors.forename: Teresa
authors.initials: T
authors.lastname: Guerrero-Urbano
authors.affiliation: Department of Head & Neck Surgery, Guy's & St Thomas' NHS Foundation Trust, London, UK. richard.oakley@gstt.nhs.uk.
authors.forename: Richard
authors.initials: R
authors.lastname: Oakley
authors.affiliation: Department of Head & Neck Surgery, Guy's & St Thomas' NHS Foundation Trust, London, UK. ricard.simo@gstt.nhs.uk.
authors.forename: Ricard
authors.initials: R
authors.lastname: Simo
authors.affiliation: Department of Head & Neck Surgery, Guy's & St Thomas' NHS Foundation Trust, London, UK. jean-pierre.jeannon@gstt.nhs.uk.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Jeannon
authors.affiliation: Department of Head & Neck Surgery, Guy's & St Thomas' NHS Foundation Trust, London, UK. mark.mcgurk@kcl.ac.uk.
authors.forename: Mark
authors.initials: M
authors.lastname: McGurk
authors.affiliation: Oral Pathology Department, King's College London, London, UK. edward.odell@kcl.ac.uk.
authors.forename: Edward W
authors.initials: EW
authors.lastname: Odell
authors.affiliation: Division of Imaging Sciences and Biomedical Engineering, King's College London, London, UK. michael.o'doherty@kcl.ac.uk.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: O'Doherty
authors.affiliation: Division of Imaging Sciences and Biomedical Engineering, King's College London, London, UK. paul.marsden@kcl.ac.uk.
authors.forename: Paul K
authors.initials: PK
authors.lastname: Marsden
keywords: Histopathology; Methodology; Oncology; PET; Registration