Fluorescence hyperspectral imaging (fHSI) using a spectrally resolved detector array

Anna Siri Luthman, Sebastian Dumitru, Isabel Quiros-Gonzalez, James Joseph, Sarah E. Bohndiek

Research output: Contribution to journalArticlepeer-review

27 Citations (Scopus)
68 Downloads (Pure)


The ability to resolve multiple fluorescent emissions from different biological targets in video rate applications, such as endoscopy and intraoperative imaging, has traditionally been limited by the use of filter-based imaging systems. Hyperspectral imaging (HSI) facilitates the detection of both spatial and spectral information in a single data acquisition, however, instrumentation for HSI is typically complex, bulky and expensive. We sought to overcome these limitations using a novel robust and low cost HSI camera based on a spectrally resolved detector array (SRDA). We integrated this HSI camera into a wide-field reflectance-based imaging system operating in the near-infrared range to assess the suitability for in vivo imaging of exogenous fluorescent contrast agents. Using this fluorescence HSI (fHSI) system, we were able to accurately resolve the presence and concentration of at least 7 fluorescent dyes in solution. We also demonstrate high spectral unmixing precision, signal linearity with dye concentration and at depth in tissue mimicking phantoms, and delineate 4 fluorescent dyes in vivo. Our approach, including statistical background removal, could be directly generalised to broader spectral ranges, for example, to resolve tissue reflectance or autofluorescence and in future be tailored to video rate applications requiring snapshot HSI data acquisition.

Original languageEnglish
Pages (from-to)840-853
Number of pages14
JournalJournal of Biophotonics
Issue number6
Early online date9 May 2017
Publication statusPublished - 29 Jun 2017


  • biomedicial
  • hyperspectral
  • in vivo
  • instrumentation
  • multiplexed fluorescence
  • SRDA

ASJC Scopus subject areas

  • General Chemistry
  • General Materials Science
  • General Biochemistry,Genetics and Molecular Biology
  • General Engineering
  • General Physics and Astronomy


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