Research Papers: Imaging

Recovering fluorophore concentration profiles from confocal images near lateral refractive index step changes

[+] Author Affiliations
Eleonóra Parelius Jonášová, Astrid Bjørkøy, Bjørn Torger Stokke

Norwegian University of Science and Technology (NTNU), Section for Biophysics and Medical Technology, Department of Physics, Høgskoleringen 5, Trondheim 7491, Norway

J. Biomed. Opt. 21(12), 126014 (Dec 21, 2016). doi:10.1117/1.JBO.21.12.126014
History: Received September 16, 2016; Accepted November 28, 2016
Text Size: A A A

Abstract.  Optical aberrations due to refractive index mismatches occur in various types of microscopy due to refractive differences between the sample and the immersion fluid or within the sample. We study the effects of lateral refractive index differences by fluorescence confocal laser scanning microscopy due to glass or polydimethylsiloxane cuboids and glass cylinders immersed in aqueous fluorescent solution, thereby mimicking realistic imaging situations in the proximity of these materials. The reduction in fluorescence intensity near the embedded objects was found to depend on the geometry and the refractive index difference between the object and the surrounding solution. The observed fluorescence intensity gradients do not reflect the fluorophore concentration in the solution. It is suggested to apply a Gaussian fit or smoothing to the observed fluorescence intensity gradient and use this as a basis to recover the fluorophore concentration in the proximity of the refractive index step change. The method requires that the reference and sample objects have the same geometry and refractive index. The best results were obtained when the sample objects were also used for reference since small differences such as uneven surfaces will result in a different extent of aberration.

Figures in this Article
© 2016 Society of Photo-Optical Instrumentation Engineers

Citation

Eleonóra Parelius Jonášová ; Astrid Bjørkøy and Bjørn Torger Stokke
"Recovering fluorophore concentration profiles from confocal images near lateral refractive index step changes", J. Biomed. Opt. 21(12), 126014 (Dec 21, 2016). ; http://dx.doi.org/10.1117/1.JBO.21.12.126014


Tables

Access This Article
Sign in or Create a personal account to Buy this article ($20 for members, $25 for non-members).

Some tools below are only available to our subscribers or users with an online account.

Related Content

Customize your page view by dragging & repositioning the boxes below.

Related Book Chapters

Topic Collections

Advertisement
  • Don't have an account?
  • Subscribe to the SPIE Digital Library
  • Create a FREE account to sign up for Digital Library content alerts and gain access to institutional subscriptions remotely.
Access This Article
Sign in or Create a personal account to Buy this article ($20 for members, $25 for non-members).
Access This Proceeding
Sign in or Create a personal account to Buy this article ($15 for members, $18 for non-members).
Access This Chapter

Access to SPIE eBooks is limited to subscribing institutions and is not available as part of a personal subscription. Print or electronic versions of individual SPIE books may be purchased via SPIE.org.