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A Transgenic Approach to Live Imaging of Heparan Sulfate Modification Patterns

  • Matthew Attreed
  • Hannes E. BülowEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1229)

Abstract

Heparan sulfate (HS) glycosaminoglycan chains contain highly modified HS domains that are separated by sections of sparse or no modification. HS domains are central to the role of HS in protein binding and mediating protein–protein interactions in the extracellular matrix. Since HS domains are not genetically encoded, they are impossible to visualize and study with conventional methods in vivo. Here we describe a transgenic approach using previously described single chain variable fragment (scFv) antibodies that bind HS in vitro and on tissue sections with different specificities. By engineering a secretion signal and a fluorescent protein to the scFvs and transgenically expressing these fluorescently tagged antibodies in Caenorhabditis elegans, we are able to directly visualize specific HS domains in live animals (Attreed et al. Nat Methods 9(5):477–479, 2012). The approach allows concomitant colabeling of multiple epitopes, the study of HS dynamics and, could lend itself to a genetic analysis of HS domain biosynthesis or to visualize other nongenetically encoded or posttranslational modifications.

Key words

Heparan sulfate Single chain variable fragment (scFv) antibody, Caenorhabditis elegans Live imaging Nongenetically encoded molecules 

Notes

Acknowledgements

This work was supported by NIH grants F31NS076243 & T32GM07491 (M.A.) and RC1GM090825 & R01GM101313 (H.E.B.).

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of GeneticsAlbert Einstein College of Medicine of Yeshiva UniversityBronxUSA
  2. 2.Dominick P. Purpura Department of NeuroscienceAlbert Einstein College of Medicine of Yeshiva UniversityBronxUSA

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