Distinct patterns of cerebral extravasation by Evans blue and sodium fluorescein in rats

PLoS One. 2013 Jul 5;8(7):e68595. doi: 10.1371/journal.pone.0068595. Print 2013.


The Evans blue dye (EBD; 961 Da) and the sodium fluorescein dye (NaF; 376 Da) are commonly used inert tracers in blood-brain barrier (BBB) research. They are both highly charged low molecular weight (LMW) tracers with similar lipophobic profiles. Nevertheless, the EBD binds to serum albumin (69,000 Da) to become a high molecular weight (HMW) protein tracer when injected into the circulation, whereas the NaF remains an unbound small molecule in the circulation. In this study, rats were injected with equal doses of either EBD or NaF to monitor their blood and tissue distribution. The EBD was largely confined to the circulation with little accumulation in the peripheral organ and even less accumulation in the central tissue, whereas the NaF distributed more evenly between the blood and the peripheral organ but was also largely excluded from the central tissue. Importantly, the EBD crossed the BBB most effectively at the prefrontal cortex and the cerebellum, and most poorly at the striatum. In marked contrast, the NaF was evenly distributed throughout the brain. Finally, the EBD exhibited this same peculiar tissue distribution profile when administered by either bolus injection or slow infusion. Our study suggests that different regions of the brain are equally permeable to LMW inert dyes like the NaF, but are markedly different in permeability to HMW proteins such as EBD-labelled serum albumin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blood-Brain Barrier / metabolism
  • Capillary Permeability
  • Cerebrum / metabolism*
  • Evans Blue / administration & dosage
  • Evans Blue / metabolism*
  • Extravasation of Diagnostic and Therapeutic Materials*
  • Fluorescein / administration & dosage
  • Fluorescein / metabolism*
  • Male
  • Rats


  • Evans Blue
  • Fluorescein

Grant support

This work was supported by research grants from the China Medical University Hospital (DMR-101-120), the Taiwan Department of Health Clinical Trial and Research Center of Excellence (DOH102-TD-B-111-004), and the National Research Council of Taiwan (NSC100-2632-B-039-001-MY3; NSC101-2321-B-039-008). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.