Monitoring the uptake of glycosphingolipids in Plasmodium falciparum-infected erythrocytes using both fluorescence microscopy and capillary electrophoresis with laser-induced fluorescence detection

Anal Chem. 2010 Dec 1;82(23):9955-8. doi: 10.1021/ac1021776. Epub 2010 Nov 2.

Abstract

The metabolism of glycosphingolipids by the malaria-causing parasite Plasmodium falciparum plays an important role in the progression of the disease. We report a new and highly sensitive method to monitor the uptake of glycosphingolipids in infected red blood cells (iRBCs). A tetramethylrhodamine-labeled glycosphingolipid (GM1-TMR) was used as a substrate. Uptake was demonstrated by fluorescence microscopy. The iRBCs were lysed with a 15% solution of saponin and washed with phosphate buffered saline to release intact parasites. The parasites were further lysed and the resulting homogenates were analyzed by capillary electrophoresis with laser-induced fluorescence detection. The lysate from erythrocytes infected at 1% parasitemia generated a signal 20 standard deviations larger than uninfected erythrocytes, which suggests that relatively low infection levels can be studied with this technique.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Electrophoresis, Capillary / methods*
  • Erythrocytes / parasitology*
  • Fluorescent Dyes / chemistry
  • Glycosphingolipids / chemistry
  • Glycosphingolipids / metabolism*
  • Humans
  • Lasers*
  • Microscopy, Fluorescence / methods*
  • Plasmodium falciparum / metabolism*
  • Rhodamines / chemistry

Substances

  • Fluorescent Dyes
  • Glycosphingolipids
  • Rhodamines
  • tetramethylrhodamine