Immunofluorescence staining protocol for STED nanoscopy of Plasmodium-infected red blood cells

• Verfasst von: Mehnert, Ann-Kathrin [VerfasserIn]
• Verfasst von: Simon, Caroline [VerfasserIn]
• Verfasst von: Guizetti, Julien [VerfasserIn]
• Titel: Immunofluorescence staining protocol for STED nanoscopy of Plasmodium-infected red blood cells
• Verf.angabe: Ann-Kathrin Mehnert, Caroline Sophie Simon, Julien Guizetti
• E-Jahr: 2019
• Jahr: 01 March 2019
• Umfang: 6 S.
• Fussnoten: Gesehen am 21.05.2019
• Titel Quelle: Enthalten in: Molecular and biochemical parasitology
• Ort Quelle: Amsterdam : Elsevier, 1980
• Jahr Quelle: 2019
• Band/Heft Quelle: 229(2019), Seite 47-52
• ISSN Quelle: 1872-9428
• DOI: doi:10.1016/j.molbiopara.2019.02.007
• Datenträger: Online-Ressource
• Sprache: eng
• Sach-SW: Blood stage
• Sach-SW: Immunofluorescence
• Sach-SW: Malaria
• Sach-SW: STED
• Sach-SW: Super-resolution
• K10plus-PPN: 1666008311

Abstract

Immunofluorescence staining is the key technique for visualizing organization of endogenous cellular structures in single cells. Labeling and imaging of blood stage Plasmodium falciparum has always been challenging since it is a small intracellular parasite. A widely-used standard for parasite immunofluorescence is fixation in suspension with addition of minute amounts of glutaraldehyde to the paraformaldehyde-based solution. While this maintains red blood cell integrity, it has been postulated that antigenicity of the parasite proteins was, if at all, only slightly reduced. Here we show the deleterious effect that even these small quantities of glutaraldehyde can have on immunofluorescence staining quality and present an alternative cell seeding protocol that allows fixation with only paraformaldehyde. The highly improved signal intensity and staining efficiency enabled us to carry out RescueSTED nanoscopy on microtubules and nuclear pores and describe their organization in greater detail throughout the blood stage cycle.