Abstract
Immunofluorescence labeling enables the detection and characterization of various parasite proteins presented on the surface of the infected red blood cell. Several approaches for immunofluorescence detection of red blood cell surface-presented proteins of Plasmodium spp. have been successfully established and published over the years. However, finding the right approach depends on the scientific question, and different protocols have different advantages. Here, we discuss some aspects that should be considered and present an easily applicable protocol for labeling parasite surface antigens, which subsequently can be analyzed by immunofluorescence microscopy (or flow cytometry).
Key words
- Cytoadhesion
- Infected red blood cells
- Immunofluorescence
- Microscopy
- Fixation
- STED super-resolution microscopy
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References
Bacallao RS, S.; Phillips, C. (2006) Guiding principles of specimen preservation for confocal fluorescence microscopy. In: Pawley J (ed) Handbook of biological confocal microscopy. Springer, New York, NY
Tonkin CJ, van Dooren GG, Spurck TP, Struck NS, Good RT, Handman E, Cowman AF, McFadden GI (2004) Localization of organellar proteins in Plasmodium falciparum using a novel set of transfection vectors and a new immunofluorescence fixation method. Mol Biochem Parasitol 137(1):13–21. https://doi.org/10.1016/j.molbiopara.2004.05.009
Mehnert AK, Simon CS, Guizetti J (2019) Immunofluorescence staining protocol for STED nanoscopy of Plasmodium-infected red blood cells. Mol Biochem Parasitol 229:47–52. https://doi.org/10.1016/j.molbiopara.2019.02.007
Kilian N, Srismith S, Dittmer M, Ouermi D, Bisseye C, Simpore J, Cyrklaff M, Sanchez CP, Lanzer M (2015) Hemoglobin S and C affect protein export in Plasmodium falciparum-infected erythrocytes. Biol Open 4(3):400–410. https://doi.org/10.1242/bio.201410942
Richter KN, Revelo NH, Seitz KJ, Helm MS, Sarkar D, Saleeb RS, D'Este E, Eberle J, Wagner E, Vogl C, Lazaro DF, Richter F, Coy-Vergara J, Coceano G, Boyden ES, Duncan RR, Hell SW, Lauterbach MA, Lehnart SE, Moser T, Outeiro TF, Rehling P, Schwappach B, Testa I, Zapiec B, Rizzoli SO (2018) Glyoxal as an alternative fixative to formaldehyde in immunostaining and super-resolution microscopy. EMBO J 37(1):139–159. https://doi.org/10.15252/embj.201695709
Rug M, Maier AG (2013) Transfection of Plasmodium falciparum. Methods Mol Biol 923:75–98. https://doi.org/10.1007/978-1-62703-026-7_6
Kilian N, Dittmer M, Cyrklaff M, Ouermi D, Bisseye C, Simpore J, Frischknecht F, Sanchez CP, Lanzer M (2013) Haemoglobin S and C affect the motion of Maurer's clefts in Plasmodium falciparum-infected erythrocytes. Cell Microbiol 15(7):1111–1126. https://doi.org/10.1111/cmi.12102
Doritchamou JYA, Morrison R, Renn JP, Ribeiro J, Duan J, Fried M, Duffy PE (2019) Placental malaria vaccine candidate antigen VAR2CSA displays atypical domain architecture in some Plasmodium falciparum strains. Commun Biol 2:457. https://doi.org/10.1038/s42003-019-0704-z
Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, Preibisch S, Rueden C, Saalfeld S, Schmid B, Tinevez JY, White DJ, Hartenstein V, Eliceiri K, Tomancak P, Cardona A (2012) Fiji: an open-source platform for biological-image analysis. Nat Methods 9(7):676–682. https://doi.org/10.1038/nmeth.2019
Strohm EM, Berndl ES, Kolios MC (2013) Probing red blood cell morphology using high-frequency photoacoustics. Biophys J 105(1):59–67. https://doi.org/10.1016/j.bpj.2013.05.037
Kilian N, Goryaynov A, Lessard MD, Hooker G, Toomre D, Rothman JE, Bewersdorf J (2018) Assessing photodamage in live-cell STED microscopy. Nat Methods 15(10):755–756. https://doi.org/10.1038/s41592-018-0145-5
Fernandez P, Viebig NK, Dechavanne S, Lepolard C, Gysin J, Scherf A, Gamain B (2008) Var2CSA DBL6-epsilon domain expressed in HEK293 induces limited cross-reactive and blocking antibodies to CSA binding parasites. Malar J 7:170. https://doi.org/10.1186/1475-2875-7-170
Gruring C, Heiber A, Kruse F, Ungefehr J, Gilberger TW, Spielmann T (2011) Development and host cell modifications of Plasmodium falciparum blood stages in four dimensions. Nat Commun 2:165. https://doi.org/10.1038/ncomms1169
Quadt KA, Smyrnakou X, Frischknecht F, Bose G, Ganter M (2020) Plasmodium falciparum parasites exit the infected erythrocyte after haemolysis with saponin and streptolysin O. Parasitol Res 119(12):4297–4302. https://doi.org/10.1007/s00436-020-06932-9
Acknowledgments
We would like to acknowledge the Baden-Württemberg Stiftung for funding to M.G. and the German Research Foundation (DFG) for funding to J.G. (349355339). M.G. is member of the SFB 1129. The authors further express gratitude to Patrick Pitoniak for critically proofreading this manuscript.
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Ganter, M., Guizetti, J., Kilian, N. (2022). Visualization of Infected Red Blood Cell Surface Antigens by Fluorescence Microscopy. In: Jensen, A.T.R., Hviid, L. (eds) Malaria Immunology. Methods in Molecular Biology, vol 2470. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2189-9_31
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DOI: https://doi.org/10.1007/978-1-0716-2189-9_31
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