Skip to main content

Visualization of Infected Red Blood Cell Surface Antigens by Fluorescence Microscopy

Part of the Methods in Molecular Biology book series (MIMB,volume 2470)


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

This is a preview of subscription content, access via your institution.

Buying options

USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
USD   149.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   199.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more


  1. 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

    Google Scholar 

  2. 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.

  3. 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.

  4. 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.

  5. 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.

    CrossRef  CAS  PubMed  Google Scholar 

  6. Rug M, Maier AG (2013) Transfection of Plasmodium falciparum. Methods Mol Biol 923:75–98.

  7. 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.

  8. 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.

  9. 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.

    CrossRef  CAS  PubMed  Google Scholar 

  10. Strohm EM, Berndl ES, Kolios MC (2013) Probing red blood cell morphology using high-frequency photoacoustics. Biophys J 105(1):59–67.

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  11. 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.

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  12. 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.

    CrossRef  CAS  PubMed  PubMed Central  Google Scholar 

  13. 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.

  14. 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.

    CrossRef  PubMed  Google Scholar 

Download references


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.

Author information

Authors and Affiliations


Corresponding author

Correspondence to Markus Ganter .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and Permissions

Copyright information

© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

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.

Download citation

  • DOI:

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2188-2

  • Online ISBN: 978-1-0716-2189-9

  • eBook Packages: Springer Protocols