Studying the complex realm of cellular communication and interactions by fluorescence microscopy requires sample fixation on a transparent substrate. To activate T cells, which are pivotal in controlling the immune system, it is important to present the activating antigen in a spatial arrangement similar to the nature of the antigen-presenting cell, including the presence of ligands on microvilli. Similar arrangement is predicted for some other immune cells. In this work, immune cell-stimulating platform based on nanoparticle-ligand conjugates have been developed using a scalable, affordable, and broadly applicable technology, which can be readily deployed without the need for state-of-the-art nanofabrication instruments. The validation of surface biofunctionalization was performed by combination of fluorescence and atomic force microscopy techniques. We demonstrate that the targeted system serves as a biomimetic scaffold on which immune cells make primary contact with the microvilli-mimicking substrate and exhibit stimulus-specific activation.

Department of Biophysical Chemistry J Heyrovsky Institute of Physical Chemistry Czech Academy of Sciences Dolejškova 2155 3 182 23 Prague Czechia

Department of Organic Chemistry Faculty of Chemical Technology University of Chemistry and Technology Prague 166 28 Prague Czechia

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Delcassian D., Sattler S., Dunlop I.E. T Cell Immunoengineering with Advanced Biomaterials. Integr. Biol. 2017;9:211–222. doi: 10.1039/c6ib00233a. PubMed DOI PMC

Jin W., Tamzalit F., Chaudhuri P.K., Black C.T., Huse M., Kam L.C. T Cell Activation and Immune Synapse Organization Respond to the Microscale Mechanics of Structured Surfaces. Proc. Natl. Acad. Sci. USA. 2019;116:19835–19840. doi: 10.1073/pnas.1906986116. PubMed DOI PMC

Bashour K.T., Gondarenko A., Chen H., Shen K., Liu X., Huse M., Hone J.C., Kam L.C. CD28 and CD3 Have Complementary Roles in T-Cell Traction Forces. Proc. Natl. Acad. Sci. USA. 2014;111:2241–2246. doi: 10.1073/pnas.1315606111. PubMed DOI PMC

Deeg J., Axmann M., Matic J., Liapis A., Depoil D., Afrose J., Curado S., Dustin M.L., Spatz J.P. T Cell Activation Is Determined by the Number of Presented Antigens. Nano Lett. 2013;13:5619–5626. doi: 10.1021/nl403266t. PubMed DOI PMC

Sun Y., Sun J., Xiao M., Lai W., Li L., Fan C., Pei H. DNA Origami–Based Artificial Antigen-Presenting Cells for Adoptive T Cell Therapy. Sci. Adv. 2022;8 doi: 10.1126/sciadv.add1106. PubMed DOI PMC

Fisher P.J., Bulur P.A., Vuk-Pavlovic S., Prendergast F.G., Dietz A.B. Dendritic Cell Microvilli: A Novel Membrane Structure Associated with the Multifocal Synapse and T-Cell Clustering. Blood. 2008;112:5037–5045. doi: 10.1182/blood-2008-04-149526. PubMed DOI

Saltukoglu D., Özdemir B., Holtmannspötter M., Reski R., Piehler J., Kurre R., Reth M. Plasma Membrane Topography Governs the 3D Dynamic Localization of IgM B Cell Antigen Receptor Clusters. EMBO J. 2023;42 doi: 10.15252/embj.2022112030. PubMed DOI PMC

Frey T., Petty H.R., McConnell H.M. Electron Microscopic Study of Natural Killer Cell-Tumor Cell Conjugates. Proc. Natl. Acad. Sci. USA. 1982;79:5317–5321. doi: 10.1073/pnas.79.17.5317. PubMed DOI PMC

Kim H.-R., Jun C.-D. T Cell Microvilli: Sensors or Senders? Front. Immunol. 2019;10 PubMed PMC

Orbach R., Su X. Surfing on Membrane Waves: Microvilli, Curved Membranes, and Immune Signaling. Front. Immunol. 2020;11:2187. doi: 10.3389/fimmu.2020.02187. PubMed DOI PMC

Perica K., Kosmides A.K., Schneck J.P. Linking Form to Function: Biophysical Aspects of Artificial Antigen Presenting Cell Design. Biochim. Biophys. Acta. 2015;1853:781–790. doi: 10.1016/j.bbamcr.2014.09.001. PubMed DOI PMC

Luo X., Liu J. Ultrasmall Luminescent Metal Nanoparticles: Surface Engineering Strategies for Biological Targeting and Imaging. Adv. Sci. 2022;9 doi: 10.1002/advs.202103971. PubMed DOI PMC

Eggermont L.J., Paulis L.E., Tel J., Figdor C.G. Towards Efficient Cancer Immunotherapy: Advances in Developing Artificial Antigen-Presenting Cells. Trends Biotechnol. 2014;32:456–465. doi: 10.1016/j.tibtech.2014.06.007. PubMed DOI PMC

Samanta D., Sarkar A. Immobilization of Bio-Macromolecules on Self-Assembled Monolayers: Methods and Sensor Applications. Chem. Soc. Rev. 2011;40:2567–2592. doi: 10.1039/c0cs00056f. PubMed DOI

Wauters A.C., Scheerstra J.F., Vermeijlen I.G., Hammink R., Schluck M., Woythe L., Wu H., Albertazzi L., Figdor C.G., Tel J., et al. Artificial Antigen-Presenting Cell Topology Dictates T Cell Activation. ACS Nano. 2022;16:15072–15085. doi: 10.1021/acsnano.2c06211. PubMed DOI PMC

Fasting C., Schalley C.A., Weber M., Seitz O., Hecht S., Koksch B., Dernedde J., Graf C., Knapp E.-W., Haag R. Multivalency as a Chemical Organization and Action Principle. Angew. Chem., Int. Ed. Engl. 2012;51:10472–10498. doi: 10.1002/anie.201201114. PubMed DOI

Gao S., Guisán J.M., Rocha-Martin J. Oriented Immobilization of Antibodies onto Sensing Platforms - A Critical Review. Anal. Chim. Acta. 2022;1189 doi: 10.1016/j.aca.2021.338907. PubMed DOI

Demirel G., Çaykara T., Akaoğlu B., Çakmak M. Construction of a Novel Multilayer System and Its Use for Oriented Immobilization of Immunoglobulin G. Surf. Sci. 2007;601:4563–4570. doi: 10.1016/j.susc.2007.06.034. DOI

Miranda A., Martínez L., De Beule P.A.A. Facile synthesis of an aminopropylsilane layer on Si/SiO2 substrates using ethanol as APTES solvent. MethodsX. 2020;7 doi: 10.1016/j.mex.2020.100931. PubMed DOI PMC

Zhang D.K.Y., Cheung A.S., Mooney D.J. Activation and Expansion of Human T Cells Using Artificial Antigen-Presenting Cell Scaffolds. Nat. Protoc. 2020;15:773–798. doi: 10.1038/s41596-019-0249-0. PubMed DOI

Franke C., Chum T., Kvíčalová Z., Glatzová D., Gentsch G.J., Rodriguez A., Helmerich D.A., Herdly L., Mavila H., Frank O., et al. Approach to Map Nanotopography of Cell Surface Receptors. Commun. Biol. 2022;5:218. doi: 10.1038/s42003-022-03152-y. PubMed DOI PMC

Spicer C.D., Pashuck E.T., Stevens M.M. Achieving Controlled Biomolecule–Biomaterial Conjugation. Chem. Rev. 2018;118:7702–7743. doi: 10.1021/acs.chemrev.8b00253. PubMed DOI PMC

Williams E.H., Davydov A.V., Motayed A., Sundaresan S.G., Bocchini P., Richter L.J., Stan G., Steffens K., Zangmeister R., Schreifels J.A., Rao M.V. Immobilization of Streptavidin on 4H–SiC for Biosensor Development. Appl. Surf. Sci. 2012;258:6056–6063. doi: 10.1016/j.apsusc.2012.02.137. DOI

Weiss A., Wiskocil R.L., Stobo J.D. The Role of T3 Surface Molecules in the Activation of Human T Cells: A Two-Stimulus Requirement for IL 2 Production Reflects Events Occurring at a Pre-Translational Level. J. Immunol. 1984;133:123–128. PubMed

Helassa N., Zhang X.h., Conte I., Scaringi J., Esposito E., Bradley J., Carter T., Ogden D., Morad M., Török K. Fast-Response Calmodulin-Based Fluorescent Indicators Reveal Rapid Intracellular Calcium Dynamics. Sci. Rep. 2015;5 doi: 10.1038/srep15978. PubMed DOI PMC

Glatzová D., Mavila H., Saija M.C., Chum T., Cwiklik L., Brdička T., Cebecauer M. The Role of Prolines and Glycine in the Transmembrane Domain of LAT. FEBS J. 2021;288:4039–4052. doi: 10.1111/febs.15713. PubMed DOI

Kim H.R., Mun Y., Lee K.S., Park Y.J., Park J.S., Park J.H., Jeon B.N., Kim C.H., Jun Y., Hyun Y.M., et al. T Cell Microvilli Constitute Immunological Synaptosomes That Carry Messages to Antigen-Presenting Cells. Nat. Commun. 2018;9:3630. doi: 10.1038/s41467-018-06090-8. PubMed DOI PMC

Cai E., Marchuk K., Beemiller P., Beppler C., Rubashkin M.G., Weaver V.M., Gérard A., Liu T.L., Chen B.C., Betzig E., et al. Visualizing Dynamic Microvillar Search and Stabilization during Ligand Detection by T Cells. Science. 2017;356:eaal3118. doi: 10.1126/science.aal3118. PubMed DOI PMC

Barbieri L. University of Oxford; 2021. Quantitative Biophysical Methods to Study Immune Cell Mechanobiology.https://ora.ox.ac.uk/objects/uuid:55fde4e5-7d84-4b14-a040-20cb96a5d52d

Lee A.M., Colin-York H., Fritzsche M. CalQuo 2: Automated Fourier-space, population-level quantification of global intracellular calcium responses. Sci. Rep. 2017;7:5416. doi: 10.1038/s41598-017-05322-z. PubMed DOI PMC

Nečas D., Klapetek P.G. An Open-Source Software for SPM Data Analysis. Open Phys. 2012;10:181–188. doi: 10.2478/s11534-011-0096-2. DOI

Edelstein A.D., Tsuchida M.A., Amodaj N., Pinkard H., Vale R.D., Stuurman N. Advanced Methods of Microscope Control Using μManager Software. J. Biol. Methods. 2014;1:e10. doi: 10.14440/jbm.2014.36. PubMed DOI PMC

Schindelin J., Arganda-Carreras I., Frise E., Kaynig V., Longair M., Pietzsch T., Preibisch S., Rueden C., Saalfeld S., Schmid B., et al. Fiji: An Open-Source Platform for Biological-Image Analysis. Nat. Methods. 2012;9:676–682. doi: 10.1038/nmeth.2019. PubMed DOI PMC