Christoph Herold

Technische Universität Dresden, Dresden, Saxony, Germany

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Publications (12)77.98 Total impact

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    ABSTRACT: Hematopoietic stem and progenitor cell (HSPC) transplantation has become a routine clinical procedure to treat hematological malignancies as well as acquired bone marrow failure syndromes. The mechanical properties of cells have been emerging as label-free, inherent marker of biological function and disease. Here, we present first results for the application of cell mechanical properties as a marker to study the stem cell potential of HSPCs. The characterization of HSPCs in the transplanted graft relies currently on molecular phenotyping based on certain surface proteins, e.g. CD34 and CD133, initially characterized by flow cytometry (FACS). HSPCs were isolated from G-CSF mobilized peripheral blood and from bone marrow aspirates from hematological healthy donors using Miltenyi Microbeads for CD34 and CD133. In vitro differentiation and expansion of HSPCs was carried out for bone marrow derived CD34+ cells over 21 days. Mechanical characterization of freshly isolated, frozen/ defrozen, culture expanded and differentiated cells was carried out using real-time deformability cytometry (RT-DC) with a high throughput (100 cells/ s). RT-DC allows to study mechanical properties of high numbers of single cells in real-time. HSPCs were investigated under label-free conditions for their mechanical properties which demonstrated differences in the deformability of CD34+ cells from mobilized peripheral blood and from bone marrow aspirates. Moreover, we could distinguish different populations within the CD34+ cell fraction of the bone marrow. Interestingly, CD34+ and CD133+ cells display different mechanical distributions measured by RT-DC. In contrast, flow cytometric analysis with anti-CD34 and anti-CD133 antibodies did not reveal differences between both populations. Our data suggest that CD34+ cells are more deformable than CD133+ cells. We demonstrated that monocytes, macrophages and granulocytes, differentiated from HSPCs, show different deformation at similar stress conditions. Performing RT-DC on primary human HSPCs, we demonstrate the capability of our method to distinguish a mechanical phenotype of HSPCs from different resources, between different stem cell markers and to detect lineage specific mechanical phenotypes of differentiated cells.
    ISCCR, Stockholm; 06/2015
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    ABSTRACT: Hematopoietic stem and progenitor cell (HSPC) transplantation has become a routine clinical procedure to treat hematological malignancies as well as acquired bone marrow failure syndromes. The mechanical properties of cells have been emerging as label-free, inherent marker of biological function and disease. Here, we present first results for the application of cell mechanical properties as a marker to study the stem cell potential of HSPCs. The characterization of HSPCs in the transplanted graft relies currently on molecular phenotyping based on certain surface proteins, e.g. CD34 and CD133, initially characterized by flow cytometry (FACS). HSPCs were isolated from G-CSF mobilized peripheral blood and from bone marrow aspirates from hematological healthy donors using Miltenyi Microbeads for CD34 and CD133. In vitro differentiation and expansion of HSPCs was carried out for bone marrow derived CD34+ cells over 21 days. Mechanical characterization of freshly isolated, frozen/ defrozen, culture expanded and differentiated cells was carried out using real-time deformability cytometry (RT-DC) with a high throughput (100 cells/ s). RT-DC allows to study mechanical properties of high numbers of single cells in real-time. HSPCs were investigated under label-free conditions for their mechanical properties which demonstrated differences in the deformability of CD34+ cells from mobilized peripheral blood and from bone marrow aspirates. Moreover, we could distinguish different populations within the CD34+ cell fraction of the bone marrow. Interestingly, CD34+ and CD133+ cells display different mechanical distributions measured by RT-DC. In contrast, flow cytometric analysis with anti-CD34 and anti-CD133 antibodies did not reveal differences between both populations. Our data suggest that CD34+ cells are more deformable than CD133+ cells. We demonstrated that monocytes, macrophages and granulocytes, differentiated from HSPCs, show different deformation at similar stress conditions. Performing RT-DC on primary human HSPCs, we demonstrate the capability of our method to distinguish a mechanical phenotype of HSPCs from different resources, between different stem cell markers and to detect lineage specific mechanical phenotypes of differentiated cells.
    ISCCR, Stockholm; 06/2015
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    ABSTRACT: We introduce real-time deformability cytometry (RT-DC) for continuous cell mechanical characterization of large populations (>100,000 cells) with analysis rates greater than 100 cells/s. RT-DC is sensitive to cytoskeletal alterations and can distinguish cell-cycle phases, track stem cell differentiation into distinct lineages and identify cell populations in whole blood by their mechanical fingerprints. This technique adds a new marker-free dimension to flow cytometry with diverse applications in biology, biotechnology and medicine.
    Nature Methods 02/2015; 12(3). DOI:10.1038/nmeth.3281 · 25.95 Impact Factor
  • Biophysical Journal 01/2015; 108(2):140a. DOI:10.1016/j.bpj.2014.11.774 · 3.97 Impact Factor
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    ABSTRACT: Giant unilamellar vesicles (GUVs) represent a versatile in vitro system widely used to study properties of lipid membranes and their interaction with biomacromolecules and colloids. Electroformation with indium tin oxide (ITO) coated coverslips as electrodes is a standard approach to GUV production. In the case of cationic GUVs, however, application of this approach leads to notorious difficulties. We discover that this is related to aging of ITO-coated coverslips during their repeated use, which is reflected in their surface topography on the nanoscale. We find that mild annealing of the ITO-coated surface in air reverts the effects of aging and ensures efficient reproducible electroformation of supergiant (diameter > 100 μm) unilamellar vesicles containing cationic lipids.
    Langmuir 03/2012; 28(13):5518-21. DOI:10.1021/la3005807 · 4.46 Impact Factor
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    Christoph Herold · Cécile Leduc · Robert Stock · Stefan Diez · Petra Schwille
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    ABSTRACT: We report on a minimal system to mimic intracellular transport of membrane-bounded, vesicular cargo. In a cell-free assay, purified kinesin-1 motor proteins were directly anchored to the membrane of giant unilamellar vesicles, and their movement studied along two-dimensional microtubule networks. Motion-tracking of vesicles with diameters of 1-3 μm revealed traveling distances up to the millimeter range. The transport velocities were identical to velocities of cargo-free motors. Using total internal reflection fluorescence (TIRF) microscopy, we were able to estimate the number of GFP-labeled motors involved in the transport of a single vesicle. We found that the vesicles were transported by the cooperative activity of typically 5-10 motor molecules. The presented assay is expected to open up further applications in the field of synthetic biology, aiming at the in vitro reconstitution of sub-cellular multi-motor transport systems. It may also find applications in bionanotechnology, where the controlled long-range transport of artificial cargo is a promising means to advance current lab-on-a-chip systems.
    ChemPhysChem 03/2012; 13(4):1001-6. DOI:10.1002/cphc.201100669 · 3.36 Impact Factor
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    Biophysical Journal 01/2012; 102(3):221a. DOI:10.1016/j.bpj.2011.11.1213 · 3.97 Impact Factor
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    ABSTRACT: In Escherichia coli, the pole-to-pole oscillation of the Min proteins directs septum formation to midcell, which is required for symmetric cell division. In vitro, protein waves emerge from the self-organization of MinD, a membrane-binding ATPase, and its activator MinE. For wave propagation, the proteins need to cycle through states of collective membrane binding and unbinding. Although MinD presumably undergoes cooperative membrane attachment, it is unclear how synchronous detachment is coordinated. We used confocal and single-molecule microscopy to elucidate the order of events during Min wave propagation. We propose that protein detachment at the rear of the wave, and the formation of the E-ring, are accomplished by two complementary processes: first, local accumulation of MinE due to rapid rebinding, leading to dynamic instability; and second, a structural change induced by membrane-interaction of MinE in an equimolar MinD-MinE (MinDE) complex, which supports the robustness of pattern formation.
    Nature Structural & Molecular Biology 05/2011; 18(5):577-83. DOI:10.1038/nsmb.2037 · 13.31 Impact Factor
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    Christoph Herold · Petra Schwille · Eugene P. Petrov
    Biophysical Journal 02/2011; 100(3). DOI:10.1016/j.bpj.2010.12.2967 · 3.97 Impact Factor
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    Christoph Herold · Petra Schwille · Eugene P. Petrov
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    ABSTRACT: We describe a previously unreported coil-globule transition of DNA electrostatically bound to a freestanding fluid cationic lipid membrane. The collapse of a DNA coil into a compact globule takes place after the DNA molecule attaches in an extended conformation to the membrane. DNA condensation is favored at a higher cationic lipid content, while at lower membrane charge densities coexistence of DNA random coils, partially collapsed conformations, and globules is observed.
    Physical Review Letters 04/2010; 104(14):148102. DOI:10.1016/j.bpj.2010.12.514 · 7.51 Impact Factor
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    C Herold · P Schwille · E P Petrov
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    ABSTRACT: We describe a previously unreported coil-globule transition of DNA electrostatically bound to a freestanding fluid cationic lipid membrane. The collapse of a DNA coil into a compact globule takes place after the DNA molecule attaches in an extended conformation to the membrane. DNA condensation is favored at a higher cationic lipid content, while at lower membrane charge densities coexistence of DNA random coils, partially collapsed conformations, and globules is observed. Understanding the interaction of polyelectrolytes with oppositely charged lipid membranes is an important issue of soft matter physics, which provides an insight into mechanisms of interactions of biological macromolecules with cell membranes. Although the question has been addressed during the past decade both experimentally and theoretically [1,2], the understanding is far from com-plete, and some important unresolved questions, including the effects of the membrane local curvature and bending elasticity, remain to be addressed. A perfect model polymer to study electrostatic polyelectrolyte-membrane interactions is double-stranded DNA: it is a semiflexible polyelectrolyte carrying two negative charges per base pair (bp) [3] whose length and structure can be precisely controlled using the modern biotechnological methods; in addition, it allows for easy fluorescence labeling which facilitates single-molecule microscopy experiments. These advantages were used in a seminal work [4] where it was demonstrated that DNA molecules adsorbed at a fluid cationic lipid bilayer on a flat support assume a 2D random coil conformation and exercise translational Brownian motion. These results have since become a text-book example of polymer coil dynamics in 2D [5]. A completely different picture is observed when double-stranded DNA interacts with small (20–100 nm) cationic liposomes: In this case DNA molecules wrap around lipo-somes and eventually form densely packed liquid crystal-line DNA-lipid globules [6] with the typical size of $100–200 nm [7,8]. Formation of DNA-lipid globules is an example of a more general phenomenon known as DNA condensation [9,10]. DNA condensation by cationic lipo-somes has attracted particular attention in view of its potential use in gene therapy [11] and importance for understanding the prebiotic chemistry [8]. The striking contrast between the behavior of DNA at flat supported cationic lipid bilayers and at strongly curved small cationic liposomes naturally leads to the question of what kind of behavior can be expected upon interaction of DNA with freestanding (unsupported) cationic lipid bi-layers. The main differences between the supported and freestanding lipid bilayers are (i) the ability of the free-standing membrane to respond by an elastic deformation to an external mechanical force [12], which is strongly sup-pressed in the case of a supported lipid bilayer [13], and (ii) the high lipid mobility within the freestanding fluid lipid bilayer, which can be strongly inhibited by the solid support [14]. Recent experiments on interaction of DNA with cationic membranes supported on structured surfaces demonstrated the importance of the local bilayer curvature in DNA-membrane interactions [15]. Obviously, freestand-ing bilayers, capable of bending locally in response to an external perturbation, may show new unexpected ways of interaction with charged semiflexible DNA molecules. Surprisingly, very little is known about interaction of DNA with freestanding cationic lipid bilayers. To the best of the authors' knowledge, the only study in this direction was carried out in a series of works [16]. The experimental approach used in these works could not, however, provide any information on conformation and dynamics of single DNA molecules. In this Letter we describe a previously unreported phe-nomenon of coil-globule transition of DNA molecules electrostatically bound to a freestanding fluid cationic lipid membrane. To model the flat freestanding fluid cationic lipid bilayer we used giant unilamellar vesicles (GUVs) with sizes in the range of 100–300 m (Fig. 1). At these sizes, the free membrane surface is essentially flat on the micrometer scale and can be directly confronted to the flat supported membrane. Both GUVs and DNA molecules were fluores-cently labeled at distinct spectral ranges, which allowed us to carry out single-molecule fluorescence microscopy experiments. Double-stranded DNA fragments with lengths of 5, 10, and 20 kbp, as well as -DNA (48.5 kbp), were obtained from Fermentas Life Sciences. DNA samples were fluo-rescently stained using the YOYO-1 dye (Molecular Probes) at the ratio of 0:2 dye=bp (reducing the staining ratio to 0:05 dye=bp did not affect our results within ex-perimental uncertainty). Giant unilamellar vesicles with the membrane in the fluid phase were produced from
    Physical Review Letters 04/2010; 104(14):148102. DOI:10.1103/PhysRevLett.104.148102 · 7.51 Impact Factor
  • Biophysical Journal 02/2009; 96(3):131A. DOI:10.1016/j.bpj.2008.12.594 · 3.97 Impact Factor