Specific Interaction Forces Research Articles

Single-Molecule-Level Quantification Based on Atomic Force Microscopy Data Reveals the Interaction between Melittin and Lipopolysaccharide in Gram-Negative Bacteria.

The interaction forces and mechanical properties of the interaction between melittin (Mel) and lipopolysaccharide (LPS) are considered to be crucial driving forces for Mel when killing Gram-negative bacteria (GNB). However, how their interaction forces perform at the single-molecule level and the dissociation kinetic characteristics of the Mel/LPS complex remain poorly understood. In this study, the single-molecule-level interaction forces between Mel and LPSs from E. coli K-12, O55:B5, O111:B4, and O128:B12 were explored using atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS). AFM-based dynamic force spectroscopy (DFS) and an advanced analytical model were employed to investigate the kinetic characteristics of the Mel/LPS complex dissociation. The results indicated that Mel could interact with both rough (R)-form LPS (E. coli K-12) and smooth (S)-form LPSs (E. coli O55:B5, O111:B4, and O128:B12). The S-form LPS showed a more robust interaction with Mel than the R-form LPS, and a slight difference existed in the interaction forces between Mel and the diverse S-form LPS. Mel interactions with the S-form LPSs showed greater specific and non-specific interaction forces than the R-form LPS (p < 0.05), as determined by AFM-based SMFS. However, there was no significant difference in the specific and non-specific interaction forces among the three samples of S-form LPSs (p > 0.05), indicating that the variability in the O-antigen did not affect the interaction between Mel and LPSs. The DFS result showed that the Mel/S-form LPS complexes had a lower dissociation rate constant, a shorter energy barrier width, a longer bond lifetime, and a higher energy barrier height, demonstrating that Mel interacted with S-form LPS to form more stable complexes. This research enhances the existing knowledge of the interaction micromechanics and kinetic characteristics of Mel and LPS at the single-molecule level. Our research may help with the design and evaluation of new anti-GNB drugs.

Adaptive hybrid-mode assist-as-needed control of upper limb exoskeleton for rehabilitation training

In this paper, a novel adaptive hybrid-mode assist-as-needed (AHMAAN) control algorithm is designed for the exoskeleton-assisted upper limb rehabilitation training. The overall control framework includes an outer control loop to calculate the required interaction force, and an inner control loop to drive the exoskeleton track subject’s motion, and provide specific target interaction force obtained from the outer control loop. In the outer control loop, a hybrid control mode is proposed, which consists of resistive mode and assistive mode. In this regard, a virtual tunnel is firstly established around the defined training task path, and the training mode is switched according to the deviation between the subject’s position and the boundary of the virtual tunnel. Furthermore, for tuning the strength of the resistance or assistance to the subjects with different motor capabilities, two adjustable gain factors are designed, whose values are adaptively adjusted according to the subject’s training performance by using a fuzzy logic. Then, for the inner control loop, a barrier Lyapunov function-based controller is designed to constrain exoskeleton tracking errors within the defined boundary. Meanwhile, time delay estimation (TDE) technology is used to estimate the uncertain terms of the system, and a robust adaption law is developed to compensate TDE error. Experimental tests have been performed on an upper limb exoskeleton with three healthy subjects to evaluate the effectiveness of the developed method. The results show that the proposed control scheme can be effectively applied in a variety of rehabilitation requirements and achieves better training performance than classical hybrid-mode assist-as-needed control method.

Dynamic Motion Primitives-Based Trajectory Learning for Physical Human–Robot Interaction Force Control

One promising function of interactive robots is to provide a specific interaction force to human users. For example, rehabilitation robots are expected to promote patients' recovery by interacting with them with a prescribed force. However, motion uncertainties of different individuals, which are hard to predict due to the varying motion speed and noises during motion, degrade the performance of existing control methods. This paper proposes a method to learn a desired reference trajectory for a robot based on dynamic motion primitives (DMPs) and iterative learning (IL). By controlling the robot to follow the generated desired reference trajectory, the interaction force can achieve a desired value. In our proposed approach, DMPs are first employed to parameterize the demonstration trajectories of the human user. Then a recursive least square (RLS)-based estimator is developed and combined with the Adam optimization method to update the trajectory parameters so that the desired reference trajectory of the robot is iteratively obtained by resolving the DMPs. Since the proposed method parameterizes the trajectories depending on the phrase variable, it removes the essential assumption of traditional IL methods where the iteration period should be invariant, and thus has improved robustness compared with the existing methods. Experiments are performed using an interactive robot to validate the effectiveness of our proposed scheme.

A chiral metal-organic framework synthesized by the mixture of chiral and non-chiral organic ligands for enantioseparation of drugs by open-tubular capillary electrochromatography

A chiral metal-organic framework L-Histidine-Zeolitic imidazolate framework-67 (L-His-ZIF-67) was synthesized by the mixture of chiral organic ligand L-histidine and non-chiral organic ligand 2-methylimidazole directly, and to the author's knowledge, the chiral L-His-ZIF-67 coated capillary column we prepared has still not been reported to date in the field of capillary electrophoresis. This chiral metal-organic frameworks material was used as the chiral stationary phase for enantioseparation of drugs by open-tubular capillary electrochromatography. The separation conditions such as pH value, buffer concentration and proportion of organic modifier were optimized. Under optimal conditions, the established enantioseparation system achieved good separation effect, and the resolution of five chiral drugs: esmolol (7.93), nefopam (3.03), salbutamol (2.42), scopolamine (1.08) and sotalol (0.81). In addition, the chiral recognition mechanism of L-His-ZIF-67 was elucidated by a series of mechanism experiments, and the specific interaction force was preliminarily speculated.

Single molecule localizations of voltage-gated sodium channel NaV1.5 on the surfaces of normal and cancer breast cells.

Voltage-gated sodium channels (VGSCs) are widely expressed in various types of tumor and cancer cells, and NaV1.5 is overexpressed in highly metastatic breast cancer cells. There may be positive relations between the expression levels of NaV1.5 and breast cancer recurrence and metastasis. Herein, NaV1.5 was detected and localized on the surfaces of normal and cancer breast cells by the single molecule recognition imaging (SMRI) mode of atomic force microscopy (AFM). The results reveal that NaV1.5 was irregularly distributed on the surfaces of normal and cancer breast cells. The NaV1.5 has an area percentage of 0.6% and 7.2% on normal and cancer breast cells, respectively, which indicates that there is more NaV1.5 on cancer cells than on normal cells. The specific interaction forces and binding kinetics in the NaV1.5-antibody complex system were investigated with the single molecule force spectroscopy (SMFS) mode of AFM, indicating that the stability of the NaV1.5-antibody on normal breast cells is higher than that on cancer breast cells. All these results will be useful to study the interactions of other ion channel-antibody systems, and will also be useful to understand the role of sodium channels in tumor metastasis and invasion.

Investigating the Effect of Tyrosine Kinase Inhibitors on the Interaction between Human Serum Albumin by Atomic Force Microscopy

It is important for elucidating the regulation mechanism of life activities, as well as for the prevention, diagnosis, and drug design of diseases, to study protein–protein interactions (PPIs). Here, we investigated the interactions of human serum albumin (HSA) in the presence of tyrosine kinase inhibitors (TKIs: imatinib, nilotinib, dasatinib, bosutinib, and ponatinib) using atomic force microscopy (AFM). The distribution of rupture events including the specific interaction force Fi and the non-specific interaction force F0 between HSA pairs was analyzed. Based on the force measurements, Fi and F0 between HSA pairs in the control experiment were calculated to be 47 ± 1.5 and 116.1 ± 1.3 pN. However, Fi was significantly decreased in TKIs, while F0 was slightly decreased. By measuring the rupture forces at various loading rates and according to the Bell equation, the kinetic parameters of the complexes were investigated in greater detail. Molecular docking was used as a complementary means by which to explore the force of this effect. The whole measurements indicated that TKIs influenced PPIs in a variety of ways, among which hydrogen bonding and hydrophobic interactions were the most important. In conclusion, these outcomes give us a better insight into the mechanisms of PPIs when there are exogenous compounds present as well as in different liquid environments.

Investigation of the interaction between MeCP2 methyl-CpG binding domain and methylated DNA by single molecule force spectroscopy

MeCP2 is an essential transcriptional repressor that mediates transcriptional inhibition by binding to methylated DNA. The binding specificity of MeCP2 protein to methylated DNA was considered to depend on its methyl-CpG binding domain (MBD). In this study, we used atomic force microscope based single-molecular force spectroscopy to investigate the interaction of MeCP2 MBD and methylated DNA. The specific interaction forces of the MeCP2 MBD-methylated DNA complexes were measured for the first time. The dynamics was also investigated by measuring the unbinding force of the complex at different loading rates. In addition, the distribution of unbinding forces and binding probabilities of MeCP2 MBD and different DNA were studied at the same loading rate. It was found that MeCP2 MBD had weak interaction with hemi-methylated and unmethylated DNA compared to methylated DNA. This work revealed the binding characteristics of MeCP2 MBD and methylated DNA at the single-molecule level. It provides a new idea for exploring the molecular mechanism of MeCP2 in regulating methylation signals.

Biophysical characterization of the adhesion between enveloped virus and phosphatidylserine-binding proteins

Abstract Enveloped viruses are viruses that have a lipid membrane. Many enveloped viruses, such as the emerging virus Ebola, are pathogenic and of great clinical importance. Recent studies have shown that the outer lipid leaflets of viral envelopes contain phosphatidylserine (PS), which serves as an important attachment factor for enveloped virus entry into host cells. The envelope PS interacts with cellular PS receptors in a manner that mimics apoptotic body-PS receptor interactions during the engulfment of dead cell debris by phagocytes. The PS receptors utilized by enveloped viruses for viral internalization are designated phosphatidylserine-mediated virus entry enhancing receptors (PVEERs). While the importance of PVEERs in mediating viral entry has been demonstrated, the biophysical properties of the PVEER-virus interactions, including the detailed kinetics of PVEER-virion-associated PS binding, have not been comprehensively evaluated and compared. Using Ebola virus (EBOV) as the model system, this study investigated and compared the interactions between model Ebola viral particles and PVEER family proteins that are known to mediate enveloped virus attachment and entry. We used single-molecule force spectroscopy to quantify the specific interaction forces between PS or model EBOV viral particles and PVEERs. Based on the dynamic force spectra of various PS-receptor interactions, our results suggest that PS-PVEER interactions are mechanically comparable to antibody-antigen interactions, and that Gas6 and MFG-E8 may generate more force-resistant intersections than TIM-1 and ProteinS. The findings can help explain the biophysical mechanisms underlying the PVEER-mediated entry of enveloped virus into cells.

Porous polymeric resin for adsorbing low concentration of VOCs: Unveiling adsorption mechanism and effect of VOCs’ molecular properties

The adsorption of 20 kinds of VOCs including n-alkanes, alcohols, esters, ketones, aromatics, chlorocarbons and cyclohexane on porous polymeric resin (PP-resin) were studied at low concentration by inverse gas chromatography (IGC). The adsorption partition coefficients and adsorption free energy of VOCs were calculated. A linear solvation energy relationship (LSER) model was employed to understand the adsorption mechanisms of VOCs on PP-resin. The results showed that hydroxyl groups on the surface of PP-resin resulted in the predominantly Lewis basicity on its surface. VOCs were adsorbed on the PP-resin mainly by dispersive interactions. Nevertheless, dipole-dipole/dipole-induced dipole interactions, hydrogen-bond acidity and hydrogen-bond basicity cannot be ignored especially for VOCs with polar groups. Among the adsorbates tested, methanol exhibited the highest specific interaction force (0.909) and cyclohexane was least only 0.038. It was also revealed that dispersive interactions were more important when longer carbon chain or larger molar volume of VOCs were adsorbed.

Rich lamellar crystal baklava-structured PZT/PVDF piezoelectric sensor toward individual table tennis training

Real-time monitoring of interaction between ball and racket can provide the extremely important information for high quality guidance of individual table tennis training. However, the existing visual monitoring system is hard to reflect the specific interaction force and accurate hit position. Here, we design a self-powered rich lamellar crystal baklava-structured PZT/PVDF piezoelectric sensor for real-time monitoring of table tennis training and the scalable sports. This unique baklava-structured piezocomposite could be massively manufactured by a facile two step method of nonsolvent induced phase separation and hot-press process. As a result, the as-developed sensor exhibits preeminent sensitivity (6.38 mV/N) and ultrafast response time (21 ms). On this basis, we made a smart table tennis racket embedded with 6 × 6 units, which could real-timely fetch the hit location and contact force, then it could provide evaluation and individual guidance for the athletes training. Admittedly, this work not only opens a new way for real-time monitoring of impact location and force in table tennis training beyond current visual detection system, but also paves a new prospect for artificial intelligence electronics toward smart sports.

Biomechanical characterization of TIM protein\u2013mediated Ebola virus\u2013host cell adhesion

Since the most recent outbreak, the Ebola virus (EBOV) epidemic remains one of the world’s public health and safety concerns. EBOV is a negative-sense RNA virus that can infect humans and non-human primates, and causes hemorrhagic fever. It has been proposed that the T-cell immunoglobulin and mucin domain (TIM) family proteins act as cell surface receptors for EBOV, and that the interaction between TIM and phosphatidylserine (PS) on the surface of EBOV mediates the EBOV–host cell attachment. Despite these initial findings, the biophysical properties of the TIM-EBOV interaction, such as the mechanical strength of the TIM-PS bond that allows the virus-cell interaction to resist external mechanical perturbations, have not yet been characterized. This study utilizes single-molecule force spectroscopy to quantify the specific interaction forces between TIM-1 or TIM-4 and the following binding partners: PS, EBOV virus-like particle, and EBOV glycoprotein/vesicular stomatitis virus pseudovirion. Depending on the loading rates, the unbinding forces between TIM and ligands ranged from 40 to 100 pN, suggesting that TIM-EBOV interactions are mechanically comparable to previously reported adhesion molecule–ligand interactions. The TIM-4–PS interaction is more resistant to mechanical force than the TIM-1–PS interaction. We have developed a simple model for virus–host cell interaction that is driven by its adhesion to cell surface receptors and resisted by membrane bending (or tension). Our model identifies critical dimensionless parameters representing the ratio of deformation and adhesion energies, showing how single-molecule adhesion measurements relate quantitatively to the mechanics of virus adhesion to the cell.

Quantitative spatial analysis of haematopoiesis-regulating stromal cells in the bone marrow microenvironment by 3D microscopy

Sinusoidal endothelial cells and mesenchymal CXCL12-abundant reticular cells are principal bone marrow stromal components, which critically modulate haematopoiesis at various levels, including haematopoietic stem cell maintenance. These stromal subsets are thought to be scarce and function via highly specific interactions in anatomically confined niches. Yet, knowledge on their abundance, global distribution and spatial associations remains limited. Using three-dimensional quantitative microscopy we show that sinusoidal endothelial and mesenchymal reticular subsets are remarkably more abundant than estimated by conventional flow cytometry. Moreover, both cell types assemble in topologically complex networks, associate to extracellular matrix and pervade marrow tissues. Through spatial statistical methods we challenge previous models and demonstrate that even in the absence of major specific interaction forces, virtually all tissue-resident cells are invariably in physical contact with, or close proximity to, mesenchymal reticular and sinusoidal endothelial cells. We further show that basic structural features of these stromal components are preserved during ageing.

Mapping the distribution of specific antibody interaction forces on individual red blood cells

Current blood typing methods rely on the agglutination of red blood cells (RBCs) to macroscopically indicate a positive result. An indirect agglutination mechanism is required when blood typing with IgG forms of antibodies. To date, the interaction forces between anti-IgG and IgG antibodies have been poorly quantified, and blood group related antigens have never been quantified with the atomic force microscope (AFM). Instead, the total intensity resulting from fluorescent-tagged antibodies adsorbed on RBC has been measured to calculate an average antigen density on a series of RBCs. In this study we mapped specific antibody interaction forces on the RBC surface. AFM cantilever tips functionalized with anti-IgG were used to probe RBCs incubated with specific IgG antibodies. This work provides unique insight into antibody-antigen interactions in their native cell-bound location, and crucially, on a per-cell basis rather than an ensemble average set of properties. Force profiles obtained from the AFM directly provide not only the anti-IgG – IgG antibody interaction force, but also the spatial distribution and density of antigens over a single cell. This new understanding might be translated into the development of very selective and quantitative interactions that underpin the action of drugs in the treatment of frontier illnesses.

Characterizing the effect of polymyxin B antibiotics to lipopolysaccharide on Escherichiacoli surface using atomic force microscopy.

Lipopolysaccharide (LPS) on gram-negative bacterial outer membranes is the first target for antimicrobial agents, due to their spatial proximity to outer environments of microorganisms. To develop antibacterial compounds with high specificity for LPS binding, the understanding of the molecular nature and their mode of recognition is of key importance. In this study, atomic force microscopy (AFM) and single molecular force spectroscopy were used to characterize the effects of antibiotic polymyxin B (PMB) to the bacterial membrane at the nanoscale. Isolated LPS layer and the intact bacterial membrane were examined with respect to morphological changes at different concentrations of PMB. Our results revealed that 3hours of 10μg/mL of PMB exposure caused the highest roughness changes on intact bacterial surfaces, arising from the direct binding of PMB to LPS on the bacterial membrane. Single molecular force spectroscopy was used to probe specific interaction forces between the isolated LPS layer and PMB coupled to the AFM tip. A short range interaction regime mediated by electrostatic forces was visible. Unbinding forces between isolated LPS and PMB were about 30pN at a retraction velocity of 500nm/s. We further investigated the effects of the polycationic peptide PMB on bacterial outer membranes and monitored its influences on the deterioration of the bacterial membrane structure. Polymyxin B binding led to rougher appearances and wrinkles on the outer membranes surface, which may finally lead to lethal membrane damage of bacteria. Our studies indicate the potential of AFM for applications in pathogen recognition and nano-resolution approaches in microbiology.

Direct mapping of chemical oxidation of individual graphene sheets through dynamic force measurements at the nanoscale.

Graphene oxide is one of the most studied nanomaterials owing to its huge application potential in many fields, including biomedicine, sensing, drug delivery, optical and optoelectronic technologies. However, a detailed description of the chemical composition and the extent of oxidation in graphene oxide remains a key challenge affecting its applicability and further development of new applications. Here, we report direct monitoring of the chemical oxidation of an individual graphene flake during ultraviolet/ozone treatment through in situ atomic force microscopy based on dynamic force mapping. The results showed that graphene oxidation expanded from the graphene edges to the entire graphene surface. The interaction force mapping results correlated well with X-ray photoelectron spectroscopy data quantifying the degree of chemical oxidation. Density functional theory calculations confirmed the specific interaction forces measured between a silicon tip and graphene oxide. The developed methodology can be used as a simple protocol for evaluating the chemical functionalization of other two-dimensional materials with covalently attached functional groups.

Probing fibronectin-antibody interactions using AFM force spectroscopy and lateral force microscopy.

The first experiment showing the effects of specific interaction forces using lateral force microscopy (LFM) was demonstrated for lectin–carbohydrate interactions some years ago. Such measurements are possible under the assumption that specific forces strongly dominate over the non-specific ones. However, obtaining quantitative results requires the complex and tedious calibration of a torsional force. Here, a new and relatively simple method for the calibration of the torsional force is presented. The proposed calibration method is validated through the measurement of the interaction forces between human fibronectin and its monoclonal antibody. The results obtained using LFM and AFM-based classical force spectroscopies showed similar unbinding forces recorded at similar loading rates. Our studies verify that the proposed lateral force calibration method can be applied to study single molecule interactions.

Deep eutectic solvents for the self-assembly of gold nanoparticles: a SAXS, UV-Vis, and TEM investigation.

In this work, we report the formation and growth mechanisms of gold nanoparticles (AuNPs) in eco-friendly deep eutectic solvents (DES; choline chloride and urea). AuNPs are synthesized on the DES surface via a low-energy sputter deposition method. Detailed small angle X-ray scattering (SAXS), UV-Vis, and cryogenic transmission electron microscopy (cryo-TEM) investigations show the formation of AuNPs of 5 nm diameter. Data analysis reveals that for a prolonged gold-sputtering time there is no change in the size of the particles. Only the concentration of AuNPs increases linearly in time. More surprisingly, the self-assembly of AuNPs into a first and second shell ordered system is observed directly by in situ SAXS for prolonged gold-sputtering times. The self-assembly mechanism is explained by the templating nature of DES combined with the equilibrium between specific physical interaction forces between the AuNPs. A disulfide-based stabilizer, bis((2-mercaptoethyl)trimethylammonium) disulfide dichloride, was applied to suppress the self-assembly. Moreover, the stabilizer even reverses the self-assembled or agglomerated AuNPs back to stable 5 nm individual particles as directly evidenced by UV-Vis. The template behavior of DES is compared to that of nontemplating solvent castor oil. Our results will also pave the way to understand and control the self-assembly of metallic and bimetallic nanoparticles.

Characterizing the Cell Surface Structure and Antibody Recognition Forces on Intact Microbial Cells using Scanning Probe Microscopy

Tannerella forsythia is among the most potent triggers of periodontal diseases and approaches to understand underlying mechanisms are currently intensively pursued. A ∼22‑nm thick, 2D crystallinesurface (S‑) layer that completely covers T. forsythia cells is crucially involved in the bacterium-host cross-talk. The S-layer is composed of two intercalating glycoproteins (TfsA‑GP, TfsB‑GP) that are aligned into a periodic lattice. To characterize this unique S‑layer structure at the nanometer-scale directly on intact T. forsythia cells, atomic force microscopy (AFM) based topography imaging and single-molecular force spectroscopy (SMFS) were applied. Direct high-resolution imaging of the outer surface of T. forsythia wild-type cells by AFM revealed a p4 structure with a lattice constant of ∼9.0 nm. In contrast, on S-layer mutant cells, no periodic lattice could be visualized. Additionally, SMFS was used to probe specific interaction forces between an anti-TfsA antibody coupled to the AFM tip and the S‑layer as present on T. forsythia wild-type and mutant cells displaying TfsA-GP alone. Unbinding forces between the antibody and wild-type cells were greater than with mutant cells. This indicated that the TfsA-GP is less strongly attached to the mutant cell surface when the co-assembling TfsB-GP is missing. Altogether, our data confirm the current model of the S‑layer architecture with two intercalating S-layer glycoproteins and TfsA-GP mainly outwardly orientated. This study was funded by a Marie Curie International Incoming Fellowship of the 7th European Community Framework Program (to Y. J. O.). Financial support came from the Austrian Science Fund FWF projects P20605-B12 and P24317-B22 (to C.S.).

Characterizing the S‐layer structure and anti‐S‐layer antibody recognition on intact Tannerella forsythia cells by scanning probe microscopy and small angle X‐ray scattering

Tannerella forsythia is among the most potent triggers of periodontal diseases, and approaches to understand underlying mechanisms are currently intensively pursued. A ~22-nm-thick, 2D crystalline surface (S-) layer that completely covers Tannerella forsythia cells is crucially involved in the bacterium-host cross-talk. The S-layer is composed of two intercalating glycoproteins (TfsA-GP, TfsB-GP) that are aligned into a periodic lattice. To characterize this unique S-layer structure at the nanometer scale directly on intact T. forsythia cells, three complementary methods, i.e., small-angle X-ray scattering (SAXS), atomic force microscopy (AFM), and single-molecular force spectroscopy (SMFS), were applied. SAXS served as a difference method using signals from wild-type and S-layer-deficient cells for data evaluation, revealing two possible models for the assembly of the glycoproteins. Direct high-resolution imaging of the outer surface of T. forsythia wild-type cells by AFM revealed a p4 structure with a lattice constant of ~9.0 nm. In contrast, on mutant cells, no periodic lattice could be visualized. Additionally, SMFS was used to probe specific interaction forces between an anti-TfsA antibody coupled to the AFM tip and the S-layer as present on T. forsythia wild-type and mutant cells, displaying TfsA-GP alone. Unbinding forces between the antibody and wild-type cells were greater than with mutant cells. This indicated that the TfsA-GP is not so strongly attached to the mutant cell surface when the co-assembling TfsB-GP is missing. Altogether, the data gained from SAXS, AFM, and SMFS confirm the current model of the S-layer architecture with two intercalating S-layer glycoproteins and TfsA-GP being mainly outwardly oriented.

Atomic force microscopy‐based force spectroscopy — biological and biomedical applications

The use of atomic force microscopy (AFM) applied to biological systems to generate high resolution images is gaining a wider acceptance. However, the most remarkable advances are being achieved on the use of the AFM to measure inter- and intramolecular interaction forces with piconewton resolution, not only to demonstrate this ability but also actually to solve biological and biomedical relevant questions. Single-molecule force spectroscopy recognition studies enable the detection of specific interaction forces, based on the AFM sensitivity and the possibility of manipulating individual molecules. In this review, we describe the basic principles of this methodology and some of the practical aspects involved. The ability to measure interactions at the single-molecule level is illustrated by some relevant examples. A special focus is given to the study of the fibrinogen-erythrocyte binding and its relevance as a cardiovascular risk factor. An approach to the latter problem by single-molecule force spectroscopy allowed the molecular recognition, characterization, and partial identification of a previously unknown receptor for fibrinogen on human erythrocytes.