Proteome analysis of human serum proteins adsorbed onto different titanium surfaces used in dental implants

Titanium dental implants are commonly used due to their biocompatibility and biochemical properties; blasted acid-etched Ti is used more frequently than smooth Ti surfaces. In this study, physico-chemical characterisation revealed important differences in roughness, chemical composition and hydrophilicity, but no differences were found in cellular in vitro studies (proliferation and mineralization). However, the deposition of proteins onto the implant surface might affect in vivo osseointegration. To test that hypothesis, protein layers formed on discs of both surface type after incubation with human serum were analysed. Using mass spectrometry (LC/MS/MS), 218 proteins were identified, 30 of which were associated with bone metabolism. Interestingly, Apo E, antithrombin and protein C adsorbed mostly onto blasted and acid-etched Ti, whereas the proteins of the complement system (C3) were found predominantly on smooth Ti surfaces. These results suggest that physico-chemical characteristics could be responsible for the differences observed in the adsorbed protein layer.     

Human palatal saliva: Adsorption behaviour and the role of low-molecular weight proteins

In situ ellipsometry was employed to study adsorption from human palatal saliva (HPalS) in terms of dependence on surface wettability and saliva concentration ( ? 1%). Adsorbed amounts, kinetics, and elutability with buffer and sodium dodecyl sulphate (SDS) were determined. The low-molecular weight protein content of bulk HPalS was also investigated using two-dimensional gel electrophoresis, and this revealed the presence of a large group of proteins < 100 kDa in size. Adsorption to pure (hydrophilic) and methylated (hydrophobized) silica surfaces revealed that the total adsorbed amounts were greater on hydrophobized silica. Below concentrations of 0.5 and 0.25% saliva, adsorption was concentration dependent on hydrophobized and hydrophilic surfaces, respectively. The initial adsorption ( ? 30 min) was faster on hydrophobized surfaces. Addition of SDS removed more material than buffer rinsing on both surfaces. Analysis of the adsorption kinetics indicated that the presence of low-molecular weight proteins plays a role in adsorption from HPalS.     

Adsorption of HSA,IgG and laminin-1 on model titania surfaces – effects of glow discharge treatment on competitively adsorbed film composition

This study investigated the effect of glow discharge treatment of titania surfaces on plasma protein adsorption, by means of ellipsometry and mechanically assisted SDS elution. The adsorption and film elution of three plasma proteins, viz. human serum albumin (HSA), human immunoglobulin G (IgG) and laminin-1, as well as competitive adsorption from a mixture of the three proteins, showed that the adsorbed amount of the individual proteins after 1 h increased in the order HSA <IgG <laminin-1 ≤ protein mixture. Film elutability showed that 30 min of SDS interaction resulted in almost complete removal of adsorbed films. No difference in the total adsorbed amounts of individual proteins, or from the mixture, was observed between untreated and glow discharge treated titania surfaces. However, the composition of the adsorbed films from the mixture differed between the untreated and glow discharge treated substrata. On glow discharge-treated titania the fraction of HSA increased, the fraction of laminin-1 decreased and the fraction of IgG was unchanged compared to the adsorption on the untreated titania, which was attributed to protein–protein interactions and competitive/associative adsorption behaviour.     

Nanometer Scale Titanium Surface Texturing Are Detected by Signaling Pathways Involving Transient FAK and Src Activations

Background

It is known that physico/chemical alterations on biomaterial surfaces have the capability to modulate cellular behavior, affecting early tissue repair. Such surface modifications are aimed to improve early healing response and, clinically, offer the possibility to shorten the time from implant placement to functional loading. Since FAK and Src are intracellular proteins able to predict the quality of osteoblast adhesion, this study evaluated the osteoblast behavior in response to nanometer scale titanium surface texturing by monitoring FAK and Src phosphorylations.

Methodology

Four engineered titanium surfaces were used for the study: machined (M), dual acid-etched (DAA), resorbable media microblasted and acid-etched (MBAA), and acid-etch microblasted (AAMB). Surfaces were characterized by scanning electron microscopy, interferometry, atomic force microscopy, x-ray photoelectron spectroscopy and energy dispersive X-ray spectroscopy. Thereafter, those 4 samples were used to evaluate their cytotoxicity and interference on FAK and Src phosphorylations. Both Src and FAK were investigated by using specific antibody against specific phosphorylation sites.

Principal Findings

The results showed that both FAK and Src activations were differently modulated as a function of titanium surfaces physico/chemical configuration and protein adsorption.

Conclusions

It can be suggested that signaling pathways involving both FAK and Src could provide biomarkers to predict osteoblast adhesion onto different surfaces.     

Cells,growth factors and bioactive surface properties in a mechanobiological model of implant healing

Interface conditions are of prime importance for implant fixation in the early post-operative period and modelling of specific biochemical interactions at implant surface is still missing. We hypothesized that updating osteoblast adhesion properties and growth factor source in an active zone located at the implant surface was relevant to model biochemical interactions of implant with its environment. We proposed an innovative set of diffusive–convective–reactive equations which relevant parameters were the cell decay factor, the cell motility and the growth factor balance.Initial comparison with histomorphometic results from a stable PMMA canine implant model provided an encouraging base to implement a numerical sensitivity analysis to evaluate the role of three types of bioactive surfaces: acid-etched titanium, coarse grit-blasted acid-etched titanium and coarse grit-blasted acid-etched titanium with RGDS peptide. We found that cell diffusion decrease (acid-etched+RGDS peptide vs. PMMA), and increase of local growth factor fraction (PMMA vs. acid-etched+RGDS peptide), significantly improved the amount of mineralized tissue on the implant surface. When the variation of structural fraction to cell motility and growth factor synthesis was investigated, an envelope pattern with an optimum was obtained but this could be exceeded for strong surface modifications and/or for high growth factor concentrations. The model also confirmed that implant bioactive properties should play a limited role to reduce heterogeneity of new-formed tissue. In conclusion, we suggested that our innovative theoretical approach was relevant to investigate implant fixation and could potentially help in reduction of implant revision.     

Effects of Chain Length and Saturability of Fatty Acids on Phospholipids and Proteins in Plasma Membranes of Bovine Mammary Gland

Free fatty acids (FFAs) in plasma are essential substrates for de novo synthesis of milk fat, or directly import into mammary cells. The physico-chemical properties of mammary cells membrane composition affected by FFAs with different chain lengths and saturability are unclear yet. Employing GC, FTIR and fluorescence spectroscopy, the adsorption capacity, phospholipids content, membrane proteins conformation, lipid peroxidation product, and free sulfhydryl of plasma membranes (PMs) interacted with different FFAs were determined. The mammary cells PMs at 38 and 39.5 °C showed different adsorption capacities: acetic acid (Ac) > stearic acid (SA) > β-hydroxybutyric acid (BHBA) > trans10, cis12 CLA. In the FTIR spectrum, the major adsorption peaks appeared at 2920 and 2850 cm^?1 for phospholipids, and at 1628 and 1560 cm^?1 for membrane proteins. The intensities of PMs-FFAs complexes were varied with the FFAs species and their initial concentrations. The β-sheet and turn structures of membrane proteins were transferred into random coil and α-helix after BHBA, SA and trans10, cis12 CLA treatments compared with Ac treatment. The quenching effects on the fluorescence of endogenous membrane protein, 1, 8-ANS, NBD-PE, and DHPE entrapped in PMs by LCFA were different from those of short chain FFAs. These results indicate that the adsorption of FFAs could change membrane protein conformation and polarity of head group in phospholipids. This variation of the mammary cells PMs was regulated by carbon chain length and saturability of FFAs.     

Extra tyrosine in the carbohydrate-binding module of Irpex lacteus Xyn10B enhances its cellulose-binding ability

The xylanase (Xyn10B) that strongly adsorbs on microcrystalline cellulose was isolated from Driselase. The Xyn10B contains a Carbohydrate-binding module family 1 (CBM1) (IrpCBM_Xyn10B) at N-terminus. The canonical essential aromatic residues required for cellulose binding were conserved in IrpCBM_Xyn10B; however, its adsorption ability was markedly higher than that typically observed for the CBM1 of an endoglucanase from Trametes hirsuta (ThCBM_EG1). An analysis of the CBM-GFP fusion proteins revealed that the binding capacity to cellulose (7.8 μmol/g) and distribution coefficient (2.0 L/μmol) of IrpCBM_Xyn10B-GFP were twofold higher than those of ThCBM_EG1-GFP (3.4 μmol/g and 1.2 L/μmol, respectively), used as a reference structure. Besides the canonical aromatic residues (W24-Y50-Y51) of typical CBM1-containing proteins, IrpCBM_Xyn10B had an additional aromatic residue (Y52). The mutation of Y52 to Ser (IrpCBM_Y52S-GFP) reduced these adsorption parameters to 4.4 μmol/g and 1.5 L/μmol, which were similar to those of ThCBM_EG1-GFP. These results indicate that Y52 plays a crucial role in strong cellulose binding.     

Inhibition de l'adhérence de Porphyromonas gingivalis sur la surface de titane greffé de poly(styrène sulfonate de sodium)

Dental implant-associated infections as peri-implantitis represent one of the major causes of osteointegration failures of oral implants. Adhesion of Porphyromonas gingivalis, one of the bacterial strains mainly involved in such infections, is tightly dependent on the topographical and/or physico-chemical properties of the implant surfaces. As a matter of fact, we showed that the grafting of one bioactive polymer such as poly(sodium styrene sulfonate) onto titanium implant surfaces allowed a sensitive decrease of Staphylococcus aureus adhesion (> 40%). The aim of the study consists in evaluating the adhesion of P. gingivalis onto titanium surfaces grafted with poly(sodium stryrene sulfonate) in order to elaborate implants exhibiting appropriate inhibiting properties towards the adhesion of periodontal pathogens. The grafting of poly(sodium stryrene sulfonate) onto titanium surfaces is carried out in two steps: chemical oxydation of titanium to initiate radical species then grafting of poly(sodium stryrene sulfonate) by radical polymerization. Chemical characterization of the surfaces is achieved by Fourier transformed infrared spectroscopy (FTIR). Bacterial adhesion was studied on grafted and non grafted (control) titanium surfaces, preadsorbed or not by plasmatic proteins. Protein adsorption as well as bacteria adhesion is followed by fluorescence spectroscopy by using proteins or bacteria previously labelled with fluorescence probes; the quantification of adsorption and bacteria adhesion are performed by image analysis. Results showed that protein adsorption is more important (~3 times) and that P. gingivalis adhesion is strongly inhibited (~73%) onto poly(sodium styrene sulfonate) grafted surfaces when compared to titanium control. Moreover, the inhibition of bacterial adhesion on grafted surfaces preadsorbed with plasma proteins is comparable to that observed on grafted surfaces preadsorbed with fibronectin. In conclusion, the obtained results evidenced that the grafting of titanium surface by poly(sodium styrene sulfonate) led to significant inhibition of P. gingivalis adhesion and that this inhibitory activity involved adsorbed proteins. Poly(sodium styrene sulfonate) grafted titanium surfaces present a high interest for the elaboration of oral implants in various clinical dental applications.     

Effect of adsorbed fibronectin on the differential adhesion of osteoblast-like cells and Staphylococcus aureus with and without fibronectin-binding proteins

The influence of fibronectin (Fn) coated surfaces patterned with poly(ethylene glycol) microgels having inter-gel spacings between 0.5 and 3.0 μm on the adhesion of Staphylococcus aureus strains with and without Fn-binding proteins and cellular adhesion/spreading was investigated. Quantitative force measurements between a S. aureus cell and a patterned surface showed that the adhesion force between the bacterium and the patterned surface increased substantially after Fn adsorption, regardless of the strain used, but decreased with decreasing inter-gel spacing. In flow-chamber experiments, the Fn-binding strain adhered at a higher rate after Fn adsorption than the strain lacking Fn-binding proteins. In both cases, the adhesion rates decreased with decreasing inter-gel spacing. Osteoblast-like cells could bind to patterned surfaces despite the microgels, and adsorbed Fn substantially amplified this effect. Even under highly non-adhesive conditions associated with closely spaced microgels, adsorbed Fn preserves a window of inter-gel spacing around 1 μm where the adhesion of staphylococcal cells is hindered while cells can still adhere and spread.     

OPTIMIZATION OF AFFINITY DIGESTION FOR THE ISOLATION OF CELLULOSOMES FROM Clostridium thermocellum

The affinity digestion process for cellulase purification consisting of binding to amorphous cellulose, and amorphous cellulose hydrolysis in the presence of dialysis (Morag et al., 1991), was optimized to obtain high activity recoveries and consistent protein recoveries in the isolation of Clostridium thermocellum cellulase. Experiments were conducted using crude supernatant prepared from C. thermocellum grown on either Avicel or cellobiose. While no difference was observed between Avicel-grown or cellobiose-grown cellulase in the adsorption step, differences were observed during the hydrolysis step. The optimal amorphous cellulose loading was found to be 3 mg amorphous cellulose per milligram supernatant protein. At this loading, 90–100% of activity in the crude supernatant was adsorbed. Twenty-four-hour incubation with the amorphous cellulose during the adsorption stage was found to result in maximal and stable adsorption of activity to the substrate. By fitting the adsorption data to the Langmuir model, an adsorption constant of 410 L/g and a binding capacity of 0.249 g cellulase/g cellulose were obtained. The optimal length of time for hydrolysis was found to be 3 hr for cellulase purified from Avicel cultures and 4 hr for cellulase purified from cellobiose cultures. These loadings and incubation times allowed for more than 85% activity recovery.     

The Biodegradation of Zein <Emphasis Type="Italic">In Vitro</Emphasis> and <Emphasis Type="Italic">In Vivo</Emphasis> and its Application in Implants

A unique polymer-based sustained-release implant drug delivery system was prepared by using biocompatible and biodegradable Zein as the skeleton meterial. After preparing Zein colloids, the Zein-loaded implant rods were formulated by injection molding followed by evaporating the solvent, and being coated with poly(lactic-co-glycolic) acid (PLGA) solution. Drug release kinetics was examined by using Fluorouracil (5-FU) as model drug. Nearly zero-order release was achieved for the model drugs for a period of 0–25 days when the implants were incubated in distilled water at 37°C. And then the degradation kinetics of the rods in vivo and in vitro were evaluated, which indicated that Zein could be absorbed by body and has good degradation property. The effects of different ratios of Zein/5-FU and the rods’ diameter on drug release were studied, respectively. The plasma concentration of 5-FU in the implants were determined by HPLC after implanting a single dose of the implants in rats. All data were subsequently processed by using the computer program 3P97, and the values were showed as follows: the area under the plasma concentration–time curve (AUC) value was 321.88 (μg/ml) × day, and the mean residence time (MRT) value was 23.05 days. The sustained-release implants of Zein/5-FU were successfully formulated. The uniqueness of the article is that Zein has been used as a skeleton material in implant delivery system for the first time and zero-order release kinetics has been obtained successfully.     

Laser scanning microscopy study on adsorption of biologically relevant proteins on implant materials

The adsorption of proteins at implant surfaces plays a key role in osseointegration and is therefore of great importance in biomaterial science. Laser scanning microscopy (LSM) is described, a method that is used here for the first study of the adsorption of proteins on implant surfaces. These LSM measurements provide information on the surface morphology, and the spatial distribution of adsorbed proteins can be deduced.     

Can the modeling for simplification of a dental implant surface affect the accuracy of 3D finite element analysis?

The aim of this study was to assess stress/strain of different implant modeling simplifications by 3D-FEA. Three variation of external hexagon implant (Ø3.75?×?10 mm) supporting one molar crown were simulated: A (no threads); B (slightly threads simplification); C (original design). 200 N (axial) and 100 N (oblique) were applied. Cortical bone was evaluated by maximum principal stress and microstrain qualitatively and quantitatively (ANOVA and Tukey post hoc (p < 0.05)). Higher stress levels (p < 0.05) were observed in model A. Models B and C presented similar stress transmission. It was possible to conclude that slightly simplification should be used for studies evaluating stress transferring for bone tissue.     

Botulinum Toxins Inhibit the Antidiuretic Hormone (ADH)-Stimulated Increase in Rabbit Cortical Collecting-Tubule Water Permeability

The mammalian renal collecting duct increases its water permeability in response to antidiuretic hormone (ADH). ADH causes cytoplasmic endosomes containing the water channel, aquaporin 2 (AQP2), to fuse with the apical membrane so that the water permeability of the tubule increases many times above baseline. SNARE proteins are involved in the docking and fusion of vesicles with the cell membrane in neuron synapses. Whether these proteins are involved in the fusion of vesicles to the cell membrane in other tissues is not entirely clear. In the present study, we examined the role of SNARE proteins in the insertion of water channels in the collecting-duct response to ADH by using botulinum toxins A, B and C. Toxins isolated from clostridium botulinum are specific proteases that cleave different SNARE proteins and inactivate them. Tubules were perfused in vitro with botulinum toxin in the perfusate (50 nM for A and B and 15 nM for C). ADH (200 pM) was then added to the bath after baseline measurements of osmotic water permeability (P_f) and the change in P_f was followed for one hour. Botulinum toxins significantly inhibited the maximum P_f by approximately 50%. Botulinum toxins A and C also decreased the rate of rise of P_f. Thus, SNARE proteins are involved in the insertion of the water channels in the collecting duct.     

Conditioning of self-assembled monolayers at two static immersion test sites along the east coast of Florida and its effect on early fouling development

Among the first events after immersion of surfaces in the ocean is surface ‘conditioning’. Here, the accumulation and composition of the conditioning films formed after immersion in the ocean are analyzed. In order to account for different surface chemistries, five self-assembled monolayers that differ in resistance to microfouling and wettability were used. Water samples from two static immersion test sites along the east coast of Florida were collected at two different times of the year and used for experiments. Spectral ellipsometry revealed that conditioning films were formed within the first 24 h and contact angle goniometry showed that these films changed the wettability and rendered hydrophobic surfaces more hydrophilic and vice versa. Infrared reflection adsorption spectroscopy showed that the composition of the conditioning film depended on both the wettability and immersion site. Laboratory and field assays showed that the presence of a conditioning film did not markedly influence settlement of microorganisms.     

Waterborne copper disrupts circadian rhythm in red seabream (Pagrus major)

We investigated the effect of copper (Cu) on circadian rhythms in red seabream, Pagrus major, under various concentrations of Cu (10, 20, 30 and 40 μg/L). To examine variability in circadian rhythms, we measured changes in the period 2 (Per2), cryptochrome 1 (Cry1), serotonin and arylalkylamine N-acetyltransferase (AANAT2) proteins. We found that circadian rhythm-related plasma proteins were significantly lower in a high-Cu environment (30–40 μg/L) than in low-Cu concentration (0, 10, or 20 μg/L). Our results indicate that environmental Cu at concentrations greater than 30 μg/L can have deleterious effects on fish circadian rhythms.     

Flexible growing rods: a biomechanical pilot study of polymer rod constructs in the stability of skeletally immature spines

Background

Surgical treatments for early onset scoliosis (EOS) correct curvatures and improve respiratory function but involve many complications. A distractible, or ‘growing rod,’ implant construct that is more flexible than current metal rod systems may sufficiently correct curves in small children and reduce complications due to biomechanical factors. The purpose of this pilot study was to determine ranges of motion (ROM) after implantation of simulated growing rod constructs with a range of clinically relevant structural properties. The hypothesis was that ROM of spines instrumented with polymer rods would be greater than conventional metal rods and lower than non-instrumented controls.

Methods

Biomechanical tests were conducted on six thoracic spines from skeletally immature domestic swines (35–40 kg). Paired pedicle screws were used as anchors at proximal and distal levels. Specimens were tested under the following conditions: control, then dual rods of polyetheretherketone (PEEK) (diameter 6.25 mm), titanium (4 mm), and cobalt-chrome alloy (CoCr) (5 mm). Lateral bending (LB) and flexion-extension (FE) moments were applied, and vertebral rotations were measured. Differences were determined by two-tailed t-tests and Bonferroni for four primary comparisons: PEEK vs control and PEEK vs CoCr, in LB and FE (α?=?0.05/4).

Results

In LB, ROM of spine segments after instrumenting with PEEK rods was lower than the non-instrumented control condition at each instrumented level. ROM was greater with PEEK rods than with Ti and CoCr rods at every instrumented level. Combining treated levels, in LB, ROM for PEEK rods was 35 % of control (p?<?0.0001) and 270 % of CoCr rods (p?<?0.01). In FE, ROM with PEEK was 27 % of control (p?<?0.001) and 180 % of CoCr (p?<?0.01). At proximal and distal adjacent non-instrumented levels in FE, mean ROM was lower for PEEK than for either metal.

Conclusions

PEEK rods increased flexibility versus metal rods, and decreased flexibility versus non-instrumented controls, both over the entire instrumented segment and at each individual level. Smaller mean increases in ROM at proximal and distal adjacent motion segments occurred with PEEK compared to metal rods, which may help decrease complications, such as junctional kyphosis. Flexible growing rods may eventually help improve treatment options for young patients with severe deformity.

     

Enzymatic cleaning of biofouled thin-film composite reverse osmosis (RO) membrane operated in a biofilm membrane reactor

Application of environmentally friendly enzymes to remove thin-film composite (TFC) reverse osmosis (RO) membrane biofoulants without changing the physico-chemical properties of the RO surface is a challenging and new concept. Eight enzymes from Novozyme A/S were tested using a commercially available biofouling-resistant TFC polyamide RO membrane (BW30, FilmTech Corporation, Dow Chemical Co.) without filtration in a rotating disk reactor system operated for 58 days. At the end of the operation, the accumulated biofoulants on the TFC RO surfaces were treated with the three best enzymes, Subtilisin protease and lipase; dextranase; and polygalacturonase (PG) based enzymes, at neutral pH (~7) and doses of 50, 100, and 150 ppm. Contact times were 18 and 36 h. Live/dead staining, epifluorescence microscopy measurements, and 5 μm thick cryo-sections of enzyme and physically treated biofouled membranes revealed that Subtilisin protease- and lipase-based enzymes at 100 ppm and 18 h contact time were optimal for removing most of the cells and proteins from the RO surface. Culturable cells inside the biofilm declined by more than five logs even at the lower dose (50 ppm) and shorter incubation period (18 h). Subtilisin protease- and lipase-based enzyme cleaning at 100 ppm and for 18 h contact time restored the hydrophobicity of the TFC RO surface to its virgin condition while physical cleaning alone resulted in a 50° increase in hydrophobicity. Moreover, at this optimum working condition, the Subtilisin protease- and lipase-based enzyme treatment of biofouled RO surface also restored the surface roughness measured with atomic force microscopy and the mass percentage of the chemical compositions on the TFC surface estimated with X-ray photoelectron spectroscopy to its virgin condition. This novel study will encourage the further development and application of enzymes to remove biofoulants on the RO surface without changing its surface properties.     

Prevention of biofilm formation on titanium surfaces modified with conjugated molecules comprised of antimicrobial and titanium-binding peptides

Specific binding of antimicrobial peptides to titanium (Ti) surfaces may serve to prevent biofilm formation, leading to a reduction in peri-implantitis. This study evaluated the binding behavior of conjugated molecules consisting of antimicrobial and hexapeptidic Ti-binding peptides (minTBP-1) using the quartz crystal microbalance (QCM-D) technique, and investigated the effect of modification of Ti surfaces with these peptides on the bioactivity of Porphyromonas gingivalis. Four kinds of peptide were prepared: histatin 5 (DSHAKRHHGYKRKFHEKHHSHRGY), minTBP-1 + histatin 5 (RKLPDAPDSHAKRHHGYKRKFHEKHHSHRGY), lactoferricin (FQWQRNMRKVR), and minTBP-1 + lactoferricin (RKLPDAPGGFQWQRNMRKVR). The QCM-D analysis demonstrated that significantly larger increases in peptide adsorption were observed in the conjugated peptides than in antimicrobial peptides alone. In addition, ATP activity in P. gingivalis in peptide-modified specimens significantly decreased compared to that in the Ti control. These results indicate that surface modification with conjugated molecules consisting of antimicrobial and Ti-binding peptides is a promising method for reduction of biofilm formation on Ti surfaces.     

Matrix-assisted laser desorption ionization mass spectrometry: a new tool for probing interactions between proteins and metal surfaces. Use in dental implantology.

The fixation in the bone of an artificial titanium tooth root is believed to be initiated by the rapid adsorption of the proteins present in the surgical cavity on the titanium surface. The study of this adsorption should make it possible to predict the osseointegration capacities of new implant surface treatments. We describe here a new method, based on matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS), for quantifying proteins adsorbed on titanium surfaces fully identical to these designed for implantology. The key step of this method is a new MALDI-MS sample preparation allowing the adsorbed proteins to be removed from the surface and to be homogeneously dispersed in the matrix crystals. The adsorption of a model protein (lysozyme) on two titanium surfaces (polished and sandblasted) was studied in order to evaluate the method. The absolute MALDI-MS intensity was shown to vary linearly with the amount of adsorbed lysozyme. After dipping the titanium surfaces for different times in lysozyme solutions at different concentrations, the maximum amount of adsorbed lysozyme was measured by MALDI-MS and was shown to correspond to a lysozyme monolayer, which is consistent with results described in the literature.