Journal of Colloid and Interface Science 356 (2011) 454–464

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Journal of Colloid and Interface Science

www.elsevier.com/locate/jcis

Deposition of colloid particles on protein layers: Fibrinogen on mica

Z. Adamczyk ⇑, M. Nattich, M. Wasilewska, M. Sadowska
Institute of Catalysis and Surface Chemistry, Polish Academy of Science, Niezapominajek 8, 30-239 Cracow, Poland

a r t i c l e i n f o a b s t r a c t

Article history: Colloid particle deposition was applied to characterize fibrinogen (Fb) monolayers on mica, which were
Received 21 October 2010 produced by controlled adsorption under diffusion transport. By adjusting the time of adsorption and the
Accepted 3 January 2011 bulk Fb concentration, monolayers of desired surface concentration were obtained. The surface concen-
Available online 14 January 2011
tration of Fb was determined directly by AFM enumeration of single molecules adsorbed over the sub-
strate surface. It was proven that Fb adsorbed irreversibly on mica both at pH 3.5 and at pH 7.4 with
Keywords: the rate governed by bulk transport. The electrokinetic properties of Fb monolayers produced in this
Colloid particle deposition
way were studied using the streaming potential method. The dependence of the apparent zeta potential
DLVO theory
Fibrinogen layers on mica
of Fb monolayers was determined as a function of the coverage. It was shown that for pH 3.5 the initial
Heterogeneous surface deposition negative zeta potential of the mica substrate was converted to positive for Fb coverage exceeding 0.16. On
Streaming potential measurements the other hand, for pH 7.4, the zeta potential of a Fb-covered mica remained negative for the entire cov-
Zeta potential of fibrinogen-covered mica erage range. The charge distribution in Fb monolayers was additionally studied using the colloid deposi-
tion method, in which negatively and positively charged polystyrene latex particles (ca. 800 nm in
diameter) were used. An anomalous deposition of negative latex particles on substrates exhibiting a neg-
ative zeta potential was observed. Results of these experiments were quantitatively interpreted in terms
of the fluctuation theory assuming that adsorption sites consisted of two and three Fb molecules, for pH
3.5 and 7.4, respectively. These results suggested that for pH 7.4, the distribution of charge on Fb mole-
cules was heterogeneous, characterized by the presence of positive patches, whereas the average zeta
potential was negative, equal to 19 mV. The utility of the colloid deposition method for studying Fb
monolayers was further demonstrated in deposition experiments involving positive latex particles. It
was shown that for a rather broad range of fibrinogen coverage, both the positive and the negative latex
particles can adsorb on surfaces covered by Fb, which behaved, therefore, as superadsorbing surfaces. It
was also concluded that the colloid deposition method can be used to determine the Fb bulk concentra-
tion for the range inaccessible for other methods.
Ó 2011 Elsevier Inc. All rights reserved.

1. Introduction using electron microscopy. Recently, many atomic force micros-

Adsorption of proteins at solid interfaces is important in many
fields of science and in various practical processes. It is involved
in thrombosis, artificial organ failure, plaque formation, the fouling
of contact lenses and heat exchangers, in ultrafiltration and mem-
brane filtration, where it exerts a negative influence.
On the other hand, controlled protein deposition on various sur-
faces is a prerequisite of their efficient separation by chromatogra-
phy, filtration, for biosensing, bioreactors, immunologic assays, etc.
Numerous studies were devoted to fibrinogen (Fb), because of
its fundamental role in blood clotting, platelet adhesion, thrombo-
sis, angiogenesis, wound healing, and tumor growth.
Fibrinogen size, shape, and conformations on surfaces (mostly
mica) were determined by Hall and Slayter [1] and others [2–4]
⇑ Corresponding author. Fax: +48 12 425 19 23.
E-mail address: ncadamcz@cyf-kr.edu.pl (Z. Adamczyk).
copy (AFM) studies were conducted with the aim of determining
conformations of fibrinogen on hydrophilic and hydrophobic solid
substrates [5–12].
Experimental studies on the kinetics of fibrinogen adsorption
are also abundant. Malmsten, using ellipsometry, determined the
kinetics of fibrinogen adsorption on methylated silica for physio-
logic conditions, i.e., pH 7.4 and I = 0.15 M [13].
Systematic measurements of fibrinogen deposition were per-
formed by Ortega-Vinuesa et al. [5,6], who measured the thickness
of the protein layer on silicon plates by ellipsometry as a function of
pH and ionic strength, for a fibrinogen bulk concentration of
70 ppm.
Precise kinetic measurements of fibrinogen adsorption on sili-
con and modified glass surfaces forming parallel-plate channels
were performed using the in situ fluorescent TIRF technique by
Santore and co-workers [11,14,15]. These results were supple-
mented with an interesting AFM determination of the kinetics of
fibrinogen adsorption for a low coverage range.

0021-9797/$ - see front matter Ó 2011 Elsevier Inc. All rights reserved.
doi:10.1016/j.jcis.2011.01.009
 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464
A significant spread in the maximum coverage of fibrinogen
was reported in these works with the lowest and highest value
ranging between 1.4 and 11 mg m2. It seems that this discrepancy
can be attributed to different adsorption mechanisms of fibrinogen
on various substrates and for various concentration regimes. For
hydrophilic surfaces, and the low concentration regime, fibrinogen
adsorption occurs according to the side-on mechanism with the
maximum amount of adsorbed protein equal to 1.4–1.7 mg m2,
depending on the degree of hydration. This is in accordance with
recent theoretic results discussed in Ref. [16].
For hydrophobic surfaces and higher fibrinogen concentrations,
the end-on adsorption of the protein becomes feasible with the
maximum coverage many times higher than for the side-on config-
uration. However, since the binding energy in the end-on configu-
ration is much lower than for the side-on orientation, a partial
reversibility of adsorption occurs, which can explain the source
of discrepancies.
Contrary to kinetic aspects, little is known about the electrostat-
ics of fibrinogen monolayers on solid substrates under wet condi-
tions, which is of vital significance. This kind of information can be
derived from electrokinetic studies, e.g., streaming potential mea-
surements performed previously for lysosyme [17,18], bovine ser-
um albumin (BSA), immuno-gamma globulins IgG [19] and for
chymotrypsin [20]. Interestingly, a significant protein adsorption
was observed in the latter work despite the fact that both the mica
substrate and the protein molecules were negatively charged. This
effect was explained by a dipolar distribution of the charge on the
protein.
There are only few works devoted to electrokinetic measure-
ments of fibrinogen adsorption on solid substrates. Zembala and
Dejardin [21] determined variations in the streaming potential of
silica capillaries on fibrinogen adsorption as a function of time
for pH 7.3 and the ionic strength of 102 M. A systematic increase
in the negative potential was observed on fibrinogen adsorption,
which was also negatively charged under these conditions.
Similar studies were performed in the work noted above by Kal-
asin and Santore [15] who determined the zeta potential of nega-
tively charged silica spheres (1 lm of diameter) covered by a
controlled amount of fibrinogen using microelectrophoresis. A
monotonic increase in the negative zeta potential of silica particles
was observed on fibrinogen adsorption, which attained a limiting
value of 20 mV for fibrinogen coverage larger than 1 mg m2
(for the ionic strength of 5–26 mM, NaCl).
Recently, systematic studies of fibrinogen adsorption on mica,
monitored by AFM and streaming potential measurements, were
reported [22]. The dependence of the apparent zeta potential of
mica on the coverage of fibrinogen was determined. A quantitative
interpretation of these experimental data was achieved in terms of
a theoretic model considering a 3D adsorption of fibrinogen mole-
cules as discrete particles, approximated as strings of touching
beads. The Gouy–Chapman model, based on the concept of a con-
tinuous charge distribution, was proven to be inadequate. It was
also suggested that anomalous adsorption occurring at pH 7.4
could be explained in terms of a heterogeneous charge distribution
on fibrinogen, whose molecules exhibited patches of positive
charge, even if the net charge was negative.
Although the streaming potential method offers the unique pos-
sibility of direct, in situ measurements, it is tedious, and does not
provide one with information on the local distribution of coverage
and charge within the protein monolayer.
Therefore, in this work, a complementary method of analyzing
protein layers on solid substrates under wet, in situ conditions is
used. The technique, referred sometimes to as the colloid enhance-
ment (CE) [23], consists in unspecific deposition of larger colloid
particles onto protein monolayers. This enables one to determine
a functional relationship between the amount of adsorbed protein
455
and the amount of adsorbed colloid, which can be assessed quan-
titatively via microscopic counting. Using the colloid deposition
method, in conjunction with electrokinetic characteristics of pro-
tein-covered substrates, one can determine mechanisms of colloid
deposition on heterogeneous surfaces, which is the primary goal of
this work.
As shown previously [14,24–27], in the case of heterogeneous
surfaces the standard DLVO theory is not applicable because the
net kinetics of colloid particle deposition is governed by favorable
areas (patches), whose zeta potential deviates from the average va-
lue pertinent to the entire substrate surface. In view of these pre-
dictions, another goal of our work is to determine the range of
applicability of this theory for predicting deposition kinetics of col-
loid particles on heterogeneous surfaces produced by controlled
fibrinogen adsorption.
The research presented in this work has also a practical impact
since the colloid deposition method can be exploited for a quanti-
tative determination of protein coverage and consequently for
detecting the bulk protein concentration.
We focus our attention on fibrinogen whose bulk characteristics
and surface electrokinetic properties have been precisely deter-
mined, which enables a quantitative analysis of the experimental
data.
2. Materials and methods
2.1. Materials
Fibrinogen from bovine plasma, fraction I, type IV (65% protein,
containing approximately 20% sodium chloride and 15% sodium
citrate) was purchased from Sigma (F4753) and used without fur-
ther purification. Protein was 93% clottable.
Ruby muscovite mica obtained from Continental Trade was
used as substrate, and was freshly cleaved immediately prior to
use without any pretreatment.
Water was purified using a Millipore Elix 5 apparatus. Chemi-
cals and reagents (Trizma base, sodium chloride, hydrochloric acid,
sodium hydroxide) were commercial products of Sigma–Aldrich
and used without further purification.
Positively charged amidine polystyrene latex (Invitrogen) and
negatively charged sulfonate polystyrene latex (synthesized in
our laboratory) were used in this work.
2.2. Experimental methods
The solutions of Fb used in subsequent measurements were
prepared as follows: a known amount of protein powder was dis-
solved in NaCl aqueous solution having an ionic strength of
1  103 or 1  102 M. The pH of these Fb solutions was regulated
by the addition of HCl or NaOH. Then, the suspension was gently
mixed using a magnetic stirrer, and filtered through the Millex-
GS 0.45-lm filter to eliminate aggregates and impurities. The
experimental temperature was kept constant at 293 ± 0.1 K.
The true concentration of Fb in these solutions was determined
using a high precision densitometer (Anton Paar, type DMA5000 M)
by measuring the specific density of the fibrinogen solutions (nom-
inal Fb content of 200–2000 ppm) and the specific density of the
supernatant solution acquired by a membrane filtration.
These concentrated stock Fb solutions were then diluted prior
to the adsorption experiments to a desired degree (usually 0.5–
5 ppm).
The purity of the Fb solutions was checked by the dynamic sur-
face tension measurements carried out using the pendant drop
shape method. There was practically no change in the surface ten-
sion of the 10 ppm solution of Fb within a 2-h time period, which is
456 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464
much longer than the typical time of DLS and deposition measure-
ments, which usually last 15–60 min.
The diffusion coefficient of Fb under various conditions was
determined by dynamic light scattering (DLS), using the Zetasizer
Nano ZS Malvern instrument (measurement range of 0.6 nm–
6 lm). This technique measures the time-dependent fluctuations
in the intensity of scattered light which occur because particles un-
dergo Brownian motion. Analysis of these intensity fluctuations
enables determination of the diffusion coefficients of particles
which are converted into size distributions.
The microelectrophoretic mobility le of Fb was measured using
the Zetasizer Nano ZS Malvern instrument deploying the laser
Doppler velocimetry (LDV) technique (measurement range of
3 nm–10 lm). The zeta potential was calculated using the equation
3g
f¼ l;
2eFðjaÞ e
where f is the zeta potential of proteins, g is the dynamic viscosity
of water, e is the dielectric constant of water, and F(ja) the function
of the dimensionless parameter ja = a/Le, Le ¼ ðekT=2e2 IÞ1=2 is the
double-layer thickness, e the elementary charge, P k2 the Boltzman
constant, T the absolute temperature, I ¼ 12 i c i zi is the ionic
strength, ci the ion concentrations, zi is the ion valency, and a the
characteristic dimension of a particle or a protein molecule (e.g.,
its hydrodynamic radius).
AFM measurements of Fb layers were carried out using the NT-
MDT Solver PRO device with the SMENA SFC050L scanning head.
The measurements were performed in semicontact mode using a
silicon probe (polysilicon cantilevers with resonance frequencies
of 240 kHz +/10% or 140 kHz +/10%, a typical tip curvature of
10 nm, and a cone angle less than 20°). The protein solution of a
desired concentration was allowed to adsorb on the surface for
an appropriate time interval. The substrate was then removed,
rinsed for 30 s using double distilled water, and dried under a gen-
tle nitrogen stream.
The latex particle size, as a function of ionic strength and pH,
was determined using a laser diffractometer (LS 13 320 particle
size analyzer, Beckman Coulter) with an accuracy of a few percent
and independently by the DLS method using the Malvern Zetasizer
Nano ZS.
The zeta potential of the latex was determined as a function of
the ionic strength and pH using the ZetaPals Brookhaven and the
Zetasizer Nano ZS.
Streaming potential measurements were carried out using a
home-made apparatus, exploiting the parallel-plate channel flow,
according to the procedure described previously [22,28–30].
Fibrinogen adsorption was carried out in a diffusion cell of a
volume equal to 40 ml, filled with a protein suspension of a desired
concentration, ionic strength, and pH. Freshly cleaved mica sheets
(usually 4–8) were placed vertically in the cell for a desired time.
Afterward, some of the mica sheets (2–4) were rinsed with a
stream of pure buffer solution for 30 s and dried under a gentle
stream of nitrogen. Afterward, the number concentration of fibrin-
ogen was directly determined by AFM examination.
The rest of the mica sheets were used in latex deposition exper-
iments performed in another diffusion cell, having the dimensions
of 2  2  5 cm. It was filled with the latex suspension, typically
having the number concentration of 1–2  1010 cm3. Wet mica
sheets, precovered by Fb monolayers as described above, were
then immersed vertically into the latex suspension. Latex article
deposition proceeded over a desired time. The true coverage of
particles was determined by a direct optical microscope counting
under wet conditions, as described in our previous works
[28–31]. Particles were counted over 10–20 equal sized areas cho-
sen randomly on the mica sheet. The net number of deposited
particles considered was ca. 2000, which ensured that these mea-
surements had a relative precision below 2%, as determined by var-
iance analysis. It is worthwhile noting that the coverage was
determined in absolute terms, i.e., H ¼ Sg hNi, where hNi is the
average surface concentration of particles (number of particles
per unit area) and Sg ¼ pa2 is the cross section of the particle (a
is the particle radius). Consequently, the coverage represents the
fraction of the entire surface area occupied by particles.
However, because of multiple scattering, the optical microscope
method became less accurate for coverage exceeding 0.3. In this
case the AFM method was used under dry conditions to determine
the surface concentration of particles and their coverage.
3. Results and discussion
ð1Þ The physicochemical characteristics of Fb, latex particles, and
the mica substrate, needed for a quantitative interpretation of
fibrinogen adsorption and latex deposition data, were initially
determined. These comprised the diffusion coefficient of Fb
(hydrodynamic diameter), microelectrophoretic mobility, and
electrokinetic (zeta) potential.
It was found by DLS measurements that the diffusion coefficient
of Fb was independent of its bulk concentration (within the range
of 100–2000 ppm), assuming an average value of 2.1  107
cm2 s1 for the ionic strength range of 103–102 M and for pH
3.5 and 7.4. Using this value the Stokes hydrodynamic radius of
Fb was calculated using the known dependence [32].
kT
RH ¼ ; ð2Þ
6pgD
where RH is the hydrodynamic radius, and D is the diffusion
coefficient.
It was calculated from Eq. (2), that RH = 12.1 nm (see Table 1). It
is interesting to observe that this is much higher than the radius of
the equivalent sphere equal to 4.5 nm [32] and reflects an elon-
gated shape of Fb determined from crystallographic data (see
Table 1).
The electrophoretic mobility of fibrinogen as a function of pH
and the ionic strength was also determined according to the meth-
od described above. Knowing the electrophoretic mobility of a par-
ticle, one can calculate the average number of uncompensated
electrokinetic charges per molecule Nc from the Lorenz–Stokes
relationship [32] and the zeta potential of fibrinogen ff from Eq.
(1). It was determined that ff was positive, equal to 24 and
28 mV for pH 3.5 and the ionic strength of 102 and 103 M,
respectively (see Table 1). On the other hand, for pH 7.4, ff became
negative, equal to 19 and 21 mV for the ionic strength of 102
and 103 M, respectively (see Table 1). From the electrophoretic
mobility measurements, it was also determined that the isoelectric
point of fibrinogen (zero value of its zeta potential) was pH 5.8.
The size of the latex particles was determined by laser diffrac-
tometry and the DLS method. Both values fell within the experi-
mental margins of error estimated to be 5%. Thus, the average
diameter of the A800 (positive) latex for the pH range 3–7.4 was
810 nm (ionic strength 103–102 M) and for the L800 (negative)
latex 800 nm, for the same range of pH and ionic strength (see
Table 1). The size and shape of latex particles were also checked
by electron microscopy (SEM); see Fig. 1. The average diameter
of the A800 latex (determined from ca. 200 particles) was practi-
cally the same as that determined by the laser diffractometer.
The zeta potential of both latex particles was also determined as
a function of pH and ionic strength. As can be seen (see Table 1),
the A800 zeta potential was fairly independent of pH, assuming
77 mV for I = 103 M and 95 mV for I = 102 M. The zeta potential
of the L800 latex was also practically independent of pH assuming
 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464 457

Table 1
Physicochemical properties of particles and the mica substrate.

Substance Property
Mica Latex A 800 Latex L 800 Fibrinogen
Zeta potential f (mV) pH 3.5
103 M 63 77 82 28
102 M 52 95 102 24
Zeta potential f (mV) pH 7.4
103 M 112 75 80 21
102 M 80 95 105 19
2 RH (nm) averaged I = 102–103 M – 810 800 24.2
47 nm

Fibrinogen molecular shape

7nm

Fig. 1. Latex particles on mica, SEM images, (a) the L800 latex, (b), the A800 latex.

an average value of 82 mV (for I = 103 M) and 102 mV for

I = 102 M.
The zeta potential of the mica substrate was determined via the
streaming potential measurements according to the procedure de-
scribed elsewhere [22,28–30]. For the ionic strength of 103 M, the
zeta potential of mica was 63 mV for pH 3.5 and 112 mV at pH
7.4. For the ionic strength of 102 M, the zeta potential of mica was Fig. 2. Fibrinogen monolayers on mica (deposition conditions: cb = 0.3 ppm, AFM
less negative, equal to 52 mV for pH 3.5 and 80 mV for pH = 7.4 images, semicontact mode, air). (a) Surface concentration of fibrinogen
(see Table 1). N = 125 lm2 (pH 3.5). (b) Surface concentration of fibrinogen N = 123 lm2 (pH
These results suggest that Fb should adsorb on mica for pH 3.5, 7.4).
due to favorable electrostatic interactions, because the zeta poten-
tial of Fb is positive and the zeta potential of mica strongly nega- atic Fb adsorption experiments were conducted under diffusion-
tive. For pH 7.4, both the Fb and the mica zeta potentials were controlled transport, according to the procedure described above.
negative, which is thought to prevent Fb adsorption because of The number of adsorbed Fb molecules on the mica substrate im-
electrostatic repulsion. In order to verify this hypothesis, system- mersed in the Fb suspension was determined by a direct AFM
458 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464
counting. The topology of fibrinogen monolayers on mica deter-
mined this way is shown in Fig. 2 for pH 3.5 and 7.4. As can be seen,
fibrinogen molecules appear as isolated entities, which facilitates
their enumeration by AFM. Therefore, using this counting proce-
dure the kinetics of Fb adsorption can be determined.
It is interesting to note that a similar AFM counting procedure of
isolated fibrinogen molecules was used by Tsapikouni and Missirlis
[12], who determined its coverage on mica for pH 7.4 and ionic
strength of 1.5  102, 0.15, and 0.5 M.
In Fig. 3, the kinetics is shown as the dependence of the reduced
surface concentration of Fb (number of Fb molecules per lm2 for
unit bulk concentration of 1 ppm) N/cb on the square root of the
adsorption time t1/2 (I = 103 M, pH 3.5, and 7.4, T = 293 K). As
can be seen, the experimental data in Fig. 3 are agreement with
theoretic results (depicted by the solid line) stemming from the
formula, pertinent to an irreversible, diffusion-controlled adsorp-
tion mechanism [33,34],
 1=2
D methods.
N¼2 t1=2 nb ð3Þ
p Knowledge of the surface concentration of Fb is necessary for a
where nb = Cb Av  106/Mw is the number concentration of Fb mol-
ecules in the bulk (Cb is the Fb concentration in ppm, and Av is the
Avogadro constant).
This suggests that Fb adsorption on mica for pH 3.5 was irre-
versible and bulk transport controlled with negligible surface
transport resistance. This seems quite natural considering the
strong attraction between the Fb molecules and the oppositely
charged mica substrate.
However, similar Fb adsorption kinetics was also observed in
the case of pH 7.4 (see Fig. 3), where a strong electrostatic repul-
sion was expected due to the same sign of zeta potential of Fb
and mica. Such anomalous fibrinogen deposition under physiologic
conditions (pH 7.4) was also previously observed by Zembala and
Dejardin [21], Malmsten [13], Ortega-Vinuesa et al. [5,6], Toscano
and Santore [11] and Kalasin and Santore [15].
As suggested in Refs. [9,15], the attractive interactions at this
pH may originate from the heterogeneous charge distribution over
the fibrinogen molecule. Hence, besides being negatively charged,
the Fb molecule exhibits positively charged patches, which can
form stable electrostatic bonds with the mica surface. This effect
was confirmed in a recent publication [22].
800
600
N/cb [µm ppm ]
-1
-2
400
200
0
0 2 4 6 8 10
t1/2 [min1/2]
Fig. 3. The dependence of the reduced surface concentration of fibrinogen N/cb
[lm2 ppm1] on the square of adsorption time t1/2, I = 103 M. The solid points
denote experimental results obtained by a direct AFM enumeration for pH 3.5
(various bulk concentrations of fibrinogen) and the hollow points denote results
obtained for pH 7.4. The solid line shows exact theoretic results obtained using the
RSA model.
The results shown in Fig. 3 have interesting practical implica-
tions showing that one can determine in a quite precise and conve-
nient way the bulk concentration of Fb if it is unknown. This can be
done by plotting N determined by AFM vs. the square root of
adsorption time, t1/2. Knowing the slope of this dependence
sf ¼ DN=Dt1=2 , one can determine the concentration of fibrinogen
in the bulk (in ppm) from the simple relationship.
 p 1=2
Mw
cb ¼  1013 sf ð4Þ
Av 2:4D
where cb is expressed in ppm and sf is expressed in lm2 min1/2.
Using the value of the diffusion coefficient and molecular
weight pertinent to fibrinogen one can express Eq. (4) in the simple
form
C b ¼ 0:014sf : ð5Þ
From Eq. (5) one can directly determine the bulk concentration
of Fb even below 0.1 ppm, which is rather difficult using other
proper interpretation of results for colloid deposition
measurements.
Other data needed for this purpose are the zeta potential f of
mica covered by Fb monolayers, which were acquired via stream-
ing potential measurements. It is to be emphasized that our
streaming potential measurements were carried out under wet,
in situ, conditions, which excludes any conformation changes of
fibrinogen monolayers induced by drying.
Results are presented in Fig. 4 showing the dependence of f on
the coverage of Fb, calculated from the equation
H ¼ Sg N; ð6Þ
where Sg is the characteristic cross section of the protein molecule.
As demonstrated in Refs. [16,22] the characteristic cross-
sectional Sg for the side-on adsorption of Fb on mica is equal to
128 nm2. Using the characteristic cross section for the side-on
adsorption, the jamming coverage for fibrinogen becomes 0.29
[16].
It can be seen in Fig. 4a and b that for pH 3.5, where Fb is pos-
itively charged, a steep, quasilinear increase in the zeta potential of
mica is observed for Hf < 0.1. Then, for Hf > 0.16, the zeta potential
became positive and approached saturation values, which were
markedly lower than the bulk values of the zeta potential of fibrin-
ogen (equal to 28 and 24 mV for 103 and 102 M, NaCl, respec-
tively). It is interesting to note that the f on Hf dependencies
shown in Fig. 4 are quite similar for both ionic strengths, i.e.,
103 and 102 M. They can be adequately reflected in terms of
the theoretic model (depicted by solid lines in this figure) postulat-
ing a irreversible, side-on adsorption of Fb [16,22]. In this theoretic
approach, a 3D distribution of charge over Fb molecules is consid-
ered, as well as the flow damping effect due to adsorbed molecules
[35,36]. It is interesting to note that this model was successfully
applied before for interpretation of zeta potential of surfaces cov-
ered by colloid particles and polyelectrolytes [28–31,37,38]. An
interesting feature, which was confirmed in these experiments is
that the limiting value of the zeta potential of particle-covered sur-
faces is 0.71 fp (where fp is the bulk zeta potential of the particle or
protein molecule) rather than fp as intuitively expected.
Analogous zeta potential data obtained for pH 7.4 are shown in
Fig. 4c. (I = 102 M). As can be seen, no inversion of the mica zeta
potential was observed, because the bare mica zeta potential and
the bulk zeta potential of Fb were both negative. However, in this
case, theoretic results calculated by assuming a uniform charge
distribution over fibrinogen molecules corresponding to the bulk
value of ff = 19 mV (solid line in Fig. 4c) deviate from experimen-
tal points, which lie below. This suggests that the effective zeta
 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464 459

(a) 40 itively charged patches on the fibrinogen molecule, which could be

responsible for the favorable electrostatic interactions with mica.
20
Because of the presence of positive patches, the local zeta potential
of negative patches, exposed to the liquid, becomes more negative,
than the average bulk value.
0
In order to check this prediction, latex deposition experiments
were performed according to the procedure described above. These
ζ -20 studies were performed under diffusion-controlled conditions at
pH 3.5 and 7.4. The number of latex particle adsorbed as a function
-40 of deposition time was determined by the direct counting proce-
dure using the optical microscopy and AFM.
A major advantage of the optical microscopy is that it enables
-60
measurements to be carried out under wet conditions, where par-
ticle positions over the substrate surface remain undisturbed. This
-80 allows one to reliably determine not only the coverage of particles
0.0 0.1 0.2 0.3 0.4 0.5
Θf but also their surface concentration and distributions over bare
and fibrinogen-covered substrates.
(b) 40 The statistical uniformity of colloid particle distributions was
confirmed quantitatively using variance analysis of the number
20 of particles deposited over equal-sized surface areas. This enabled
one to determine the coverage of latex particles on mica surfaces
precovered by fibrinogen sublayers and consequently the relation-
0
ship between the maximum (saturation) latex coverage HL and the
fibrinogen coverage Hf.
ζ -20 The procedure for determining HL was as described above. First,
fibrinogen monolayers of an appropriate density (determined by
-40 AFM) were produced on mica sheets under diffusion transport con-
ditions. Then, the wet sheets were immersed in the latex suspen-
-60 sion having a bulk concentration of typically 1.77  1010 cm3.
After completing the deposition run, typically lasting 24 h, the la-
tex distribution and coverage were independently determined by
-80
0.0 0.1 0.2 0.3 0.4 0.5 optical microscopy and AFM. It should be noted that in all these
Θf experiments, latex deposition was irreversible, as confirmed in
separate desorption runs, where the pure latex-covered mica
(c) 40 sheets were immersed for a prolonged time period (24 h) in pure
electrolyte having the same ionic strength and pH.
20 The dependencies of the maximum latex coverage HL on the
fibrinogen coverage Hf derived from these kinetic experiments
are plotted in Fig. 5 for pH 3.5 and in Fig. 6, for pH 7.4, respectively.
0
In order to compare results obtained for different ionic strengths,
the reduced maximum coverage HL/Hmx was used as the depen-
ζ -20 dent variable (where Hmx is the maximum coverage for a homoge-
neous surface) depending slightly on the ionic strength.
-40 As can be seen in Figs. 5 and 6, HL/Hmx was slightly dependent
on the ionic strength and increased monotonically with Hf . For pH
3.5 the increase of HL/Hmx with the fibrinogen coverage was quite
-60
abrupt, whereas for pH 7.4, the maximum latex coverage increased
at a much slower rate. Hence, an apparent threshold value of the
-80 Hf was needed to achieve significant latex deposition. For pH 7.4,
0.0 0.1 0.2 0.3 0.4 0.5
Θf this threshold value was approximately 0.04, whereas for pH 3.5,
it was significantly smaller, equal to 0.01. A similar behavior was
Fig. 4. The dependence of the zeta potential of mica f on the coverage of fibrinogen previously observed for latex particle adsorption on polyelectro-
Hf = SgN. The points denote experimental results obtained from the streaming lyte-covered mica [37,38].
potential measurements: (a) I = 103 M, pH 3.5, fi = 63 mV, ff = 28 mV; (b)
Analogous results, pertinent to the initial deposition rate of sil-
I = 102 M, pH 3.5, fi = 52 mV, ff = 24 mV. The solid lines represent exact theoretic
results calculated according to the electrokinetic model developed in Refs. [35,36]. ica particles on pDMAEMA and fibrinogen-covered silica slides,
(c) I = 102 M, pH 7.4. The solid lines represent exact theoretic results calculated were also reported [15]. In the latter case, the threshold value for
according to the electrokinetic model developed in Refs. [34,35] for fi = 80 mV, silica particle adsorption was roughly. 0.1 for I = 2.6  102 M
ff = 19 mV, and the dashed line for fi = 80 mV, ff = 30 mV. and 0.15 for I = 5  103 M. This difference stems from the fact that
our deposition experiments lasted 24 h, whereas the kinetic exper-
potential of adsorbed fibrinogen molecules is smaller (more nega- iments of Kalasin and Santore [15] were performed under convec-
tive) than in the bulk. Indeed, a good agreement between experi- tion conditions (in the parallel-plate channel) and lasted typically
mental data and the theoretic model is observed if the effective 5 min. These conditions were, therefore, less favorable for an effec-
zeta potential of fibrinogen equal to 30 mV is assumed (dashed tive attachment of silica particles under strong shearing forces pre-
line in Fig. 4c). As discussed in Ref. [22] this behavior can be ex- vailing at the channel’s walls.
plained in terms of a heterogeneous charge distribution over The results shown in Figs. 5 and 6 indicate unequivocally that
fibrinogen molecules. Thus, at pH 7.4, there apparently exists pos- latex particle immobilization was not possible at adsorption sites
460 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464

formed by only one fibrinogen molecule. If this were the case, HL/ 1.0
 1=2
2
Hmx should increase initially as k2 Hf =Hmx (where k ¼ pSag ), i.e.,
proportionally to Hf. Considering that in our case Sg = 1.28  0.8
1012 cm2 and pa2 = 5.03  109 cm2 one obtains k2 = 3.93  103.
Thus, the slope of the HL/Hmx vs. Hf dependence should be initially
very large initially and independent of pH. 0.6

ΘL/Θmx
It is interesting to note that the data shown in Figs. 5 and 6 are
anomalous in comparison to measurements of colloid particle
deposition on homogeneous surfaces [31,33]. This becomes evi- 0.4
dent if one notes that for pH 3.5 (see Fig. 6) and Hf < 0.15, where
a significant latex deposition was measured, the averaged zeta po-
0.2
tential of the fibrinogen-covered mica substrate remained negative
(see Fig. 4), i.e., of the same sign as the zeta potential of latex. This
should result in a strong electrostatic repulsion preventing particle 0.0
deposition. Qualitatively, this discrepancy could be explained in 0.0 0.1 0.2 0.3 0.4
terms of a heterogeneous distribution of positive charge due to Θf
natural fluctuations in adsorbed protein density. Consequently,
Fig. 6. Same as in Fig. 5 but for pH 7.4, I = 102 M. The points denote the
the local charge of the substrate could become positive, enabling
experimental results obtained by optical microscopy (d) and AFM (s).
latex particle deposition.
However, the results obtained for pH 7.4 (see Fig. 6) are even
more intriguing, because in this case all components involved in
the adsorption event, i.e., the bare mica substrate, fibrinogen mol-
is observed for negative values of the substrate surface zeta poten-
ecules, and latex particles exhibited negative zeta potentials.
tial, being of the same sign as the latex particle zeta potential.
Hence, paradoxically, the conjuncture of three negative interac-
It is interesting to compare this behavior with predictions
tions produced a positive effect, i.e., irreversible deposition of latex
stemming from the standard DLVO theory, which was previously
particles. This phenomenon can also be explained in terms of the
applied for successfully predicting particle deposition on homoge-
density fluctuation concept if a heterogeneous charge distribution
neous surfaces [31]. The main assumption of this theoretic ap-
over adsorbed fibrinogen molecules is assumed. Therefore, the
proach is that the van der Waals and electric double-layer
simple latex deposition experiments shown in Fig. 6 suggest quite
interactions are additive, which can be expressed for the spherical
unequivocally that at pH 7.4, where the averaged zeta potential of
particle/flat interface by the equation [34].
fibrinogen is negative (19 mV for I = 102 M), there were positive
patches on its surface. This supports the previous conclusion de- A123 a
/ðhÞ ¼  þ /0 eh=Le ; ð7Þ
rived from the streaming potential measurements [22]. 6h
The results shown in Figs. 5 and 6 can be interpreted in a more
where /ðhÞ is the particle / interface interaction energy, A123 is the
quantitative way if the normalized maximum coverage HL/Hmx is
Hamaker constant for the polystyrene/water/mica interactions, h is
plotted against the average zeta potential of fibrinogen-covered
the gap with between the particle and the interface, /0 ¼
mica f, rather than the fibrinogen coverage. The transformation
16aeðkT=eÞ2 tan hðfe=4kTÞ tan hðfp e=4kTÞ is the characteristic energy
from Hf to f was done using the results shown in Fig. 4.
for the linear superposition approximation (LSA) double-layer mod-
As seen in Fig. 7, a quite universal graph was obtained in this
el [39,40].
way, with experimental values of HL/Hmx fairly independent of
Assuming a typical value of A123 = 5  1014 kT, and f = 5 mV,
the ionic strength and pH. However, the results shown in Fig. 7
fp = 82 mV (see Table 1), I = 103 M one obtains from Eq. (7) that
are anomalous because a considerable deposition of latex particles
/ = 278 kT for h = 7 nm (fibrinogen diameter) and / = 0.1 kT for
h = 60 nm. Analogously, for I = 102 M, f = 5 mV, fp = 102 mV,
/ = 59 kT for h = 7 nm and / = 0.5kT for h = 15 nm. These estima-
tions indicate unequivocally that the energy barrier of considerable
1.0 height and extension of db = 15–60 nm is predicted to appear even
for such a low zeta potential of the substrate.
It is interesting to note that even for such low zeta potential of
0.8
the interface, the height of the barrier is mainly governed by the
electrostatic interactions.
ΘL/Θ mx

0.6 Knowing the specific energy distribution, one can calculate the
adsorption rate constant from the definite integral [33,34].
"Z #1
0.4 e/ðhÞ=kT
ka ¼ dh ; ð8Þ
db DðhÞ
0.2
where D(h) is the position-dependent diffusion coefficient of the la-
tex particles.
0.0 Knowing Ka, the latex coverage as a function of deposition time
0.00 0.05 0.10 0.15 0.20 0.25 0.30 and bulk concentration can be theoretically calculated by solving
Θf the governing diffusion equation using the finite-difference
scheme [33].
Fig. 5. The dependence of the reduced coverage of negative latex particles L800 HL/
Hmx on the fibrinogen coverage Hf. The points denote the experimental results
It is interesting to note that for a barrier height exceeding 10 kT
obtained by optical microscopy (d) and AFM (s) for ionic strength 103 M, (N, 4) (which is the case for f > 2 mV), the kinetics of latex particle
for ionic strength 102 M, pH 3.5, T = 293 K. deposition can be approximated by the expression.
 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464
1.0
0.8
0.6
ΘL /Θmx
0.4
0.2
0.0
-60 -40 -20 0 20
ζ 1 þ 0:314a2 þ 0:45a3
Fig. 7. The dependence of the reduced coverage of negative latex particles L800,
HL/Hmax on the zeta potential of fibrinogen-covered mica f determined by the
streaming potential method. The points denote the experimental results obtained
by optical microscopy and AFM for: (d) I = 103 M, pH 3.5; (N) I = 102 M, pH 3.5;
(4) I = 102 M, pH 7.4. The dashed line denotes theoretic results calculated from the
DLVO theory for a continuous zeta potential distribution.
 
D /b /b =kT
HL ¼ Sg ka nb t ffi e Sg nb t: ð9Þ
a kT
The results calculated using Eqs. (7)–(9), referred to as the DLVO
theory predictions, are shown by the dashed line in Fig. 7. As can be
seen, for f < 0, the DLVO theory predicts negligible latex deposition,
after 24 h. This deviates completely from experimental results,
indicating a significant deposition for substrate zeta potential as
low as 50 mV.
It is interesting to note that such behavior was observed before
for heterogeneous surfaces produced by controlled cationic poly-
electrolyte adsorption [37,38].
As postulated in these works, the deviation from the DLVO the-
ory is a direct manifestation of a discrete charge distribution
caused by natural fluctuations in the fibrinogen density over the
mica substrate. Because fibrinogen molecules exhibited a net posi-
tive zeta potential at pH 3.5, a local increase in their concentration
at mica could lead to formation of favorable adsorption sites, capa-
ble of immobilizing latex particles.
Recently, deviations from the classical DLVO theory were theo-
retically analyzed in Ref. [27] using the grid surface integration
(GSI) technique. Both the surface roughness effect and the charge
heterogeity were quantitatively evaluated. The surface heterogene-
ities were assumed to have the form of nanopillars having opposite
sign of zeta potential as the surface and colloid particles depositing
on it. A significant decrease in the energy barrier height with the
size of the nanopillars was predicted for their coverage of 0.25. This
seems in a qualitative accordance with our experimental observa-
tions show in Fig. 7. However, the effect of the nanopillars coverage
has not been studied in Ref. [27], which prohibits a quantitative
comparison with our experimental data. Moreover, these theoretic
results cannot account for the situation arising for pH 7.4, where the
net zeta potential of fibrinogen was negative. Since a significant
deposition of latex was also observed in this case (see Fig. 6) one
can suspect that there were positive patches on fibrinogen, which
were used to make permanent contacts with latex particles.
These effects can be properly analyzed in terms of the charge
fluctuation theory previously developed to interpret particle depo-
sition on surfaces covered by polyelectrolytes [36,37] and fibrino-
gen molecules [23]. In a recent work [26], this approach was
exploited to interpret the influence of ionic strength on silica par-
ticle deposition rates on polyelectrolyte-covered silica surfaces.
461
The basic assumption of the charge fluctuation approach is that
particle adsorption occurs at point-like sites distributed at random
over an otherwise homogeneous surface [41]. It is also assumed
that one site is capable of immobilizing one particle only. The basic
parameter of this approach, characterizing the density of adsorp-
tion sites (fibrinogen molecules in our case), is defined as
pa2
a ¼ pa2 Ns ¼ Hs ¼ k2 Hs ; ð10Þ
Sg
where Ns is the surface concentration of adsorption sites. As calcu-
lated above, in our case, k2 = 3.93  103.
Knowing the a parameter one can calculate the maximum cov-
erage of latex particles on a heterogeneous surface covered by sites
from the analytical dependence [23,41] modified in this work to
account for the fact that Hmx is a variable, rather than a constant.
!
HL ¼ Hmx 1  : ð11Þ
1 þ H1mx a þ 0:66a3 þ a7=2
In the limit of low coverage of sites, where a  1, Eq. (11) simplifies
to
HL ¼ k2 Hs : ð12Þ
This approach was generalized in Refs. [37,38] assuming that an
effective particle immobilization can occur on sites formed by ns
closely spaced molecules (polyelectrolytes or proteins). It was
shown in these works, assuming the Poisson statistics that the cov-
erage of adsorption sites composed of ns or more particles is given
by the expression.
X1
ðSHf Þn ðSHf Þns
Hs ðns Þ ¼ eSHf ffi ; ð13Þ
n¼ns
n ns !
where S ¼ S =Sg and S is the effective interaction area, whose size is
comparable with the cross-sectional area of the site, thus S  1.
As can be noted, the coverage of sites composed of two Fb mol-
ecules is proportional to H2f , three molecules H3f , etc. This means
that the number of such sites is very low for Fb coverage not
exceeding 0.1.
Using Eqs. (13) and (11) one can predict that latex coverage in-
creases initially according to the relationship
Hnf s
HL ¼ k2 : ð14Þ
ns !
Thus, in the case of ns = 2, a parabolic dependence of H L on the
Fb coverage is initially predicted, described by the equation
1 1
HL ¼ kHs ð2Þ ¼ k2 H2f : ð15Þ
2 2
In general, for arbitrary and ns, the latex coverage can be calcu-
lated from Eq. (11) using Eq. (13) to calculate Hs.
As can be seen in Fig. 8, the experimental data are not properly
reflected by theoretic results derived from these equations if ns = 1
is assumed, since a much steeper increase of HL/Hmx with the Fb
coverage was observed initially. This indicates that adsorption of
latex particles on sites consisting of a single Fb molecule was not
possible due to too low attractive interaction energy. An analogous
phenomenon was previously observed in the case of poly(allyla-
mine) hydrochloride and poly(ethylene imine) polyelectrolytes
[37,38], where latex particle deposition occurred at adsorption
centers composed of two or more closely spaced polyelectrolyte
chains.
However, a quantitative agreement of theoretic results with the
experimental data shown in Fig. 8 is attained when ns = 2 is as-
sumed (see curve 2). In particular, the parabolic dependence, de-
scribed by Eq. (15) for low Hf coverage, is confirmed. This
462 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464

suggests that the CE experiments of latex deposition on fibrinogen

1
monolayers can be exploited for an efficient determination of the 1.0
bulk concentration of the protein.
2
Analogous results obtained for pH 7.4 are shown in Fig. 9. As can
0.8
also be seen, the negative latex adsorbed efficiently for Hf > 0.04
(threshold value of fibrinogen coverage). As noted above, this can
be explained by assuming a heterogeneous charge distribution 0.6

ΘL/Θ mx
over Fb molecules, bearing patches of positive charge on their
surfaces.
Using the fluctuation theory, one can analyze this effect in more 0.4
detail by determining the number of Fb molecules necessary for
the immobilization of latex particles. As seen in Fig. 9, the experi- 0.2
mental data for pH 7.4 are in accordance with the theoretic predic-
tions derived from Eq. (13) assuming that adsorption sites were
formed by three Fb molecules (solid line in Fig. 10). It is interesting 0.0
to observe that for Hf < 0.1, the experimental data were well de- 0.00 0.05 0.10 0.15 0.20 0.25 0.30
Θf
scribed by the limiting analytical formula derived from Eq. (14)
for ns = 3, Fig. 8. The dependence of the reduced coverage of negative latex particles L800,
HL/Hmx on the fibrinogen coverage Hf. The points denote the averaged exper-
1
HL ¼ k2 H3f : ð16Þ imental results obtained by optical microscopy and AFM (s) for ionic strength
6 103 M, (N) for ionic strength 102 M, pH 3.5, T = 293 K. The solid lines 1–2 denote
theoretic prediction derived from the fluctuation theory, Eqs. (11) and (13) for the
Thus, in this case the latex coverage HL is expected to increase
adsorption site composed of one and two fibrinogen molecules, respectively. The
proportionally to the third power of the Fb coverage for the low dashed line denotes the limiting analytical solution, i.e., HL ¼ 12 k2 H2f =Hmx .
coverage range.
Immobilization of latex particles on sites composed of three Fb
molecules is understandable because at pH 7.4 (I = 102 M) the
partial positive charge on fibrinogen equals 12e [22] being consid-
1.0
erably smaller than the positive charge pH 3.5 (I = 102 M), esti-
mated to be 24e. However, one should remember that the partial 3

positive charge at pH 7.4 is overcompensated by the negative 0.8

charge, equal to 33e. Therefore, the experimental data shown in
Fig. 9 suggest strongly that positively charged patches over Fb mol-
ecules are separated by a considerable distance from negative 0.6
ΘL/Θmx

patches leading to a dipole formation. Considering the Fb molecule

structure and conformation [7–10] this could only happen if the
core region of Fb (D–E domains) was negatively charged, whereas
the side arms (aC domains) were positively charged. Hence, they
could act as anchors for the immobilization of Fb at the mica sur-
face. This concept is in accordance with the charge distribution
over Fb proposed in Ref. [9]. If such a charge distribution is true,
clusters of three Fb molecules forming a triangle could exhibit a
considerable positive charge (of about 36e) in the middle region,
which would be sufficient to immobilize irreversibly latex
particles.
Such a heterogeneous charge distribution over Fb molecules can
also explain why the standard DLVO theory does not work in the
case of the negative latex deposition for pH 7.4, where the average
zeta potential of the mica substrate remains negative for the entire
range of Fb coverage.
It is also interesting to observe that the results in Fig. 9, showing
a more gradual increase of latex coverage with Hf , can be exploited
for a sensitive determination of the bulk Fb concentration by the
colloid deposition method. Knowing HL, this can be done using
Eq. (11) to calculate the Fb coverage Hf and then Eq. (3) to calcu-
late cb for a fixed fibrinogen adsorption time.
In order to explore further the potential of the colloid deposi-
tion method for studying Fb layers, an additional series of deposi-
tion experiments was performed using the positive latex A800. The
results of these measurements are presented in Fig. 10 for pH 3.5
and in Fig. 11 for pH 7.4. As can be seen, for pH 3.5 and Hf < 0.16
the positive latex coverage remained practically the same as the
limiting value for homogeneous surfaces. For Hf > 0.16, the latex
coverage decreased abruptly to negligible values. Hence, in this
case, the latex deposition corresponded with the average zeta po-
tential of the mica substrate. Therefore, the DLVO theory proved
adequate (see dashed line in Fig. 10). This was so, because for the
0.4
0.2
0.0
0.0 0.1 0.2 0.3 0.4
Θf
Fig. 9. Same as in Fig. 8, but for pH 7.4, 102 M. The solid line 3 denote theoretic
prediction derived from the fluctuation theory, Eqs. (11) and (13) for the adsorption
site composed of three fibrinogen molecules. The dashed line denotes the limiting
analytical solution, i.e., HL ¼ 16 k2 H3f =Hmx .
high coverage range, fluctuations in the Fb molecule density (local
charge) within the area of interaction with the latex particles
(whose cross-sectional area is k2 = 3.8  103 times larger than the
fibrinogen molecule cross-sectional area) become negligible. As a
result, the local zeta potential of Fb-covered mica remained close
to the averaged zeta potential.
The strict correlation of positive latex deposition with the zeta
potential of fibrinogen-covered mica demonstrated in Fig. 10 can
be exploited for a precise determination of the isoelectric point
of protein-covered surfaces, which is of essential practical
significance.
The results shown in Fig. 10 also prove that the negative and po-
sitive latex deposition on Fb layers is a complementary process be-
cause for Hf < 0.16 the positive latex adsorbs fully (making a
negative replica of the Fb monolayer) and for Hf > 0.16 the nega-
tive latex adsorbs fully, making a positive replica. This behavior
can be utilized for performing a qualitative but very sensitive
 Z. Adamczyk et al. / Journal of Colloid and Interface Science 356 (2011) 454–464 463

1 1
1.0 1.0
2

0.8 2 0.8

0.6 0.6
ΘL/Θ mx

ΘL/Θmx
0.4 0.4

0.2 0.2

0.0 0.0
0.00 0.05 0.10 0.15 0.20 0.25 0.30 0.0 0.1 0.2 0.3 0.4
Θf Θf

Fig. 10. The dependence of the reduced coverage of latex particles HL/Hmx on the
fibrinogen coverage Hf for pH 3.5, T = 293 K. The points denote the averaged
experimental results obtained by optical microscopy and AFM for (d) the L800
latex, I = 103 M and (N) 102 M; (}) the A800 latex, I = 103 M and (O) 102 M. The
solid line 2 denotes theoretic predictions derived from the fluctuation theory, Eqs.
(11) and (13), and the dashed line 1 denotes the results predicted from the DLVO
theory for the A800 latex.
Fig. 11. Same as in Fig. 10 but for pH 7.4, 102 M. The points denote the averaged
experimental results obtained by optical microscopy and AFM for (d) the L800
latex, and (O) the A800 latex. The solid line 2 denotes theoretic predictions derived
from the fluctuation theory, Eqs. (11) and (13), and the dashed line 1 denotes the
results predicted from the DLVO theory for the A800 latex.

It was also concluded that the colloid deposition method can be

used for a quantitative determination of the Fb bulk concentration,
detection test of fibrinogen layers on mica, and, therefore fibrino-
for a range inaccessible for other methods.
gen presence in the bulk.
Another practical significance which can be drawn from these
results is that for fibrinogen coverage range of 0.02 < Hf < 0.16, Acknowledgments
both the negative and the positive latexes will deposit with a
comparable efficiency. Hence, such surfaces can be named super- This work was supported by the COST Action D43 Special Grant
adsorbing surfaces, because they are able to remove both posi- and Grant No. POIG.01.01.02-12-028/09-01. The authors are
tive and negatively charged colloid particles from their indebted to Dr. E. Bilańska for performing SEM measurements of
suspensions. This could be of vital significance for various filtra- latex monolayers.
tion processes.
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