Dental Materials (2006) 22, 533–544

www.intl.elsevierhealth.com/journals/dema

Extension of a one-step self-etch adhesive

into a multi-step adhesive
K.L. Van Landuyt, M. Peumans, J. De Munck,
P. Lambrechts, B. Van Meerbeek*

Leuven BIOMAT Research Cluster, Department of Conservative Dentistry, School of Dentistry,

Oral Pathology and Maxillo-Facial Surgery, Catholic University of Leuven, Kapucijnenvoer 7,
B-3000 Leuven, Belgium

Received 21 March 2005; accepted 11 May 2005

KEYWORDS Summary One-step self-etch adhesives are undoubtedly the most user-friendly
Adhesion; adhesives, but have been associated with lower bonding effectiveness as compared
Adhesive; to two-step and three-step adhesives. Conversion of a one-step self-etch system into
Enamel; a two-step self-etch adhesive by adding a bonding step, or into a three-step
Dentin; etch&rinse adhesive by adding a beforehand etching step and a bonding step might be
Monomer; tempting in order to improve bond strength.
Solvent; Objectives: The objective of this study was to investigate whether adding
Phase separation; application steps influences the bond strength of a one-step self-etch adhesive.
Extension; Methods: In this study, the bonding effectiveness of two experimental one-step self-
Multi-step; etch adhesives and three different commercial adhesives to enamel and dentin was
Self-etch determined using a micro-tensile bond-strength protocol. This procedure was
repeated for the experimental adhesives transformed into two-step self-etch and
three-step etch&rinse adhesives. In addition, their interaction with tooth tissue was
investigated using TEM and Feg-SEM.
Results: Transforming a one-step into a two-step self-etch adhesive did improve the
bond strength to enamel and dentin, though not significantly. By adding a preceding
etching step, the bond strength to enamel was significantly improved, but that to
dentin was decreased considerably. The latter must be attributed to hampered resin
infiltration of the one-step self-etch adhesive within the relatively deeply exposed
collagen fibril network.
Significance: Additional application of a hydrophobic bonding agent slightly
improved bonding effectiveness. Adding a preceding etching step is beneficial for
enamel but should be avoided for dentin as this will decrease bond strengths, and
may even jeopardize the bonding durability.
Q 2005 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

* Corresponding author. Tel.: C32 16 337 587; fax: C32 16 332 752.
E-mail address: bart.vanmeerbeek@med.kuleuven.ac.be (B.V. Meerbeek).

0109-5641/$ - see front matter Q 2005 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.dental.2005.05.010
534 K.L. Van Landuyt et al.

Introduction a rapid decline in bonding effectiveness has been

reported for simplified adhesives [20–22], and these
Current adhesive research focuses on the simplifi- findings are also confirmed by a recent review
cation of application procedure [1]. Reduction of concerning clinical effectiveness of adhesives [23].
the number of application steps should reduce The objective of this study was to investigate
manipulation time, and abate technique sensitivity, whether the bonding effectiveness of a 2-SEA
thus improving bonding effectiveness. This trend in consisting of a 1-SEA followed by a hydrophobic
adhesive dentistry has led to the introduction of bonding, and of a 3-E&R consisting of the same
self-etch adhesives, of which the one-step self-etch 1-SEA preceded by an etching step and followed by
adhesives (1-SEAs) or the so-called all-in-one a hydrophobic bonding is comparable to that of
adhesives are the most user-friendly adhesive commercial 2-SEAs and 3-E&Rs, and whether adding
systems nowadays on the market. Their application application steps influences the bond strength.
procedure involves a single step, combining etch- Therefore, the bonding effectiveness of two exper-
ing, priming and bonding [2,3]. imental 1-SEAs applied according to different
Research, however, so far has demonstrated that application procedures and three commercial
simplified systems do not bring the expected adhesives (one 1-SEA, one 2-SEA and one 3-E&R)
improvement in bonding effectiveness, in spite of was tested using a micro-tensile bond-strength
(mTBS) protocol. The interaction of the adhesives
their assumed reduced technique-sensitivity. Sev-
with dentin was comparatively characterized using
eral authors have reported lower bonding effec-
transmission electron microscopy (TEM) and scan-
tiveness for simplified adhesive systems [4–9].
ning electron microscopy (Feg-SEM). In addition,
Moreover, 1-SEAs in particular have been associated
the effect of acetone or ethanol used as solvent was
with considerable shortcomings. Tay et al. [10–13]
investigated in the experimental 1-SEAs.
have demonstrated that polymerized 1-SEAs are
The hypotheses tested in this study were (1) that
porous structures that can act as semi-permeable
extension of a 1-SEA into a 2-SEA and into a 3-E&R
membranes after polymerization, permitting bi-
by using the 1-SEA as a primer solution and by
directional water movement across the adhesive
adding application steps improves the bonding
layer if not lined by a cured composite. Nanometer-
effectiveness correspondingly; and (2) that there
sized voids in the adhesive layer, revealed by silver
is no difference in bonding effectiveness between
tracer, are suspected to function as water ducts. ethanol and acetone when used as solvent.
Reticular patterns of nanoleakage, so-called water-
trees, have been observed in the adhesive layer of
1-SEAs and are considered as sites of incomplete
water removal and sub-optimally polymerized resin Material and methods
[14]. Concern about accelerated degradation of the
tooth-resin bonds of these adhesives exists, as this This study involved two experimental ‘mild’ 1-SEAs
permeability may increase the hydrolytic degra- (exp-Eth and exp-Ac) containing both a carboxylate
dation and leaking of resin components [15–17]. and phosphate-based functional monomer (Table 1)
More recently, complex processes of phase separ- [1]. Both adhesives had a similar composition, but
ation have been shown to occur in one-component differed in solvent (ethanol and acetone, respect-
HEMA-free 1-SEAs [18]. This phase separation leads ively). Their application procedure involved strong
to the formation of multiple water droplets that can air-drying before light curing.
become entrapped in the adhesive layer upon light By adding a hydrophobic bonding agent (UB;
curing. from Unifil Bond, GC, Tokyo, Japan) after the
Van Meerbeek et al. [1,19] have proposed a application of exp-Eth or exp-Ac (which was not
classification of contemporary adhesives, based on light-cured, thereby serving as a self-etching
the adhesion strategy and application procedure. As primer), experimental 2-SEAs were formed (2-
differences regarding bonding effectiveness have SEA/exp-Eth and 2-SEA/exp-Ac). Similarly, by
been found between different adhesive categories, adding a preceding etching step with 35% H3PO4
this classification can also serve as a qualitative (Scotchbond etchant, 3M ESPE) and the same
classification of adhesives. In general, 3-step hydrophobic bonding agent, 1-SEAs were trans-
etch&rinse adhesives (3-E&R) have been found to formed into three-step etch&rinse adhesives (3-
perform better than 2-step etch&rinse adhesives E&R/exp-Eth and 3-E&R/exp-Ac).
(2-E&R); similarly, research has reported better As a control group, commercial adhesives
results for two-step self-etch adhesives (2-SEAs) representing 1-SEAs (iBond; Heraeus-Kulzer,
than for 1-SEAs [4–6,9]. Regarding durability, Hanau, Germany), 2-SEAs (Clearfil SE Bond;
 Extension of simplified adhesive into multi-step adhesive
Table 1 List of adhesives investigated along with their composition and application procedure.
Adhesive Classa Manufacturer Compositionb Application
Experimental exp-Eth 1-SEA GC, Tokyo, Japan 4-MET, phA-m, DMA, ethanol, water, filler, Apply adhesive to the entire surface with a
adhesives photoinitiator, stabilizer (pHw2) disposable applicator
Keep dentin wet (shiny surface) with
adhesive for at least 10 s; Strongly air-dry
Light-cure for at least 10 s
exp-Ac 1-SEA 4-MET, phA-m, DMA, acetone, water, filler, Idem as above
photoinitiator, stabilizer (pHw2)
2-SEA/exp-Eth 2-SEA Primer: exp-Eth. Bonding: UDMA, HEMA, Apply adhesive to the entire surface with
TEGDMA, photo-initiator (bonding agent of disposable applicator
Unifil Bond, GC) Keep dentin wet with adhesive for at least
10 s.; strongly air-dry; do not light-cure
Apply bonding with disposable applicator;
gently air-dry
Light-cure for 20 s
2-SEA/exp-Ac 2-SEA Primer: exp-Ac. Bonding: UDMA, HEMA, Idem as above
TEGDMA, photo-initiator (bonding agent of
Unifil Bond, GC)
3-E&R/exp-Eth 3-E&R Etchant: 35% phosphoric acid. Primer and Apply etchant for 15 s, rinse thoroughly and
bonding: see 2-SEA/exp-Eth gently air-dry
Apply adhesive to the entire surface with
disposable applicator
Keep dentin wet with adhesive for at least
10 s; strongly air-dry; do not light-cure
Apply bonding with disposable applicator;
gently air-dry
Light-cure for 20 s
3-E&R/exp-Ac 3-E&R Etchant: 35% phosphoric acid. Primer and Idem as above
bonding: see 2-SEA/exp-Ac
Control iBond 1-SEA Heraeus-Kulzer, UDMA, 4-MET, glutaraldehyde, acetone, Apply three layers of adhesive; wait for
adhesives Hanau, Germany water, photo-initiator, stabilizer 30 s, while slightly agitating
Gently air-dry for few seconds
Light-cure for 20 s
Clearfil SE Bond 2-SEA Kuraray, Osaka, Primer: 10-MDP, HEMA, hydrophilic DMA, Apply primer for 20 s; gently air-dry
Japan photo-initiator, aromatic tertiary amine, Apply bonding agent Light-cure for 20 s
water. Bonding: 10-MDP; Bis-GMA, HEMA,
Hydrophobic DMA, photo-initiator,
aromatic tertiary amine, silanated col-
loidal silica
(continued on next page)

535
536 K.L. Van Landuyt et al.

Kuraray, Osaka, Japan) and 3-E&R adhesives

glycidyl methacrylate; HEMA, 2-hydroxyethyl methacrylate; DMA, dimethacrylate; phA-m, phosphoric acid ester monomer; GPDM, glycerol phosphate dimethacrylate; PAMM, phtalic acid
Apply etchant for 15 s, rinse thoroughly and

Abbreviation of monomers in numerical/alphabetical order: 4-MET, 4-methacryloxyethyltrimellitic acid; 10-MDP, 10-methacryloyloxydecyl dihydrogen phosphate; Bis-GMA, Bisphenol-
(Optibond FL; Kerr, Orange, Ca, USA) were tested
as well.

Apply primer for 15 s; gently air-dry

Apply bonding agent, gently air-dry
mTBS-Testing

Human third molars (gathered following informed

consent approved by the Commission for Medical

Italicized words refer to difference in either composition or application procedure of the experimental adhesive as compared to the Exp-Eth master adhesive.
Ethics of KU-Leuven) were used within 1 month

Light-cure for 30 s
after extraction. They were stored in 0.5% chlor-
gently air-dry

amine/water at 4 8C until used. To prepare dentin

Application

samples, the occlusal crown third was removed

with a diamond saw (Isomet 1000, Buehler, Lake
Bluff, IL, USA), thereby exposing a flat mid-coronal
dentin surface. A bur-cut smear layer was produced
by removing a thin layer of the surface using a
Micro-Specimen Former (University of Iowa, Iowa
photoinitiator, ethanol, water. Bonding:
thickener. Primer: HEMA, GPDM, PAMM,

TEGDMA, UDMA, GPDM, HEMA, bis-GMA,

Etchant: 37.5% phosphoric acid, silica

City, IA, USA), equipped with a high-speed regular-

grit (100 mm) diamond bur (842, Komet, Lemgo,
Germany). For enamel, a flat surface was ground
using the same bur at the buccal and lingual surface
of a tooth. After application of the experimental
and control adhesives according to the instructions
filler, photoinitiator

listed in Table 1, dentin was built up without delay

monoethyl methacrylate; TEGDMA, triethylene glycol dimethacrylate; UDMA, urethane dimethacrylate.

using Gradia Direct Anterior (GC).

After storage overnight in distilled water (37 8C),
Compositionb

rectangular sticks (2!2 mm wide; 8–9 mm long)

were sectioned perpendicular to the adhesive–tooth
interface using the Isomet saw. Only the four central
sticks were used to eliminate substrate regional
variability [24,25]. The sticks were trimmed at
the interface into an hourglass shape (diameter of
Kerr, Orange, CA,

G1.1 mm) using the MicroSpecimen Former,

Manufacturer

equipped with a fine-grit (30 mm) diamond

(5835KREF, Komet) in a high-speed handpiece
According to classification proposed by Van Meerbeek et al. [1].

under air/water coolant. The specimens were

USA

fixed to Ciucchi’s jig with cyanoacrylate glue

(Model Repair II Blue, Dentsply-Sankin, Ohtawara,
Japan) and stressed in tension at a crosshead speed
of 1 mm/min using a universal testing device (LRX,
3-E&R
Classa

Lloyd, Hampshire, UK). The mTBS was derived by

dividing the imposed force at the time of fracture by
the bond area (mm2). When a specimen failed during
processing (pre-testing failure), the mTBS was set at
Optibond FL

0 MPa [26,27]. One-way ANOVA and the Tukey HSD

Adhesive

multiple comparison test were employed to evalu-

ate the statistical differences in mTBS between the
Table 1 (continued)

different application methods and experimental

and control 1-SEAs, 2-SEAs and 3-E&R adhesives
(aZ0.05). The mode of failure was determined with
a stereomicroscope at 50! magnification.
Representative dentin and composite mTBS-
fracture planes, exhibiting the most frequently
observed failure mode, and a mTBS close to
b
a

the mean were processed for field-emission gun

Extension of simplified adhesive into multi-step adhesive 537

scanning electron microscopy (Feg-SEM; Philips Results

XL30, Eindhoven) using common specimen proces-
sing described previously [28]. The micro-tensile bond strengths of exp-Eth and
exp-Ac varied significantly over the three differ-
TEM interface characterization ent application protocols (Figs. 1 and 2). With
regard to enamel, each additional application
Each adhesive was applied to bur-cut dentin step increased the bond strength. However, this
according to the application procedure mentioned increase was significant only when the exper-
in Table 1. The specimens were processed for TEM imental 1-SEAs were extended to three-step
according to the procedure described in detail by etch&rinse adhesives. For dentin, the bond
Van Meerbeek et al. [29]. Non-demineralized and strength increased when a 2-SEA protocol was
lab-demineralized (10% formaldehyde–formic acid followed. This rise in mTBS, however, was not
for 36 h) ultrathin sections were cut (Ultracut UCT, significant. When dentin was etched prior to the
Leica, Vienna, Austria), and examined unstained application of primer (1-SEA) and bonding (UB),
and positively stained (5% uranyl acetate for there was a significant drop in bond strength of
20 min/saturated lead citrate for 3 min) using TEM both exp-Eth and exp-Ac.
(Philips CM10, Eindhoven, The Netherlands). In Among the control adhesives (Fig. 2), iBond
order to reveal the defects in acid-etched hybrid performed significantly worse than the others. On
layers, additional specimens of Optibond FL and enamel, Optibond FL achieved significantly higher
3-E&R/exp-Ac stained with 50 wt% ammoniacal bond strengths than iBond and Clearfil SE Bond.
silver nitrate solution were prepared according to On dentin, however, mTBS of Optibond FL was
a nanoleakage detection protocol previously not significantly different from Clearfil SE Bond.
described by Tay et al. [14]. Significant differences were observed between

Figure 1 Box-whisker plot denoting the mean mTBS (diamonds) and median mTBS (line in bar) to enamel (top) and to
dentin (bottom). The bar indicates the lower and upper quartile and the whiskers define minimum and maximum. On the
right, the mean mTBSs with standard deviation are shown (mean (standard deviation); n, total number of specimens; ptf,
pre-testing failure) to enamel (top) and to dentin (bottom). One-way ANOVA and Tukey HSD multiple comparison test
were employed to evaluate statistical differences between the different application methods of exp-Ac and exp-Eth.
A separate test was performed for enamel and dentin.
538 K.L. Van Landuyt et al.

Figure 2 Micro-tensile bond strength results for enamel (top) and dentin (bottom). Bars denote mean mTBS and
whiskers define standard deviation. One-way ANOVA and Tukey HSD multiple comparison test were employed to
evaluate the statistical differences between the commercial and experimental 1-SEAs, 2-SEAs and 3-E&R adhesives. A
separate test was performed for enamel and dentin. Different letters inside the boxes signify statistical differences.

the experimental 2-SEAs and C-SE and between the ving enamel fracture. As for dentin, all adhesives
experimental 3-E&Rs and Optibond FL (Fig. 2). failed mixed-adhesively, except for 3-E&R/exp-Eth
Failure analysis on enamel revealed a predomi- and 3-E&R/Ac, which all failed completely adhe-
nantly mixed adhesive failure pattern for all sively (Fig. 3).
adhesives, sometimes involving cohesive fracture TEM observations of dentin showed the
of enamel or composite (Fig. 3). 3-E&R/exp-Eth and formation of a hybrid layer with different charac-
3-E&R/Ac exhibited a mixed failure usually invol- teristics for each class of adhesive (Figs. 4–6).
Extension of simplified adhesive into multi-step adhesive 539

Figure 3 Field-emission gun scanning electron photomicrographs showing fractured dentin specimens after micro-
tensile bond strength testing (top pictures: dentin side; bottom pictures: composite side). (a and b) Feg-SEM overview
photomicrograph of fractured surfaces with 2-SEA/exp-Ac (a, dentin side; b, composite side), revealing a typical mixed
failure involving interface, adhesive resin and composite. Note the air-bubbles in the composite. (c and d) Detail by
magnification of a and b, respectively, localized at the interfacial failure, showing the interaction of 2-SEA/exp-Ac with
dentin. Note that the dentinal tubules remained sealed with smear plugs. As collagen fibrils can be observed both on the
dentin and composite side, the failure must have occurred at the bottom of the small hybrid layer formed by 2-SEA/exp-
Ac. Fractured specimens of exp-Eth, exp-Ac and 2-SEA/exp-Eth (not displayed) did not substantially differ from this one.
(e and f) Feg-SEM overview photomicrograph of fractured surfaces with 3-E&R/exp-Ac, showing that the failure was
chiefly interfacial, although some remnants of adhesive resin can be seen. (g and h) Details by magnification of e and f,
respectively. Note the widened tubules orifices, and the broken resin tags, both on the dentin and composite side.
Again, collagen on the dentin and composite side indicates failure at the bottom of the hybrid layer (Ar, adhesive resin;
Coll, collagen; C, composite; Dt, dentinal tubule; Hy, hybrid layer; Rt, resin tag; Sp, smear plug).

All self-etch adhesives in this study created a hybrid practice the choice between a simplified and a
layer with a thickness of 0.5–1 mm, in which conventional, more elaborate adhesive. Such a
hydroxyapatite crystals could still be found universal adhesive could be adapted to the clinical
(Figs. 4 and 5a–c). When dentin was etched, a situation (e.g. deciduous teeth, child, available
hybrid layer of 3 mm that was completely devoid of time, etc.) and would eliminate the need for
hydroxyapatite and distinct resin tags was formed stocking several adhesives. However, the merit of
(Fig. 5d–f). A similar but overall thicker hybrid layer a system that consists of a basic priming solution
(5 mm) was produced by Optibond FL (Fig. 6). All that can be used both as an individual adhesive and
tested 1-SEAs showed droplets in their adhesive as part of a 2-SEA and 3-E&R depends on the
layers (Fig 4). However, no droplets were found in usefulness of adding application steps. This study
the 2-SEAs or the 3-E&Rs (Fig. 5). Applying a clearly showed that the tested experimental one-
hydrophobic bonding after application of exp-Eth step self-etch adhesives could not be extended
or exp-Ac enlarged the thickness of the adhesive without suffering the consequences. Even though
layer from 10 to 50 mm. After silver-staining, the the application of a hydrophobic bonding onto the
hybrid layer of 3-E&R/exp-Ac exhibited a spot-like non-cured 1-SEAs yielded higher bond strengths
pattern of nanoleakage, whereas Optibond FL (although not significant) both to enamel and to
intermittently exhibited dense deposits of silver dentin and a prior etching step significantly
throughout the hybrid layer (Fig. 6). increased the bond strength to enamel, the bond
strength to dentin was influenced adversely by this
preceding etching step. Therefore, the hypothesis
Discussion that adding an application step improves the
bonding effectiveness must be rejected when the
A 1-SEA that can be extended with a corresponding bond strength to dentin is regarded, but accepted
gain in bonding effectiveness would give a dentist in for bond strengths to enamel.
540 K.L. Van Landuyt et al.

Figure 4 Transmission electron photomicrographs of exp-Eth and exp-Ac on dentin. No morphological difference
could be observed between both adhesives. (a) Non-demineralized not-stained TEM showing an overview of the
interface of exp-Ac with dentin. Note the distinct oxygen inhibition layer on top of the thin adhesive layer (7–12 mm).
Strong air-blowing the adhesive before light curing reduced the number of droplets in the adhesive layer considerably.
(b) Non-demineralized not-stained TEM of the hybrid layer created by exp-Ac on smear-layer covered dentin. Dentin was
only partially demineralized, and hydroxyapatite crystals remain scattered throughout the shallow (0.5–1 mm) hybrid
layer. (c) Non-demineralized not-stained TEM of exp-Eth revealing small droplets in the adhesive layer, in spite of strong
air-blowing the adhesive after application. (d) Demineralized, stained TEM of the submicron hybrid layer formed by exp-
Eth (Ar, adhesive resin; C, composite; Hy, hybridized layer; O2-I, oxygen inhibition layer; Ud, unaffected dentin).

In this study, no statistical differences were applied for the two experimental 1-SEAs. TEM and
found between the two experimental 1-SEAs SEM observations revealed that not all droplets had
adhesives, exp-Eth and exp-Ac (Fig. 1). Even when been removed, but their number had been drasti-
these adhesives were applied according to a 2-SEA cally decreased. When iBond was applied according
protocol or a 3-E&R protocol, the obtained mTBS to the manufacturer’s instructions, the adhesive
was similar within each application group. Any layer also contained droplets.
effect of the solvent on the bonding effectiveness On enamel, all 3-E&R adhesives obtained higher
was hence excluded. This is in accordance with an bond strengths than the self-etch adhesives.
earlier study regarding the phase separation However, this increase in bond strength is only
phenomenon in these 1-SEAs. This study did show significant when compared to the 1-SEAs. The
a distinct effect of the solvent on the course of the higher bond strengths for acid-etched enamel can
phase-separation reaction, but no effect on the be explained by the more micro-retentive enamel
bond strength [18]. As a consequence, the second surface obtained when enamel is etched with
part of the null hypothesis stating that there is no phosphoric acid as compared to when enamel is
difference between use of ethanol or acetone must etched by the self-etch adhesive. The self-etch
be accepted. Nonetheless, acetone is preferred as a adhesives in this study belong to the category of
solvent medium, because of the better hydrolytical mild self-etch adhesives with a pH of approximately
stability of the functional monomers in acetone 2. Several authors have reported that mild self-etch
than in ethanol (unpublished data). adhesives only shallowly demineralize enamel,
HEMA-free 1-SEAs are prone to phase-separation resulting in a very thin micro-retentive pattern
reactions, giving rise to multiple droplets in the without formation of distinct macro- and micro-
adhesive layer, which can get entrapped upon light resin tags [30,31] This ill-defined etching pattern
curing [18]. In order to reduce the number of has been associated with a lower bond strengths
droplets in the adhesive and to remove water from [32–35]. An additional factor influencing the bond
the adhesive, a strong air-blowing protocol was strength is the short application time, which was
Extension of simplified adhesive into multi-step adhesive 541

Figure 5 Transmission electron photomicrographs of 2-SEA/exp-Eth and 2-SEA/exp-Ac (a, b and c); and 3-E&R/exp-
Eth and 3-E&R/exp-Ac (d–f) on dentin. (a) Non-demineralized, not-stained TEM of the hybrid layer of 2-SEA/exp-Ac. No
droplets could be found in the adhesive layer. The morphological appearance of the hybrid layer is similar to that of exp-
Ac, which is consistent. The adhesive layer, however, is much thicker (up to 50 mm) than the one of exp-Ac (not in
picture visible). (b) Non-demineralized not-stained TEM of 2-SEA/exp-Eth, showing a partially demineralized hybrid
layer. (c) Demineralized, stained TEM of 2-SEA/exp-Ac showing a shallow hybrid layer, similar to that in Fig. 4d. (d) Non-
demineralized, not stained TEM giving an overview of 3-E&R/exp-Ac on dentin. A hybrid layer that is characteristic for
phosphoric-acid etching is formed and the dentinal tubules are sealed by resin tags. (e) Non-demineralized, not stained
TEM of 3-E&R/exp-Ac, showing the hybrid layer of approximately 3 mm, which is rather small for phosphoric-acid etched
dentin. After air-drying, the demineralized collagen network collapsed, but exp-Ac must have been incapable of re-
expanding it completely. (f) Demineralized, stained TEM of the hybrid layer created by 3-E&R/exp-Eth. Note the
accumulation of remaining silica filler particles from the etching gel on top of the hybrid layer, which can also be seen in
d and e (Ar, adhesive resin; Hy, hybridized layer; Rt, resin tag; Ud, unaffected dentin).

only 10 s for the experimental adhesives. However Transforming a 1-SEA into a 2-SEA did yield
difficult to compare, the longer priming time of 20 higher bond strengths although this effect was not
or 30 s for Clearfil SE Bond and iBond, respectively, statistically significant. Several reasons for this
did not involve higher bond strengths. moderate rise in bond strength may be considered.

Figure 6 Transmission electron photomicrographs of 3-E&R/Exp-Ac (left) and Optibond FL (right) on dentin, after
silver nitrate staining. (a) TEM of 3-E&R/Exp-Ac on dentin showing a spotted pattern of nanoleakage throughout the
whole hybrid layer, which could be seen in all samples. (b) TEM of Optibond FL showing a similar hybrid layer as 3-
E&R/Exp-Ac. However, only intermittently, dense deposits of silver were observed in Optibond FL. Whereas some parts
of the hybrid layer exhibited nanoleakage (in this image), other locations were devoid of silver-staining (not in this
image) (Ar, adhesive resin; Hy, hybridized resin; Rt, resin tag; Ud, unaffected dentin).
542 K.L. Van Landuyt et al.

The most plausible explanation must be found in obtained significantly higher bond strengths than
the different proportional composition of one-step the two custom-made 2-SEAs and than all 1-SEAs.
and two-step adhesives. Although one- and two- Moreover, Clearfil SE Bond performed equally well
step adhesives generally contain the same com- as the commercial 3-E&R Optibond FL on dentin.
ponents (they both contain functional monomers, These observations are in agreement with the
cross-linking monomers, solvent, inhibitors and results of other, both in vitro and in vivo studies
activators), the amounts of ingredients applied on [4,9,44–46]. Its good performance on dentin can be
the tooth surface differ considerably among one- explained by its specific and adapted composition
and two-step adhesives. Whereas 2-SEAs consist of and the use of the functional monomer 10-MDP,
a pure priming solution containing only functional which has been shown to exhibit high chemical
etching monomers dissolved in organic solvent and interaction capacity to hydroxyapatite [47].
water, and a solvent-free bonding containing When dentin had been etched prior to the
hydrophobic cross-linking monomers (such as application of 1-SEA and hydrophobic bonding, a
UDMA, TEGDMA), 1-SEAs are complex mixtures of decrease in bond strength was observed. Additional
both these hydrophobic and hydrophilic ingredi- phosphoric acid etching even negated the beneficial
ents. Usually, one-component 1-SEAs contain acidic effect on the bonding effectiveness of the appli-
functional monomers dissolved in high concen- cation of an additional hydrophobic bonding agent.
trations of organic solvent and/or water blended 3-E&R/exp-Eth and 3-E&R/exp-Ac exhibited signifi-
with hydrophobic cross-linking monomers. As the cantly lower bond strengths than Optibond FL. This
solvent and functional monomers usually make up drop in bond strength to dentin must be attributed
almost 50% of the adhesive, the concentration of to suboptimal infiltration of the demineralized
hydrophobic monomers is drastically reduced. Since collagen network and subsequent poor adaptation
the mechanical strength of the adhesive is mainly of the bonding resin to the collagen fibrils. TEM and
provided by the polymerization of cross-linking nanoleakage observations confirmed the formation
monomers, relatively less hydrophobic monomers of an inferior and porous hybrid layer by 3-E&R/exp-
are available on the tooth surface after application Ac. Even though some nanoleakage could be
of the 1-SEA, which impairs the bond strength observed locally in the hybrid layer of Optibond FL
[34,36]. When the 1-SEA was applied according to a under the form of dense silver deposits, the whole
2-SEA protocol, the additional application of the hybrid layer of Exp-Ac/E&R exhibited a consistent
hydrophobic bonding must have increased the spotted pattern, suggesting a porous hybrid layer
concentration of the hydrophobic monomers, that is prone to silver stain uptake (Fig. 6). This
which explains increased bond strength. In pattern is suggestive of inadequate resin infiltration
addition, by applying a hydrophobic bonding, the and poorly enveloped collagen fibrils. When the
uncured 1-SEA that was applied as primer was demineralized collagen network is not optimally
diluted, leading to a thicker and more uniform layer impregnated, the exposed collagen fibrils are not
with lower concentrations of retained water and protected by resin, making them susceptible to
solvent. Sites of remaining water and solvents are tensile forces [48]. On the longer run, hydrolytic
thought to weaken the adhesive layer [37–40]. breakdown may jeopardize the bonding effective-
Another explanation is the effect of HEMA in the ness [49]. SEM observations confirmed that failure
bonding agent on the phase separation in the 1-SEA. under tensile stress occurred adhesively at the
When this 1-SEA is not light-cured, HEMA in the bottom of the hybrid layer (Fig. 3). The dense silver
bonding counteracts the phase-separation reaction deposits that can be observed in the hybrid layer of
by bringing the adhesive’s ingredients back into Optibond FL are also an indication for poor resin
solution. This was also confirmed by SEM and TEM impregnation and gaps in the hybrid layer, but as
showing that 2-SEA/exp-Eth and 2-SEA/exp-Ac their appearance is less prevalent, the bond
did not exhibit droplets in their adhesive layer. As strength of Optibond FL was probably not influ-
any kind of flaws in the adhesive layer may act as enced to the same extent as the bond strength of
stress raisers, droplets are bound to affect the 3-E&R/exp-Eth and 3-E&R/exp-Ac.
bond strength adversely [41]. Absence of droplets This suboptimal resin infiltration can be
in the experimental 2-SEAs is likely to lead to accounted to different causes [50]. As the major
higher bond strengths. Furthermore, the beneficial function of primer solutions in 3-E&R is to ensure
effect of a thicker adhesive layer in the 2-SEAs sufficient wetting of the exposed collagen fibrils, a
may also explain the slight increase in bond good primer is usually characterized by high
strength [42,43]. wetting properties and by low viscosity and
Regarding the bonding effectiveness of Clearfil usually contains low-molecular agents, enabling
SE Bond to dentin, this commercial adhesive good diffusion [1,49,51]. 1-SEAs in this study that
Extension of simplified adhesive into multi-step adhesive 543

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