Effect of Saliva Contamination and CleansingSolutions on the Bond Strengths of Self-EtchAdhesives to Dentinjerd_374 402..410

HUMA SHEIKH, BDS, MS*

HARALD O. HEYMANN, DDS, MED†

EDWARD J. SWIFT, Jr., DMD, MS‡

THOMAS L. ZIEMIECKI, DDS, MS§

ANDRÉ V. RITTER, DDS, MS¶

ABSTRACT

Purpose: This study determined the effect of saliva contamination and cleansing solutions onmicrotensile bond strengths of self-etch adhesives to dentin.

Materials and Methods: Seventy-five human molars were ground flat to expose mid-coronaldentin and randomly assigned to five groups (N = 15): no contamination, saliva contaminationwithout cleansing, saliva and cleansing with water, saliva and cleansing with 2% chlorhexidine,and saliva and cleansing with 5% sodium hypochlorite. One-third of the specimens in eachgroup of 15 were bonded with Adper Prompt L-Pop (all-in-one self-etch adhesive; 3M ESPE,St. Paul, MN, USA), one-third with Adper Easy Bond (all-in-one self-etch adhesive; 3M ESPE),and one-third with Clearfil SE Bond (self-etch primer system; Kuraray America, New York,NY, USA). Specimens were restored with composite and processed for microtensile bondstrength testing (5–6 rods/tooth).

Results: Mean bond strengths ranged from 17.3 MPa for Adper Prompt L-Pop after watercleansing to 69.3 MPa for Clearfil SE Bond after water cleansing. For all three adhesives, therewas no statistically significant difference in bond strengths between the saliva contaminatedgroup, the cleansing groups, and the no contamination groups.

Conclusions: Neither saliva nor the cleansing solutions adversely affected bond strengths of theself-etch adhesive systems.

CLINICAL SIGNIFICANCESaliva contamination of dentin does not seem to adversely affect bonding with self-etch adhe-sive systems. These results should be considered preliminary and need confirmatory studiesbefore conclusive recommendations can be made for clinical practice.

(J Esthet Restor Dent 22:402–411, 2010)

*Graduate student, Department of Operative Dentistry, University of North Carolina, Chapel Hill, NC, USA†Professor, Department of Operative Dentistry, University of North Carolina, Chapel Hill, NC, USA

‡Professor and chair, Department of Operative Dentistry, University of North Carolina, Chapel Hill, NC, USA§Associate professor, Department of Prosthodontics, University of North Carolina, Chapel Hill, NC, USA

¶Associate professor, Program Director, Graduate Operative Program, Department of Operative Dentistry,University of North Carolina, Chapel Hill, NC, USA

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I N T R O D U C T I O N

The effect of saliva contamina-tion of the tooth on dentin

bonding has been investigated, butthe results are somewhat inconclu-sive. Several studies report thatsaliva contamination has a detri-mental effect on resin bonding todentin.1–3 In one study, reapplica-tion of the adhesive after drying orrinsing off the saliva reestablishedbond strengths to control levels.1

Two studies suggest that completedrying of the saliva-contaminatedsurface should be avoided, as thismay decrease bond strengths.4,5

Others have reported that adhe-sives perform remarkably welleven when the saliva is notremoved after acid-etching,4,6,7

which might be explained by thehigh percentage of hydrophilic sol-vents these agents contain, such asacetone or alcohol.8 Hydrophilicbonding agents may be attractedby the saliva moisture, promptlyspreading upon moist dentin ordisplacing the adsorbed saliva.9

Therefore, the outcome may berelated not only to the contami-nant but also to the type ofadhesive used.

When saliva contamination of thepreparation occurs after acid-etching in anticipation of using anetch-and-rinse adhesive, phosphoricacid can be briefly reapplied andrinsed to effectively cleanse thesite.5 However, the same strategycould be detrimental to the

performance of self-etch adhesives,as it has been shown that pretreat-ment of dentin with phosphoricacid significantly reduces bondstrengths of self-etch adhesives.10

Given the paucity of studies on theeffect of saliva contamination andof different cleansing solutions ondentin bond strengths when con-temporary self-etch adhesivesystems are used, the purpose ofthis study was to examine theeffect of saliva contamination anddifferent cleansing solutions on thebond strengths of self-etch adhe-sives to dentin. Water, chlorhexi-dine, and sodium hypochloritewere used as cleansing agents. Thenull hypotheses tested were (1) thatsaliva contamination has no effecton dentin bond strengths of self-etch adhesive systems, and (2) thatrinsing saliva-contaminated dentinwith water, chlorhexidine, orsodium hypochlorite has no effecton the dentin bond strengths ofself-etch adhesive systems.

M AT E R I A L S A N D M E T H O D S

A pilot study using 10 extractedintact human molars determinedthat 75 teeth would be required topower the study at 80%. Seventy-five intact human molars were col-lected and stored in a solution of0.5% chloramine trihydrate at4°C. The occlusal surfaces of thespecimens were sectioned to exposemid-coronal dentin and polishedusing 600-grit silicon carbide paper

under water to create a uniformsmear layer.11 Peripheral enamelwas removed using diamond rotaryinstruments in water-cooled high-speed handpiece. The specimenswere randomly divided into fivegroups of 15 specimens each:

1. Group A—Dentin was not con-taminated (positive control)

2. Group B—Dentin was contami-nated with human saliva for5 seconds (no cleansing,negative control)

3. Group C—Dentin was contami-nated with human saliva for 5seconds and the contaminatedsurface was rinsed using dis-tilled water for 2 seconds

4. Group D—Dentin was contami-nated with human saliva for 5seconds and the contaminatedsurface was cleansed using 2%chlorhexidine digluconate solu-tion (Cavity Cleanser, Bisco,Schaumburg, IL, USA) appliedby lightly scrubbing with amicrobrush for 5 seconds. Thenthe surface was rinsed with dis-tilled water for 2 seconds

5. Group E—Dentin was contami-nated with human saliva for 5seconds and the contaminatedsurface was cleaned with 5%sodium hypochlorite (ACROSOrganics, Somerville, NJ, USA)applied by lightly scrubbingwith a microbrush for 5seconds. Then the surface wasrinsed with distilled water for2 seconds.

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The specific cleansing agents wereselected as they are readily avail-able to clinicians and would belikely choices for removingsaliva contamination.

One-third of the specimens in eachgroup (N = 5) were bonded with atwo-component all-in-one self-etchadhesive (Adper Prompt L-Pop,3M ESPE, St. Paul, MN, USA).One-third of the specimens in eachgroup (N = 5) were bonded withanother all-in-one self-etch adhe-sive system (Adper Easy Bond, 3M

ESPE), and one-third with a two-step self-etch adhesive system(Clearfil SE Bond, Kuraray Dental,Kurashiki, Japan). The composi-tions of all products used are listedin Table 1, and the flowchart ofthe experimental procedures isdepicted in Figure 1.

The adhesives were applied andlight-activated according to manu-facturers’ recommendations, afterthe surfaces were dried using acompressed air syringe at a dis-tance of 5 cm from the tooth

surface for 5 seconds. (A pilotstudy showed no significant differ-ence between air and blot dryingthe prepared tooth surface prior tobonding.) Adper Prompt L-Popwas applied with a rubbing motionfor 15 seconds, then gently butthoroughly air-dried to remove theaqueous solvent. A second coatwas then applied (no waiting timefor the second layer) and gentlybut thoroughly air-dried to removethe aqueous solvent. Finally, theadhesive was light cured for10 seconds.

TA B L E 1 . T R A D E N A M E S , M A N U FA C T U R E R S , L O T N U M B E R S , A N D C O M P O S I T I O N O F T H E P R O D U C T S U S E D I N T H I S

S T U D Y.

Product Composition

5% sodium hypochlorite solution 5% NaOClACROS OrganicsSomerville, NJ, USALOT A0248559Cavity cleanser 2% Chlorhexidine Digluconate (C22H30Cl2N10 · 2 C6H12O7)Bisco, Schaumburg, IL, USA2% Chlorhexidine SolutionLOT 0700007926Adper Prompt L-Pop methacrylated phosphoric esters, Bis-GMA, initiators based on camphorquinone,

stabilizers, water, 2-hydroxyethyl methacrylate (HEMA), polyalkenoic acid3M ESPE, St. Paul, MN, USALOT 332454Adper Easy Bond 2-hydroxyethyl methacrylate, bisphenol a diglycidyl ether dimethacrylate, water,

ethanol, phosphoric acid-6-methacryloxy-hexylesters, silane treated silica,1,6-hexanediol dimethacrylate, copolymer of acrylic & itaconic acid,(dimethylamino)ethyl methacrylate, camphorquinone,2,4,6-trimethlybenzoyldiphenylphosphine oxide

3M ESPELOT 299001

Clearfil SE Bond 10-methacryloyloxydecyl dihydrogen phosphate, 2-hydroxyethyl methacrylate,hydrophilic dimethacrylate, dl-Camphorquinone, N,N-Diethanol-p-toluidine, water,bis-phenol A diglycidylmethacrylate, silinated colloidal silica

Kuraray Dental, Kurashiki, JapanLOT 61832Filtek Supreme Plus silane treated ceramic, silane treated silica, bisphenol a polyethylene glycol diether

dimethacrylate, diurethane dimethacrylate bisphenol a diglycidyl ether methacrylate,triethylene glycol dimethacrylate, water

Universal RestorativeA2 Body Shade3M ESPELOT 20070802

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Adper Easy Bond was applied for atotal of 20 seconds by lightlyscrubbing with a microbrush,then dried for 5 seconds at a 5-cmdistance, and light-cured for10 seconds.

Clearfil SE Bond primer wasapplied and left for 20 seconds,then dried with gentle air flow at a5-cm distance. The Clearfil SEBond bonding agent was appliedand dispersed with mild air flow toevenly distribute on the surface of

the tooth. The adhesive was thenlight-cured for 10 seconds.

Composite resin (Filtek SupremePlus, 3M ESPE) was used incre-mentally to build-up the specimento a thickness of 4 mm. Each incre-ment was light-activated using ahigh intensity L.E.Demetron II(Kerr Corporation, Orange, CA,USA) unit operating at >800 mW/cm2 for 20 seconds. The specimenswere stored in distilled water for24 hours.

Each specimen was fixed in anepoxy resin block with stickywax. Specimens were sectionedmesiodistally using a water-cooledlow-speed Isomet 1,000 diamondmicro-slicing saw (Buehler, LakeBluff, IL, USA) to obtain 0.9-mmthick sections. The sections werefurther cut faciolingually to obtain6-mm-long, 0.9-mm-thick rods,with the dentin-composite inter-face located at the center. Eachspecimen had a cross-sectionalarea of 0.9 � 0.2 mm2, measuredwith a digital caliper (DigimaticIP67, Mitutoyo Co., Kawasaki,Japan). Between five and ninerods were obtained foreach tooth.

Each specimen was fixed to aCiucchi Jig (EZ-Test, Shimadzu,Kyoto, Japan) using acyanoacrylate-based adhesive(Zapit Base and Accelerator,Dental Ventures of America Inc,Corona, CA, USA). The specimenswere carefully placed on the jigso that the composite-dentininterface was exactly perpendicu-lar to the axis of the testingassembly. The microtensile bondstrengths of all specimens weretested using a universal testingmachine (EZ-Test, Shimadzu) witha crosshead speed of 1 mm/min.The bond strength (MPa) of eachspecimen was determined as thefailure load (N) divided by thecross-sectional area of thebonded interface.

Ground

Specimens

Contaminated

w/ saliva

n=60

Did not

contaminate

w/ saliva

(+ control)

n=15

Cleansed with:

(1) Water n=15

(2) CHX n=15

(3) NaOCl n=15

Did not

cleanse

(- control)

n=15

Rinsed (2)

and (3) with

water

TEST

Bonded 1/3 of the specimens

with Adper Prompt L-Pop, 1/3

with Adper Easy Bond, and 1/3

with Clearfil SE Bond.

Figure 1. Flowchart of experimental procedures.

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Data were plotted using MS Excel2007 (Microsoft, Mountain View,CA, USA) and analyzed using Stata10 (StataCorp, College Station,TX, USA). Microtensile bondstrength (MTBS) was the primaryoutcome (dependent variable),whereas surface treatment (salivacontamination and cleansing proto-col) and adhesive were the inde-pendent variables, with tooth, rod,area, width, and thickness as inde-pendent co-variables. Means wereadjusted for area and tooth toaccount for variations in measure-ments and potential differences inspecimens. We examined the corre-lations between MTBS and rodarea, width, and thickness usingpairwise correlation coefficients. Toexplore interactions and comparemeans between groups, data weresubjected to factorial analysis ofvariance (ANOVA) and Tukey’sHSD post hoc tests, where

indicated, with p = 0.05 signifi-cance level. MTBS results for rodsoriginating from the same toothwere pooled and averaged, so thatthe tooth was the unit of analysis.

R E S U LT S

A negative linear relationship wasnoted between MTBS and area(r = -0.8094, p < 0.0001, Figure 2),MTBS and width (r = -0.7658,p < 0.0001), and MTBS and thick-ness (r = -0.7658, p < 0.0001).These negative relationships indi-cate that, as expected, the smallerthe rod area, width, and thickness,the higher the MTBS. Using thetooth as the unit of analysis, therewas no significant interactionbetween adhesive and treatment(p = 0.076). The results of the fac-torial ANOVA and Tukey’s HSDpost hoc tests (mean MTBS andstandard deviations) are presentedin Table 2.

For all three adhesives, therewas no statistically significantdifference in mean MTBSbetween surface treatments,indicating that neither saliva con-tamination nor the cleansing pro-tocols negatively affected MTBSobtained with the positive control,uncontaminated dentin surface.When comparing adhesives withinsurface treatments, in general, theMTBS results were higher forClearfil SE Bond. Clearfil SE Bondhad significantly higher meanMTBS than Adper Easy Bond andAdper Prompt L-Pop for bothpositive and negative controltreatments. For the three cleansingprotocols (water, chlorhexidine,and sodium hypochlorite), MTBSobtained with Clearfil SE Bondwas similar to those obtainedwith Adper Easy Bond, andsignificantly higher than theMTBS obtained with AdperPrompt L-Pop.

D I S C U S S I O N

The purpose of this study was toexamine the effect of saliva con-tamination and of different cleans-ing agents on the dentin bondstrengths of self-etch adhesives tosaliva-contaminated dentin. Theresults showed that the testedadhesives were not affected bysaliva contamination and that alltested cleansing solutions, afterbeing rinsed with water, did notnegatively affect the bond strengthsof the adhesives tested. We

020

40

60

80

100

Mic

rote

nsile

Bond S

trength

.7 .8 .9 1 1.1

Rod area

Figure 2. Relationship between rod area and microtensilebond strength (r = -0.8094, p < 0.0001).

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therefore failed to reject the nullhypotheses tested.

Saliva contamination of tooth sur-faces and its impact on the bondstrengths of self-etch adhesivesystems are potentially importantclinical problems, with few andinconclusive studies addressingthem. Currently available optionsto manage saliva contamination oftooth surfaces remain inadequate.The use of phosphoric acid gel tocleanse the tooth surface aftersaliva contamination results inover-etching of the dentin surfaceof the tooth, resulting in significantreduction of the microtensilebond strengths of self-etchadhesive systems.10

Water is an obvious and simpleoption to address saliva contami-nation of a prepared tooth surface.The water syringe may be the firstdevice that many dentists reach forin anticipation of cleansing a con-taminated tooth surface. In a studyby Sattabanasuk and colleagues, asimilar hypothesis was tested.1 In

one of the test groups, the pre-pared dentin surface was contami-nated with saliva, rinsed withwater, and the self-etch adhesivewas reapplied. Bonding procedureswere performed according tomanufacturers’ directions, and acomposite resin was bonded ontothe surface of the prepared tooth.The results showed that rinsing thecontaminated tooth surface withwater reestablished the bondstrengths to a value similar to thatof no contamination at all.1 Ourstudy also showed that for AdperPrompt L-Pop, Adper Easy Bond,and Clearfil SE Bond, cleansingwith water did not adversely affectthe bond strengths.

We selected 2% chlorhexidinedigluconate solution as one of thecleansing agents for our study,given its established potential tomaintain or even strengthen themicrotensile bond strengths of self-etch adhesive systems.12 An in vivostudy by Carrilho and colleaguestested the hypothesis that chlo-rhexidine could be used to inhibit

the degradation of resin-dentinbonds by blocking the action ofmatrix metalloproteinases.12 Theseauthors showed that the syntheticprotease inhibitor, chlorhexidine,stabilized the bond strengths of thetreated dentin surfaces as com-pared with the untreated toothsurfaces.12 Our study concludedthat chlorhexidine does not haveany negative effect on the bondstrengths of self-etch adhesives,although we were also unable todetermine if chlorhexidine wouldimprove the bond strengths of self-etch adhesives. Additional long-term studies may have to be donein order to determine the long-termeffects of chlorhexidine on thebond strength of self-etchadhesive systems.

Sodium hypochlorite is anothercleansing alternative, given itspopular application as a bacterialreducing agent in intracanalpreparations.13 The fact that thepH of sodium hypochlorite ishighly alkaline (~11) makes it apotentially acceptable alternative

TA B L E 2 . M I C R O T E N S I L E B O N D S T R E N G T H M E A N S ( S D ) B Y A D H E S I V E A N D S U R FA C E T R E AT M E N T S ( N = 5 ; R E S U LT S

E X P R E S S E D I N M P a ) . *

Adhesive systems Surface treatments/cleansing agents

No contamination Saliva Water Chlorhexidine Sodium hypochlorite

Adper Prompt L-Pop 20.1 (7.9)aB 26.7 (14.1)aB 17.3 (4.6)aB 17.7 (4.3)aB 20.6 (8.7)aB

Adper Easy Bond 37.9 (4.4)aB 51.0 (9.2)aB 63.9 (13.8)aA 49.3 (12.0)aA 61.3 (11.3)aA

Clearfil SE Bond 60.5 (7.0)aA 72.0 (11.5)aA 69.3 (10.2)aA 62.5 (29.2)aA 54.8 (20.3)aA

*Same lowercase superscript letters indicate means that are not significantly different (p > 0.05) within rows (adhesives); same uppercasesuperscript letters indicate means that are not significantly different (p > 0.05) within columns (cleansing agents).

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as a cleansing agent, as it doesnot have etching potential andwould have substantial benefitover the use of acidic agents suchas phosphoric acid. These effectswere highlighted in a study byMountouris and colleagues whereit was demonstrated that sodiumhypochlorite has the potential todeproteinate the coronal dentinsurface without affecting the car-bonates and the phosphates.14

This reiterates the fact that thereis no dissolution of the mineralcontent of the tooth that takespart in the chemical adhesion tothe restorative material. Our studyshowed that sodium hypochloritefollowed by water rinsing doesnot have any detrimental effecton bond strengths of theadhesives tested.

The rationale for choosing AdperPrompt L-Pop, Adper Easy Bond,and Clearfil SE Bond as the self-etch adhesive systems is as follows.Adper Prompt L-Pop was chosenbecause its acidic pH (0.7) wouldallow us to observe the response ofa strongly acidic all-in-one typeself-etch adhesive to a contaminat-ing agent. Adper Easy Bond waschosen because it is less acidic,with a pH of 2.7 (technical infor-mation, 3M ESPE), and it was ofinterest for this study to observehow a comparatively less acidicadhesive would respond to a con-taminating agent. Clearfil SEBond is a two-step self-etch system,

has been used in many previousstudies as the “standard” self-etchsystem, and has a relatively mildpH (2.0).

Human saliva was used as the con-taminating agent rather than anartificial saliva or saliva substitute.Fresh whole human saliva is anacceptable substance in testingsaliva contamination and adsorp-tion.15 It was pertinent to thisstudy to make sure that there wasa “real” contaminating agent used,or otherwise run the risk of con-ducting a study that had little orno clinical significance.

To standardize the contaminant,the saliva was collected fromonly one person, a healthy26-year-old female, collected atfasting level early in the morning,before any oral hygiene regimen.Fasting-level saliva was collectedin order to provide less variabilityin pH of the saliva, as well asaltered electrolyte, enzyme, orprotein content seen after consum-ing a food or drink.16 Using a pHmeter, it was determined that thepH of this saliva was an averageof 7.4.

Typically, microtensile bondstrength uses individual specimensas the unit of analysis. One poten-tial problem with this approach isthat specimens originating from thesame tooth are not independentobservations. Additionally, dentin

from specimens obtained near thedentinoenamel junction is not thesame compositionally as that ofspecimens obtained near the pulp.Using individual specimens as theunit of analysis can therefore resultin biased data. To avoid thisproblem, in this study the toothwas used as the unit of analysis.MTBS values obtained from eachspecimen originating from thesame tooth are averaged.

Although this was not our intentwhen planning the study and theseresults cannot be generalized toother adhesives, we noticed a nega-tive correlation between the pH ofthe self-etch adhesive systems andthe MTBS data. This observationsuggests that less acidic self-etchadhesive systems tend to showhigher microtensile bond strengthsto dentin.

We also noted that, for all threeself-etch adhesive systems tested,there was a numerical increase inmicrotensile bond strengths whenapplied to saliva-contaminateddentin without any rinsing, whencompared with the positivecontrol (no contamination). Thisunexpected finding may suggestthat the tested self-etch adhesivesmay work acceptably in the pres-ence of saliva contamination. Fur-thermore, if the numerical resultsare considered, saliva might evenimprove dentin bond strengths.This suggests that there might be

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no need to cleanse the preparationsurface after contamination withsaliva prior to bonding with aself-etch adhesive. Simply dryingthe surface may suffice. It is pos-sible that the inherent acidity ofself-etch adhesive systems allowsthem to not only modify/penetratethe smear layer but also breakthrough the mucopolysaccharidesin the saliva and develop bondstrengths comparable with thoseobtained on a noncontaminateddentin surface. This observationcan possibly prove that, in a clini-cal situation, when saliva contami-nation of the prepared toothsurface occurs, it may be possibleto maintain bond strengths afterdrying the surface. There may beno need to cleanse with anyother agent.

As this was an in vitro study, theresults cannot be generalized tothe clinical setting. Although itmay be possible that some self-etch adhesive systems are notaffected by saliva contamination,as long as the tooth surface isdried, clinical studies are neededto confirm these results beforebroad generalizations can bemade. Also, as only three self-etchadhesive systems were tested, wecannot generalize our results forall self-etch adhesives availablein the market. Additionally, onlyone person’s saliva was used asthe contaminating agent, andnot every person’s saliva would

necessarily have the same effectsas the one used.

Because we rinsed the cleansingagents with water before proceed-ing with the bonding protocol, weare unable to determine the effectof the cleansing agents had theynot have been removed but onlydried before the application of theadhesives. We elected to rinsethe cleansing agents because weassumed that clinicians wouldrinse the cleansing agent off thecontaminated surface after it isapplied. Nevertheless, futureresearch could study theeffect of chlorhexidine andsodium hypochlorite assurface contaminants.

C O N C L U S I O N S

Under the conditions of this study,the following conclusions canbe made:

1. Saliva contamination of dentindid not adversely affect bondingwith the three self-etch adhesivesystems tested

2. Cleansing saliva-contaminateddentin surfaces with water, chlor-hexidine, or sodium hypochlo-rite had no negative effect onthe bond strengths of these self-etch adhesive systems.

D I S C L O S U R E A N D

A C K N O W L E D G E M E N T S

The authors have no personal orfinancial affiliation with any

company producing the productsused in this study that might beconsidered a conflict of interest inperforming this research.

The authors thank 3M ESPE fortheir support for this study, and3M ESPE, Bisco, and Kuraray forthe donation of materials.

R E F E R E N C E S

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2. Johnson ME, Burgess JO, Hermesch CB,Buikema DJ. Saliva contamination ofdentin bonding agents. Oper Dent1994;19(6):205–10.

3. Yoo HM, Oh TS, Pereira PN. Effect ofsaliva contamination on the microshearbond strength of one-step self-etchingadhesive systems to dentin. Oper Dent2006;31(1):127–34.

4. Fritz UB, Finger WJ, Stean H. Salivarycontamination during bonding procedureswith a one-bottle adhesive system.Quintessence Int 1998;29(9):567–72.

5. Park JW, Lee KC. The influence of sali-vary contamination on shear bondstrength of dentin adhesive systems.Oper Dent 2004;29(4):437–42.

6. el-Kalla IH, Garcia-Godoy F. Saliva con-tamination and bond strength of single-bottle adhesives to enamel and dentin.Am J Dent 1997;10(2):83–7.

7. Hitmi L, Attal JP, Degrange M. Influenceof the time-point of salivary contamina-tion on dentin shear bond strength of 3dentin adhesive systems. J Adhes Dent1999;1(3):219–32.

8. Rosa B. Bond strength of composite resinto contaminated enamel and dentin. PhDthesis, University of North Carolina,Chapel Hill; 2000.

9. Baier RE, Shafrin EG, Zisman WA. Adhe-sion: mechanisms that assist or impede it.Science 1968;162(860):1360–8.

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10. Torii Y, Itou K, Nishitani Y, et al. Effectof phosphoric acid etching prior to self-etching primer application on adhesion ofresin composite to enamel and dentin.Am J Dent 2002;15(5):305–8.

11. Pashley DH, Tao L, Boyd L, et al. Scan-ning electron microscopy of the substruc-ture of smear layers in human dentine.Arch Oral Biol 1988;33(4):265–70.

12. Carrilho MR, Geraldeli S, Tay F, et al.In vivo preservation of the hybrid layerby chlorhexidine. J Dent Res2007;86(6):529–33.

13. McGurkin-Smith R, Trope M, Caplan D,Sigurdsson A. Reduction of intracanal

bacteria using GT rotary instrumentation,5.25% NaOCl, EDTA, and Ca(OH)2.J Endod 2005;31(5):359–63.

14. Mountouris G, Silikas N, Eliades G.Effect of sodium hypochlorite treatmenton the molecular composition and mor-phology of human coronal dentin.J Adhes Dent 2004;6(3):175–82.

15. Gu F, Li Y, Zhou C, et al. Bacterial16S rRNA/rDNA profiling in the liquidphase of human saliva. Open Dent J2009;3:80–4.

16. Johansson I, Ericson T, Steen L. Studiesof the effect of diet on saliva secretionand caries development: the effect of

fasting on saliva composition of femalesubjects. J Nutr 1984;114(11):2010–20.

Reprint requests: Huma Sheikh, BDS, MS,School of Dentistry, University of NorthCarolina at Chapel Hill, 433 Brauer Hall,Chapel Hill, NC 27599, USA; Tel.: (919)966-2770; email: sheikhh@dentistry.unc.edu

This article is accompanied by commentary,“Effect of Saliva Contamination andCleansing Solutions on the Bond Strengthsof Self-Etch Adhesives to Dentin,” DanielL. Leonard, DDS, ABGD, DOI 10.1111/j.1708-8240.2010.00375.x.

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