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Year : 2012  |  Volume : 2  |  Issue : 1  |  Page : 15-18

A comparative analysis of sandblasted and acid etched and polished titanium surface on enhancement of osteogenic potential: An in vitro study

Department of Prosthodontics and Implantology, Meenakshi Ammal Dental College and Hospital, Chennai, India

Date of Web Publication24-May-2012

Correspondence Address:
Raj G Singh
3A/59, Rachna Vaishali, Ghaziabad, Uttar Pradesh- 201 010
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/0974-6781.96558

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Purpose: Establishment of titanium surface treatment and comparison of surface roughness and osteogenic potential of commercially pure titanium.
Materials and Methods: Twenty commercially pure grade 2 titanium disks of similar design and dimensions were divided into two groups. Smooth samples were assigned as control group (Group S). The second group (Group SLA) samples were sand-blasted and etched with different acids. Scanning Electron Microscopy (SEM) at ×500 was performed to observe the surface morphology. The surface roughness of samples was evaluated with surface profilometer. To evaluate the effect of surface treatment, samples from each group underwent cell culture study using human osteosarcoma osteoblast cell lines (HOS). SEM of one sample from each group was performed at ×500 to observe the cell morphology and cell attachment.
Results: Sand-blasted and acid-etched (SLA) surface was rougher in comparison to the smooth surface. SEM result of sand-blasted and acid-etched surface (Group SLA) showed that cell sheets were able to adhere inside the valleys suggesting excellent attachment osseointegration to rough surface.
Conclusion: This approach to develop sand-blasted and acid-etched surface was successful in uniform distribution of human osteosarcoma osteoblast cell lines (HOS) suggesting excellent osseointegration.

Keywords: Human osteosarcoma osteoblast cells, SLA, smooth surface

How to cite this article:
Singh RG. A comparative analysis of sandblasted and acid etched and polished titanium surface on enhancement of osteogenic potential: An in vitro study. J Dent Implant 2012;2:15-8

How to cite this URL:
Singh RG. A comparative analysis of sandblasted and acid etched and polished titanium surface on enhancement of osteogenic potential: An in vitro study. J Dent Implant [serial online] 2012 [cited 2023 May 31];2:15-8. Available from:

   Introduction Top

Titanium (Ti) is the implant material of choice for use in dental and orthopedic applications. [1],[2] The stable oxide that forms readily on titanium surfaces is documented to attributes to its excellent biocompatibility. [3],[4] However, it was also found that bone response to implant surface was dependent on the chemical and physical properties of titanium surfaces thereby affecting implant success. The surface characteristics of dental implants are considered to play an important role in their clinical success. One of the more important surface characteristics of implant is the surface topography or roughness. [5],[6],[7] The degree of roughness is determined either by the machining process during the preparation of the implant, or by the subsequent modification of the surface.

Bone formation around an implant is a complex process, and it is not fully understood. Apart from patient metabolism, physicochemical and topographical surface characteristics are some of the most influential factors in the improvement of osseointegration. [8],[9] The mechanical interlocking of micro and nano-irregularities with the tissue plays an important role in bioactivity results.

The present study optimizes a method of surface treatment for commercially pure titanium (Ti) and compared the human osteosarcoma osteoblast cell response to the smooth and new surface.

   Materials and Methods Top

Machining of samples

Approximately 2.5-mm thickness samples were cut from grade 2 commercially pure Titanium 6mm diameter rod (Central training institute, Chennai). To attach a fixture mount, a 1mm diameter hole was prepared at the circumference of the sample. Twenty samples were selected for the study. Samples were abraded with 500, 800, 1200 grit silica carbide paper for 1min subsequently. The samples underwent an ultrasonic cleaning procedure for 180 s using acetone to remove surface residues.

Grouping of samples

Twenty samples were selected for the study and divided into two groups: " Group S" (smooth surface) and "Group SLA" (sand-blasted and acid-etched).

Sample preparation

The samples were placed in a burn-out furnace at 100 o C for 1min to remove surface moisture. Samples were handled carefully after wearing latex gloves. The samples were held with tweezers taking care not to damage or contaminate the flat surface. The samples were attached to fixture mount and sandblasting was done with sand-blaster equipment [Figure 1]. Distance between nozzle and the sample was standardized at 2cm. Sand blasting was performed for 1min at a constant pressure of 4kg/cm 2 with 50 μm alumina (Al 2 O 3 ) particles. [10] Ultrasonic cleaning procedure was done using acetone for 180 s (twice) in order to remove any residual alumina particle. At room temperature each sample was dipped in three different acids, namely - 15% hydrofluoric acid (HF), 96% sulfuric acid (H 2 SO 4 ) and 37% hydrochloric acid (HCl). Each sample was attached to screw soldered with metal rod. Forty milliliter of each acid was transferred in a glass tumbler. Twenty percent sodium bicarbonate (Na 2 CO 3 ) solution and deionized water were housed separately. Acid etching was performed by dipping the samples in solutions following this sequence:

15% HF for 1min

Rinsed in deionized water for 5 s

20% Na 2 CO 3 for 30 s

96% H 2 SO 4 for 3min

Rinsed in deionized water for 5 s

20% Na 2 CO 3 for 30 s

37% HCl for 3min

Rinsed in deionized water for 5 s

20% Na 2 CO 3 for 30 s
Figure 1: Sand blasting of sample

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All the samples again underwent ultrasonic cleaning in acetone for 180 s to remove any residue of chemicals. All samples were kept in a burn out furnace at 400 o C for 2 min to kill all bacteria spores. The samples were transferred to glass vials that had been ultrasonically cleaned previously. Each vial had a minute perforation in the cap to allow steam entry during autoclaving. The vials with samples were autoclaved at 121°C, 16 min, 1 bar for 20min and followed by ultraviolet light for 48 h before cell culture test. Sample from each group underwent scanning electron microscopy (SEM) at 500× to evaluate surface morphology (Model S-3400, Hitachi, Japan). Surface roughness of samples was measured with surface profilometer (Horizon noncontact optical Profilometer, Burleigh Instruments, NY).

Cell culture study

Cell culture study was performed using procedure described by Anil Kumar et al.[11],[12] Human osteosarcoma osteoblast cells (HOS, National Centre for Cell Science, Pune, India) were cultured and maintained in Iscove's Modified Dulbecco's Medium (IMDM) supplemented with 5% fetal bovine serum (FBS) and 100 IU ml -1 penicillin and 100 μg ml-1 streptomycin in a CO 2 incubator (Nuaire, USA) at 37°C with 95% humidified atmosphere containing 5% CO2 . HOS cells were seeded at a density of 5 × 10 3 per test material and control cover slip [Figure 2]. Cells were seeded on cover slip to assess cell behavior during the test. Cells were incubated at 37 ± 2°C with test samples for 48 h, then medium was removed and cells were fixed in 3% glutaraldehyde for minimum 1 h. Samples were processed by dehydrating in graded alcohol (Ethanol), critical point drying (Hitachi, HCP-2), and gold sputtering followed by observation under a scanning electron microscope (Model S-3400, Hitachi, Japan) at 500×.
Figure 2: Trypsinization of the samples

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   Results Top

Surface morphology of the samples was evaluated using SEM at 15 kV acceleration voltage and magnification of 500×. SEM-revealed Sample S (control) showed parallel grooves with smooth surface. Irregular micro-pits were irregularly distributed over the surface [Figure 3]. Sample SLA (sand-blasted and acid-etched) showed irregular rough surface. Numerous elevations and depressions were present [Figure 4]. Surface profilometer (Horizon noncontact optical Profilometer, Burleigh Instruments, NY) result showed that sand-blasted and acid-etched samples were rougher than smooth samples [Table 1]
Figure 3: Sample S (×500)

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Figure 4: Sample SLA (×500)

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Table 1: Surface roughness

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Cell adhesion test result

Sample S (Control) at 500× revealed cells were irregularly present on the surface. Cells were loosely attached to the sample and irregular in shape [Figure 5]. Sample SLA (sand-blasted and acid-etched) at 500× showed homogenous spread of cells on the surface. Cells were able to penetrate into depressions suggesting excellent osseointegration [Figure 6].
Figure 5: Cells on sample S (control) at ×500

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Figure 6: Cells on sample SLA (sand-blasted and acid-etched) at ×500

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   Discussion Top

As further evidence of the importance of surface topography, numerous experimental animal studies have pointed towards a more rapid and stronger bone response to roughened surfaces than to smoother polished or machined surfaces. [13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23] An optimal roughness value of Sa 1.4 μm has even been suggested by Wennerberg et al.[16] However, a surface which is too rough has not been shown to result in a better bone response than intermediate or moderately rough surfaces. [16],[17],[22]

Surface topography has been considered by many workers to play a critical role in osseointegration. Davies [24] and Berglundh et al. [25] have postulated that a rougher surface is osteoconductive since it anchors the fibrin scaffold through which differentiating cells migrate towards the implant surface. Terms such as contact guidance and rugophilia have been used to describe the interaction of cells and tissues with textured surfaces. The former refers to the directional guidance provided by a substrate. Rugophilia literally means "rough-loving". Whereas some type of cells like fibroblasts will accumulate on smooth surfaces, others, such as macrophages prefer roughened surfaces.

Most implant surfaces exhibit mechanical integration providing resistance to shear forces but poor resistance in tension. On the other hand, chemical integration (i.e. osseocoalescence) provides good resistance to both shear and tensile forces and is the major thrust of future research in implant surface development. [26]

   Conclusion Top

Sand-blasted and acid-etched (SLA) samples were rougher in comparison to the control group. Group SLA showed homogenous spread of cell sheet on the prepared surface; cell sheets were able to penetrate into the pores and adhered inside suggesting excellent osseointegration.

   Acknowledgments Top

The author would like to thank Dr. E. Munirathnam Naidu, MDS and Dr. D. Sendhilnathan for their kind support and guidance. Author also thanks Dr. P.R. Anil Kumar (Implant Biology Section, Sri Chitra Institite, Kerala) for cell culture work.

   References Top

1.Branemark PI, Hansson BO, Adell R, Breine U, Lindstrom U, Hellan O, et al. Osseointegrated implants in the treatment of the edentulous jaw. Experience from a 10-year period. Scand J Plast Reconstr Surg 1977;16(Suppl. 1):7-127.  Back to cited text no. 1
2.Alberktsson T, Branemark PI, Hansson HA, Lindstrom J. Osseointegrated titanium implants. Requirements for ensuring a long-lasting, direct bone-to-implant anchorage in man. Acta Orthop Scand 1981;52:155-70.  Back to cited text no. 2
3.Stanford CM, Keller JC. The concept of osseointegration and bone matrix expression. Crit Rev Oral Biol Med 1991;2:83-101.  Back to cited text no. 3
4.Lausmaa J, Mattsson L, Rolander U. Chemical composition and morphology of titanium surface oxides. In: Williams JM, editor. Biomedical Materials. Pittsburgh, USA: Materials Research Society; 1986.  Back to cited text no. 4
5.Martin JY, Schwartz Z, Hummert TW, Schraub DM, Simpson J, Lankford J Jr, et al. Effect of titanium surface on proliferation, differentiation, and protein synthesis of human osteoblast like cells (MG63). J Biomed Mater Res 1995;29:389-401.  Back to cited text no. 5
6.Kasemo B. Biocompatibility of titanium implants: Surface science Aspect. J Prosthet Dent 1983;49:832-7.  Back to cited text no. 6
7.Puleo DA, Thomas MV. Implant surfaces. Dent Clin N Am 2006;50:323-38.  Back to cited text no. 7
8.Rosa AL, Beloti MM. Rat bone marrow cell response to titanium and titanium alloy with different surface roughness. Clin Oral Impl Res 2003;14:43-8.  Back to cited text no. 8
9.Cooper LF, Masuda T, Yliheikkilä PK, Felton DA. Generalization regarding the process and phenomenon of osseointegration: Part II- in vivo studies. Int J Oral Maxillofac Implants 1998;13:163-74.  Back to cited text no. 9
10.Lim YJ, Oshida Y, Andres CJ, Barco MT. Surface Characterizations of Variously Treated Titanium Materials. Int J Oral Maxillofac Implants 2001;16:333-42.  Back to cited text no. 10
11.Anil Kumar PR, Varma HK, Kumary TV. Rapid and complete cellularization of hydroxyapatite for bone tissue engineering. Acta Biomaterialia 2005;1:5 545-52.  Back to cited text no. 11
12.Anil Kumar PR, Varma HK, Kumary TV. Cell patch seeding and functional analysis cellularized scaffolds for tissue engineering. Biomed Mater 2007;1:48-54.  Back to cited text no. 12
13.Kasemo B. Biocompatibility of titanium implants: Surface science aspacts. J Prosthet Dent 1983;49:832-7.  Back to cited text no. 13
14.Brånemark PI. Osseointegration and its experimental background. J Prosthet Dent 1983;50:399-410.  Back to cited text no. 14
15.Albrektsson T, Jacobsson M. Bone-metal interface in osseointegration. J Prosthet Dent 1987;57:597-607.  Back to cited text no. 15
16.Wennerberg A, Ektessabi A, Albrektsson T, Johansson C, Andersson B. A 10 year follow-up of differing surface roughness placed in rabbit bone. Int J Oral Maxillofac Implants 1997;12:486-94.  Back to cited text no. 16
17.Wennerberg A, Hallgren C, Johansson C, Danielli S. A histomorphometric evaluation of screw-shaped implants each prepared with two surface roughnesses. Clin Oral Imp Res 1998;9:11-9.  Back to cited text no. 17
18.Buser D, Nydegger T, Hirt HP, Cochran DL, Nolte LP. Removal torque values of titanium implants in the maxilla of miniature pigs. Int J Oral Maxillofac Implants 1998;13:611-9.  Back to cited text no. 18
19.PiateIli A, Manzon L, Scarano A, Paolantonio M, Piatelli M. Histologic and histomorphometric analysis of the bone response to machined and sandblasted titanium implants: An experimental study in rabbits. Int J Oral Maxillofac Implants 1998;13:805-10.  Back to cited text no. 19
20.Baker D, London RM, O'Neal R. Rate of pull-out strength gain of dual-etched titanium implants: A comparative study in rabbits. Int J Oral Maxillofac Implants 1999;14:772-28.  Back to cited text no. 20
21.Cordiolli G, Majzoub Z, PiateUi A, Scarano A. Removal torque and histomorphometric investigation of 4 different titanium surfaces: An experimental study in rabbit tibia. Int J Oral Maxillofac Implants 2000;15:668-74.  Back to cited text no. 21
22.London M, Roberts FA, Baker DA, Rohrer MD, O'Neal RB. Histologic comparison of a thermal dual-etched implant surface to machined, TPS, and HA surface: Bone contact in vivo in rabbits. Int J Oral Maxillofac Implants 2002;17:369-76.  Back to cited text no. 22
23.Marinho V, Celletti R, Brachetti G, Petrone G, Minkin C, Piatelli A. Sand-blasted and acid-etched dental implants: A histological study in rats. Int J Oral Maxillofac Implants 2003;18:75-81.  Back to cited text no. 23
24.Davies JE. Mechanisms of endosseous integration. Int J Prosthodont 1998;11:391-401.  Back to cited text no. 24
25.Berglundh T, Abrahamsson I, Lang NP, Lindhe J. De-novo alveolar bone formation adjacent to endosseous implants. Clin Oral Impl Res 2003;14:251-62.  Back to cited text no. 25
26.Areva S, Aäritalo V, Tuusa S, Jokinen M, Lindén M, Peltola T. Sol-Gel-derived TiO 2 -SiO 2 implant coatings for direct tissue attachment. Part II: Evaluation of cell response. J Mater Sci Mater Med 2007;18:1633-42.  Back to cited text no. 26


  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]

  [Table 1]

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