JADA Continuing Education
The Influence of Zero-Value Subtraction on the Performance of Two Laser Fluorescence Devices for Detecting Occlusal Caries In Vitro
Jonas de A. Rodrigues, MSc, DMD, PhD,
Isabel Hug,
Michele B. Diniz, MSc,
Rita C.L. Cordeiro, MSc, PhD and
Adrian Lussi, DMD
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ABSTRACT
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Background. The aim of this study was to evaluate the influence of zero-value subtraction on the performance of two laser fluorescence (LF) devices developed to detect occlusal caries.
Methods. The authors selected 119 permanent molars. Two examiners assessed three areas (cuspal, middle and cervical) of both mesial and distal portions of the buccal surface and one occlusal site using an LF device and an LF pen. For each tooth, the authors subtracted the value measured in the cuspal, middle and cervical areas in the buccal surface from the value measured in the respective occlusal site.
Results. The authors observed differences among the readings for both devices in the cuspal, middle and cervical areas in the buccal surface as well as differences for both devices with and without the zero-value subtraction in the occlusal surface. When the authors did not perform the zero-value subtraction, they found statistically significant differences for sensitivity and accuracy for the LF device. When this was done with the LF pen, specificity increased and sensitivity decreased significantly.
Conclusions. For the LF device, the zero-value subtraction decreased the sensitivity. For this reason, the authors concluded that clinicians can obtain measures with the LF device effectively without using zero-value subtraction. For the LF pen, however, the absence of the zero-value subtraction changed both the sensitivity and specificity, and so the authors concluded that clinicians should not eliminate this step from the procedure.
Clinical Implications. When using the LF device, clinicians might not need to perform the zero-value subtraction; however, for the LF pen, clinicians should do so.
Key Words: Occlusal caries; DIAGNOdent 2095; DIAGNOdent pen 2190; laser fluorescence; zero valueAbbreviations: A: Cuspal • B: Middle • C: Cervical • D: Distal • D0: Free of caries • D1: Caries extending up to halfway through the enamel • D2: Caries extending into the inner one-half of enamel • D3: Caries in dentin • D4: Deep dentinal caries • LF: Laser fluorescence • M: Mesial • Subtr.: Zero-value subtraction
Detecting occlusal caries is a challenging task for clinicians because conventional diagnostic methods are more specific than sensitive.1,2 Therefore, researchers have developed new detection methods to help dentists determine the best treatments and preventive management strategies for dental caries.
Caries lesions emit stronger fluorescence than do sound tissues when stimulated by a red laser visible light with a wavelength of 655 nanometers.3 Several studies have examined the performance of a laser fluorescence (LF) device (DIAGNO-dent 2095, KaVo, Biberach, Germany)1–5; on the basis of the principle of fluorescence, the DIAGNO-dent 2095 has shown good or excellent results in occlusal caries detection.
Recently,5 the manufacturer introduced a new LF pen device (DIAGNOdent pen 2190, KaVo) for detecting occlusal and interproximal caries. It is because of the DIAGNOdent pen 2190’s new design and its tips that users also can assess interproximal surfaces. Both signs (fluorescence emission and excitation) run in the same optical path of propagation, but in opposite directions.6–8 This is one of the pen’s differences from the DIAGNOdent 2095 device.
For both devices, in accordance with the manufacturer’s instructions, users should perform two calibration steps: standard and individual calibration. The standard calibration consists of placing the tip on a porcelain object with known fluorescence and pressing the "CAL" button on the device. Users also should perform an additional individual calibration (placing the tip and pressing the CAL button) against a noncarious spot on the buccal surface, allowing the automatic subtraction of the zero value from the carious site of interest.3,9,10 Investigators have reported this step of individual automatic calibration as being a time-consuming procedure in the clinical practice9,11 and one that many practicing dentists do not routinely follow. Despite that, some authors5–8 have reported manually measuring the noncarious spot on the buccal surface to obtain the zero value of fluorescence, instead of doing this automatically. Then, after the user assesses the caries lesion, he or she manually subtracts the zero value (zero-value subtraction).
Preliminary studies from our laboratory showed higher values when we assessed occlusal sites after individual calibration, compared with the results we obtained after subtracting the zero value (J.de A. Rodrigues, unpublished data, 2007). This means that the values obtained from the buccal surface and automatically subtracted by the device are even lower than the values subtracted manually. Furthermore, we found no mention in the literature of the area in which clinicians should record the zero value when using the DIAGNOdent devices on occlusal surfaces. Clinicians can assume that the incorrect measurement or the lack of a measurement might influence the assessment of the caries status and, hence, their treatment decision.
The results of studies that evaluated the protocol of individual calibration are controversial. While Braun and colleagues9 suggested that clinicians should perform individual calibration because they found a difference of six units of LF in their assessments done using LF, Braga and colleagues10 did not find an influence of calibration on performance in their study using primary teeth. Owing to these divergent results and before we could indicate in vivo use of the LF devices, we decided to carry out this in vitro investigation of the calibration process. Furthermore, we found that no investigators had evaluated the influence of zero-value subtraction on LF performance using the DIAGNOdent pen 2190 for occlusal caries detection.
The aim of this study was to evaluate the influence of zero-value subtraction on the performance of two LF devices (DIAGNOdent 2095 and DIAGNOdent pen 2190) developed to detect occlusal caries in vitro.
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MATERIALS AND METHODS
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We used 119 human extracted maxillary and mandibular third molars (frozen at –20 C until use) suspected to have initial caries lesions on an occlusal surface. Preliminary studies in our laboratory showed that this method of storage does not change the red fluorescence signal significantly.12 Dental practitioners in Switzerland—a country with no water fluoridation and 250 parts per million of fluoride in table salt—had extracted all the teeth used for this study (which already had been scheduled for extraction for reasons other than the study). Before undergoing the extraction, the patients received information about the use of the teeth for research purposes and provided consent.
We removed calculus and debris using a scaler (Cavitron, Dentsply, York, Pa.). We cleaned the teeth for 15 seconds with a toothbrush and water and then for 10 seconds with Prophyflex (KaVo) with sodium bicarbonate. To avoid leaving powder remnants in the fissure, we rinsed the teeth off with a three-in-one syringe for 10 seconds.13 We took photographs of the occlusal and buccal surfaces (magnification x 6.25) and chose the site giving the highest LF value (using the DIAGNOdent 2095 device) on the occlusal surface from each tooth (test site).9 Additionally, we divided the buccal surfaces of the selected teeth into six noncarious areas (sites 1 to 6) (Figure 1
) for the zero-value measurements: three sites on the distal (D) portion and three on the mesial (M) portion, which we called cuspal (A), middle (B) and cervical (C) areas.
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Figure 1. Six sites on the buccal surface that the authors measured by using the DIAGNOdent 2095 (KaVo, Biberach, Germany) and the DIAGNOdent pen 2190 (KaVo). A: Cuspal. B: Middle. C: Cervical. D: Distal. M: Mesial.
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Before taking the LF measurements, we defrosted the teeth for three hours at room temperature. During the assessments, we stored the teeth individually in 100 percent humidity with drops of water to prevent dehydration.
Investigators have described in detail the mode of function of both LF devices (DIAGNOdent 2095 and DIAGNOdent pen 2190).1,5,6 Both LF systems function on the same principles and are based on the phenomenon of fluorescence, in which the absorption of light of a given wavelength by a fluorescent molecule is followed by the emission of light at longer wavelengths. These LF systems emit red light at a wavelength of 655 nm (approximately 1 milliwatt of power). A photodetector quantifies the fluorescent light passing through the filter and digitally displays a real-time (moment) and a maximum (peak) value. For the DIAG-NOdent pen 2190 device, the excitation and the emission of fluorescence follow the same solid fiber tip, but in opposite directions. This is the pen’s main difference from the DIAGNOdent 2095, which has different fibers for light excitation and emission.
After performing standard calibration, two experienced dentists (J.A.R. and M.B.D.) independently assessed each test site in the occlusal surface and each delimited area in the buccal surface twice, using the same tips for both surfaces according to their respective device, and recorded the values. For the measurements, the dentists used specific tips for occlusal caries detection: the A tip for DIAGNOdent 2095 and the cylindrical sapphire fiber tip for DIAGNOdent pen 2190 (with a diameter of 1.1 millimeters).
After the assessments, one of the examiners (J.A.R.) ground the teeth longitudinally on a Knuth-Rotor (Streurs, Copenhagen, Denmark) polishing machine using silicon carbide paper (60-micrometer grain size) and cooled the teeth in running tap water. The examiner constantly checked the progression of the grinding process under the microscope (magnification x 6.25). When the examiner reached the periphery of the site, he used silicon carbide papers of grain size 30, 18, 8 and 5 µm for further polishing.5 He then colored the cut surfaces with saturated rhodamine B (Fluka, Buch, Switzerland) and performed the examination using the dye’s penetration into the enamel, the dentinal tissues or both for feedback (Figure 2). The examiner assessed the sites for the presence of caries and, when present, for the degree of caries extension (magnification x10). We classified the sites as caries-free (D0), with caries extending up to halfway though the enamel (D1), with caries extending into the inner one-half of enamel (D2), with caries in dentin (D3) and deep dentinal caries (D4). Subsequently, we took photographs.
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Figure 2. The rhodamine B dye application. (Illustration reproduced with permission of Cristiano Faria Maciel.)
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Statistical analyses.
We rejected the normal distribution of the values measured in the buccal as well as in the occlusal sites (P < .0001), which was aided by using statistical analysis software (MedCalc for Windows, Version 9.3.0.0, MedCalc Software, Mariakerke, Belgium). As the Wilcoxon rank sum test results did not show any statistically significant difference between the mesial and distal areas of the buccal surface, we combined the values from the A, B and C portions of these two areas. For each tooth, we calculated and subtracted the average between the M and D surfaces from the A, B and C areas in the buccal surface from the value measured in the respective occlusal site. We performed a Wilcoxon rank sum test to compare the values of fluorescence obtained with and without zero-value subtraction. We calculated and compared the areas under the receiver operating characteristic (ROC) curve for each threshold.14 We assessed the performance according to the cutoff limits suggested by Lussi and Hellwig5 (both with and without zero-value subtraction), for the DIAGNOdent 2095 device as follows: 0.0 to 7.0 = sound enamel; 7.1 to 14.0 = caries in enamel; 14.1 to 24.0 = caries in dentino-enamel junction; greater than 24 = caries in dentin. Our cutoff limits for the DIAGNOdent pen 2190 device were as follows: 0.0 to 6.0 = sound enamel; 6.1 to 13.0 = caries in enamel; 13.1 to 17.0 = caries in dentinoenamel junction; greater than 17.0 = caries in dentin. We compared the values of sensitivity, specificity and accuracy by performing the McNemar test. We set the significance level at P < .05.
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RESULTS
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Table 1 shows the average, standard deviation (SD) and 95 percent confidence interval (CI) of the readings from each area using the DIAG-NOdent 2095 and DIAGNOdent pen 2190 devices in the buccal surface. Table 1 also shows the increase of values from area A (6.5 and 8.9) to area C (9.4 and 13.0, respectively for DIAGNOdent 2095 and DIAGNOdent pen 2190). For both devices, the Wilcoxon rank sum test showed a statistically significant difference (P < .05) between areas A, B and C but no difference between M and D. Furthermore, we found a statistical difference when comparing the values obtained by the two LF devices. We observed a statistically significant difference when we compared the procedures without and with zero-value subtraction (subtracting the values from the areas A, B and C) (Figure 3). Figure 4 (page 1110) shows three examples of occlusal sites, including the values obtained in the different areas in the buccal surface and the values after zero-value subtraction, as well as images obtained initially and after histologic assessments.
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TABLE 1 Average of DIAGNOdent 2095 and DIAGNOdent pen 2190 readings, standard deviation and the 95 percent confidence interval in cuspal, middle, cervical, mesial and distal areas of the teeth.
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Figure 3. Box plots for laser fluorescence values of the devices from measurements on the occlusal surface without and with zero-value subtraction. A. DIAGNOdent 2095 (KaVo, Biberach, Germany). B. DIAGNOdent pen 2190 (KaVo). Note the statistically significant differences between the procedures (indicated by lowercase letters a-d) (P < .05, Wilcoxon rank sum test). The figure shows the median, first and third quartiles, minimum and maximum values (whiskers) and outliers (marked as dots).
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Figure 4. Occlusal photographs and averaged laser fluorescence values (DIAGNOdent 2095 [KaVo, Biberach, Germany] and the DIAGNOdent pen 2190 [KaVo]) obtained in different areas of the buccal and occlusal surface, as well as the values after zero-value subtraction. Histologic assessments: tooth 1 = score 2 (caries extending into the inner one-half of enamel), tooth 2 = score 3 (caries in dentin) and tooth 3 = score 4 (deep dentinal caries). A: Cuspal. B: Middle. C: Cervical. Subtr.: Zero-value subtraction.
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Table 2 (page 1111) shows the values of sensitivity, specificity, accuracy and area under the ROC curve. For the DIAGNOdent 2095 device, the values of specificity did not show a statistically significant difference, but sensitivity and accuracy were higher when we did not perform zero-value subtraction. For the DIAGNOdent pen 2190 device, the specificity increased and the sensitivity decreased significantly when we performed zero-value subtraction. However, only the accuracy at the D2 threshold changed significantly. The area of the ROC curve varied from 0.73 to 0.81 for the DIAGNOdent 2095 device and from 0.72 to 0.80 for the DIAGNOdent pen 2190. For both devices, we found no statistically significant difference comparing the areas under the ROC curves with and without the zero-value subtraction.
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TABLE 2 Performance of the DIAGNOdent 2095 and DIAGNOdent pen 2190* devices for occlusal caries detection without and with zero-value subtraction in cuspal, middle and cervical areas.
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DISCUSSION
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As suggested by the manufacturer, those using the LF devices should locate a sound spot on the buccal surface to obtain the value of fluorescence zero, which can be either automatically subtracted from the site of interest (individual calibration)9,10 or manually subtracted after the lesion assessment (zero-value subtraction).5–8 This step of calibration is somewhat time-consuming, and many clinicians do not routinely follow it in practice. Some variation in these measurements may change the LF values and, consequently, the treatment decision. Researchers studying the first device and assessing the buccal surface found values varying by up to six units over the four quadrants,9 whereas others did not find differences in LF readings without the zero-value subtraction.10
Some investigators have explained the highest values of fluorescence in cervical areas by observing the mineralization levels in the different areas of the teeth15 and by noting the decreasing thickness of enamel toward the cementoenamel junction, which allows high fluorescence measurements of the underlying dentin.16 The high values of fluorescence found in the cervical area for both devices could respond similarly to dental caries on the occlusal surface. However, we evaluated the mesial and distal areas and noted that the values showed no statistically significant difference. This was likely because the average among the areas A, B and C of both mesial and distal portions represented similar levels of mineralization.
We found the highest values for the DIAGNOdent pen 2190 when we compared the same areas assessed by both devices with a statistically significant difference (Wilcoxon rank sum test, P < .05), as Kühnisch and colleagues7 also found when they assessed occlusal sites in vitro. By contrast, Krause and colleagues17 found lower fluorescence values for the DIAGNOdent pen 2190 in an in vivo study assessing occlusal surfaces (they obtained a median value of five units lower after zero-value calibration); they did not mention the area used for their assessment of the buccal surface.
In our study, the values measured on the buccal surface ranged from 6.5 to 9.4 for the DIAGNOdent 2095 and 8.9 to 13.0 for the DIAGNOdent pen 2190, varying in the A to the B and C areas. In light of that, we could test and confirm the influence of a specific area for the zero-value subtraction on the performance of the DIAGNOdent 2095 and DIAGNOdent pen 2190. The smaller diameter of the cylindrical tip (1.1 mm) and the different composition of each device could be factors contributing to the differences between the values we found in the performance of the DIAGNOdent 2095 and DIAGNOdent pen 2190. The cylindrical tip used for the DIAGNOdent pen 2190 assessments in occlusal surfaces is made of a solid single sapphire fiber, and the end of the tip has a prismatic shape that deflects the beam of excitation and collects it laterally along its longitudinal axis (the excitation and the emission run through the same fibers but in opposite directions, as explained earlier). For the DIAGNOdent 2095 device, the whole probe is made of a fiber rod, consisting of a bundle of single fibers, each of them with a diameter of 40 µm.5 Furthermore, statistically the high number of outliers seen via box plotting (Figure 3
) likely is due to the abnormal data distribution.
In our study, we tested different tooth areas for the zero-value subtraction to verify its influence on the performance of both devices. For the DIAGNOdent 2095 device, we found no influence when we compared the values of specificity with and without zero-value subtraction. The zero-value subtraction showed an influence on the performance when we used the cutoff limits proposed by Lussi and Hellwig,5 significantly decreasing the sensitivity and accuracy from the B toward the C surfaces. Considering that we maintained other parameters and that the observed difference was not positive, we could suppress this step for the LF device. Our study’s findings agree with those of Braga and colleagues,10 who found higher LF readings from assessments performed without the individual calibration using the center of each tooth’s buccal surface, but no influence on the LF performance when assessing primary teeth.
Some authors studying the same subject in permanent teeth obtained different results.9 They observed that individual calibration influences LF readings and indicated that clinicians should perform this procedure before performing the diagnostic measurement.9 However, clinicians must consider that in both studies,9,10 these investigators performed individual calibration and not zero-value subtraction. We suggest that future investigations compare both kinds of procedures. The histologic validation and the cutoff limits used in previous studies were different from those in our investigation, which also could explain the different results we observed. In unpublished studies from our laboratory, we observed positive correlations between the micro-hardness and dye-enhanced rhodamine B.
For the DIAGNOdent pen 2190 device, we observed the influence of the zero-value subtraction when we analyzed the specificity, and we found differences when comparing these values within the different areas. However, we observed higher values of sensitivity at D2 and D3 when we did not perform the zero-value subtraction. The difference between the high sensitivity and low specificity suggests that clinicians should not eliminate the zero-value subtraction and that any area could be suggested for the zero-value assessment. More research associating cutoff limits and zero-value subtraction using the DIAGNOdent pen 2190 for occlusal caries detection is needed.
The area under the ROC curve displays the ability of both devices to detect occlusal caries lesions (sensitivity), as well as the absence of occlusal caries lesions (specificity) with and without the zero-value subtraction. However, our analysis had several cutoff limits and did not account for a stipulated value for the calculation.18 The advantages of the ROC curve are that it includes several cutoff points, it shows the relationship between the sensitivity and specificity, and it is not affected by the prevalence of disease.19
It is important to point out that these results represent an in vitro situation, and so investigators should use caution when using these devices in clinical practice. Even though the setup of our study was such that storage should not influence the LF values,9 clinicians should consider the use of LF devices to serve as a second opinion only.6 Furthermore, the treatment decision also depends on other patient variables, such as dietary and toothbrushing habits, caries activity and use of fluoride.
Additional in vivo studies using deciduous and permanent teeth should be carried out. The use of the LF pen device also needs to be analyzed further.
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CONCLUSION
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We observed an influence of the zero-value subtraction in the DIAGNOdent 2095 device’s performance: the sensitivity decreased. However, we conclude that clinicians could perform the LF readings without the zero-value subtraction. For the DIAGNOdent pen 2190, the absence of the zero-value subtraction changed both the sensitivity and specificity, and, therefore, clinicians and researchers should not eliminate the use of zero-value subtraction when using the LF pen.
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FOOTNOTES
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Dr. Rodrigues is a postdoctoral fellow, Department of Preventive, Restorative and Pediatric Dentistry, School of Dental Medicine, University of Bern, Freiburgstrasse 7, CH 3010, Bern, Switzerland, e-mail "jorodrigues{at}hotmail.com". Address reprint requests to Dr. Rodrigues.
Ms. Hug is a laboratory technician, Department of Preventive, Restorative and Pediatric Dentistry, School of Dental Medicine, University of Bern, Switzerland.
Ms. Diniz is a PhD student, Department of Pediatric Dentistry, School of Dentistry, Araraquara, São Paulo State University, Brazil.
Dr. Cordeiro is a professor, Department of Pediatric Dentistry, School of Dentistry, Araraquara, São Paulo State University, Brazil.
Dr. Lussi is a professor, Department of Preventive, Restorative and Pediatric Dentistry, School of Dental Medicine, University of Bern, Switzerland.
Disclosure. The authors did not report any disclosures.
The authors thank Cristiano Faria Maciel for providing the illustration in Figure 2.
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ABSTRACT
MATERIALS AND METHODS
