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Measuring subcutaneous fat thickness using skinfold calipers vs. high-resolution B-scan ultrasonography in healthy volunteers: A pilot study

Open AccessPublished:December 02, 2021DOI:https://doi.org/10.1016/j.nutos.2021.11.007

      Summary

      Background & Aims

      Measuring the thickness of skinfolds using calipers is a frequently used method for determining body fat in clinical practice. It is inexpensive and time saving compared to other methods and can be learned quickly. The aim of this study is to compare skinfold measurements made using calipers against measurements obtained using high-resolution ultrasonography (USG).

      Methods

      A monocentric cross-sectional observational pilot study with 69 normal-weight volunteer participants aged between 18 and 80 years (women: n = 36, men: n = 33). Subcutaneous fat thickness was measured at five different sites (chest, abdomen, thighs, triceps, and back) using calipers and via USG.
      The mean values from the calipers and the USG measurements were examined for statistical significance using the paired t-test, and the Bland–Altman plot was used to compare the two methods. We then performed an agreement analysis by calculating the percentage of the differences less than or equal to 1 mm between the measurements made using calipers and those obtained via USG.

      Results

      From the Bland–Altman analysis, the best matches among measurements for the female subjects were the skinfolds on the triceps (ULoA: 4.6 mm, mean: −0.1 mm, LLoA: −4.8 mm), the back (ULoA: 3.9 mm, mean: −0.2 mm, LLoA: −4.4 mm), and on the chest (ULoA: 5.9 mm, mean: 0.6 mm, LLoA: −4.6 mm). In contrast, for the male subjects, the best matches were the skinfolds on the triceps (ULoA: 3.3 mm, mean: −0.3 mm, LLoA −4.0 mm), the back (ULoA 3.5 mm, mean −0.3 mm, LLoA −4.0 mm), and the thigh (ULoA 2.2 mm, mean −0.5 mm, LLoA −3.2 mm).
      The best agreement for women was obtained with the measurements of the chest fold (47.2%), followed by the measurements of the back fold (38.9%). For men, it was the measurements of the thigh fold (63.6%), followed by the measurements of the triceps (51.5%) and back fold (51.5%).

      Conclusion

      The agreement between skinfold measurements made using calipers and ultrasonography depends on the skinfold thickness and the localization of the measurement, with gender-based differences.

      Keywords

      1. Introduction

      To determine the nutritional status of a patient, it is essential to examine the fat content in the body. Calculating body fat using skinfold measurement is still a clinically established method [
      • Kuo F.C.
      • Lu C.H.
      • Wu L.W.
      • Kao T.W.
      • Su S.C.
      • Liu J.S.
      • et al.
      Comparison of 7-site skinfold measurement and dual-energy X-ray absorptiometry for estimating body fat percentage and regional adiposity in Taiwanese diabetic patients.
      ], and is applicable for both normal-weight patients at risk of malnutrition and patients suffering from obesity [
      • Sengeis M.
      • Müller W.
      • Störchle P.
      • Führhapter-Rieger A.
      Body weight and subcutaneous fat patterning in elite judokas.
      ,
      • Yeşil E.
      • Köse B.
      • Özdemir M.
      Is Body Adiposity Index a Better and Easily Applicable Measure for Determination of Body Fat?.
      ]. Obesity plays an increasingly important role in daily clinical work because its prevalence has reached almost epidemic proportions [
      • Ezzati M.
      Worldwide trends in body-mass index, underweight, overweight, and obesity from 1975 to 2016: a pooled analysis of 2416 population-based measurement studies in 128.9 million children, adolescents, and adults.
      ,]. Being overweight or obese has a severe impact on quality of life and lowers the life expectancy of those affected [
      • Kolotkin R.L.
      • Williams V.S.L.
      • Ervin C.M.
      • Williams N.
      • Meincke H.H.
      • Qin S.
      • et al.
      Validation of a new measure of quality of life in obesity trials: Impact of Weight on Quality of Life-Lite Clinical Trials Version.
      ,
      • Vidra N.
      • Trias-llimós S.
      • Janssen F.
      Impact of obesity on life expectancy among different European countries: secondary analysis of population-level data over the 1975 – 2012 period.
      ]. Body fat measurement plays a crucial role in diagnosing obesity [
      • Kuo F.C.
      • Lu C.H.
      • Wu L.W.
      • Kao T.W.
      • Su S.C.
      • Liu J.S.
      • et al.
      Comparison of 7-site skinfold measurement and dual-energy X-ray absorptiometry for estimating body fat percentage and regional adiposity in Taiwanese diabetic patients.
      ]. For body fat measurement, determining body fat composition by measuring the thickness of skinfolds is cost effective, requires minimal equipment, and is a practicable method of determining body fat in everyday clinical practice [
      • Ketel I.J.G.
      • Volman M.N.M.
      • Seidell J.C.
      • Stehouwer C.D.A.
      • Twisk J.W.
      • Lambalk C.B.
      Superiority of skinfold measurements and waist over waist-to-hip ratio for determination of body fat distribution in a population-based cohort of Caucasian Dutch adults.
      ,
      • Sobotka L.
      • Allison S.
      • Forbes A.
      • Meier R.
      • Schneider S.M.
      • Soeters P.B.
      • et al.
      Basics in clinical nutrition.
      ]. Using a suitable equation, fat-free mass (FFM) and fat mass (FM) can be calculated [
      • Durnin J.V.
      • Womersley J.
      Body fat assessed from total body density and its estimation from skinfold thickness: measurements on 481 men and women aged from 16 to 72 years.
      ,
      • Jackson A.S.
      • Pollock M.L.
      Generalized equations for predicting body density of men.
      ]. Despite great care, caliper measurements have numerous limitations. Kuczmarski et al. [
      • Kuczmarski R.J.
      • Fanelli M.
      • Koch G.G.
      Ultrasonic assessment of body composition in obese adults: overcoming the limitations of the skinfold caliper.
      ] highlighted numerous problems associated with the caliper measurement method. The transition from fat to muscle cannot always be palpated. With thicker skinfolds, the measuring lever tips can slip, leading to significantly thicker or thinner erroneous skinfold measurements. Repeated measurements tend to return lower values due to compression, and edema is another factor that falsifies measurement results. Furthermore, the elasticity of fat and skin tissue varies with age and sex, and interindividual differences are also known to interfere with measurement accuracy. Pain or discomfort at certain sites can also lead to inaccurate readings [
      • Störchle P.
      • Müller W.
      • Sengeis M.
      • Lackner S.
      • Holasek S.
      • Fürhapter-Rieger A.
      Measurement of mean subcutaneous fat thickness: eight standardised ultrasound sites compared to 216 randomly selected sites.
      ,
      • Müller W.
      • Lohman T.G.
      • Stewart A.D.
      • Maughan R.J.
      • Meyer N.L.
      • Sardinha L.B.
      • et al.
      Subcutaneous fat patterning in athletes: selection of appropriate sites and standardisation of a novel ultrasound measurement technique: ad hoc working group on body composition, health and performance, under the auspices of the IOC Medical Commission.
      ].
      Diagnostic ultrasonography (USG) images are generated using sequences of ultrasound (US) beams in a pulse-echo technique. A short pulse of US waves penetrates the tissue at the speed of sound. The echo intensity of the tissue corresponds to the brightness of the screen. Through improvements to the USG method, especially by Müller [
      • Müller W.
      • Lohman T.G.
      • Stewart A.D.
      • Maughan R.J.
      • Meyer N.L.
      • Sardinha L.B.
      • et al.
      Subcutaneous fat patterning in athletes: selection of appropriate sites and standardisation of a novel ultrasound measurement technique: ad hoc working group on body composition, health and performance, under the auspices of the IOC Medical Commission.
      ,
      • Müller W.
      • Horn M.
      • Fürhapter-Rieger A.
      • Kainz P.
      • Kröpfl J.M.
      • Maughan R.J.
      • et al.
      Body composition in sport: a comparison of a novel ultrasound imaging technique to measure subcutaneous fat tissue compared with skinfold measurement.
      ], it became possible to standardize the measurement of subcutaneous fat tissue in athletes and persons of normal weight. The applicability of the USG measurement to overweight and obese persons has also been confirmed [
      • Störchle P.
      • Müller W.
      • Sengeis M.
      • Ahammer H.
      • Fürhapter-Rieger A.
      • Bachl N.
      • et al.
      Standardized Ultrasound Measurement of Subcutaneous Fat Patterning: High Reliability and Accuracy in Groups Ranging from Lean to Obese.
      ]. The age of the test subjects, as Weits et al. [
      • Weits T.
      • van der Beek E.J.
      • Wedel M.
      Comparison of ultrasound and skinfold caliper measurement of subcutaneous fat tissue.
      ] were able to show, is a known confounder. Due to skin elasticity, there is greater agreement between the results of the two methods (calipers and USG) for younger subjects than for older subjects [
      • Weits T.
      • van der Beek E.J.
      • Wedel M.
      Comparison of ultrasound and skinfold caliper measurement of subcutaneous fat tissue.
      ].
      In contrast to skinfold measurements using calipers, the USG method if used correctly should be used without any pressure and therefore does not necessarily compress the tissue because forming a skinfold is unnecessary, nor is the measurement influenced by the viscosity or elasticity of the tissue. However, measurement sites on the trunk have not proved to be optimal for USG, as the transition from subcutaneous fat tissue to muscle may be less visible under certain circumstances. Limitations to the use of USG include a limited availability of USG in epidemiological studies and the training of examiners [
      • Müller W.
      • Horn M.
      • Fürhapter-Rieger A.
      • Kainz P.
      • Kröpfl J.M.
      • Maughan R.J.
      • et al.
      Body composition in sport: a comparison of a novel ultrasound imaging technique to measure subcutaneous fat tissue compared with skinfold measurement.
      ,
      • Störchle P.
      • Müller W.
      • Sengeis M.
      • Ahammer H.
      • Fürhapter-Rieger A.
      • Bachl N.
      • et al.
      Standardized Ultrasound Measurement of Subcutaneous Fat Patterning: High Reliability and Accuracy in Groups Ranging from Lean to Obese.
      ,
      • Müller W.
      • Horn M.
      • Fürhapter-Rieger A.
      • Kainz P.
      • Kröpfl J.M.
      • Ackland T.R.
      • et al.
      Body composition in sport: Interobserver reliability of a novel ultrasound measure of subcutaneous fat tissue.
      ].
      The aim of this study is to confirm the reliability of skinfold measurements using calipers by comparing them with measurements obtained via USG. We raised the following questions: Are certain skinfolds more suitable for caliper measurement than others? Are there any gender-specific differences? Does skinfold thickness impact agreement between measurements obtained using either method?

      2. Materials and methods

      2.1 Study design

      This research is part of an explorative observational pilot study with a cross-sectional design [
      • Mair E.
      • Nösslinger H.
      Comparison of indirect calorimetry with predictive BMR formulas in normal weight subjects and development of a new formula for fat determination by means of skinfold thickness.
      ]. The study protocol was designed in compliance with the Declaration of Helsinki, 1964. Approval for this study was obtained from the Ethics Committee of the South Tyrolean Health Authority (Approval number: 54-2018).

      2.2 Study participants

      The recruitment of volunteers was not randomized. We used an exponential mostly discriminatory snowball sampling procedure. For the sample size calculation, we followed the recommendations for pilot studies [
      • Machin D.
      • Campbell M.J.
      • Tan S.B.
      • Tan S.H.
      Sample sizes for clinical, laboratory and epidemiology studies.
      ] and verified them using a formula adopted from the statistical handbook by Dallal [
      • Dallal G.E.
      The little handbook of statistical practice.
      ]. Of the 89 volunteers recruited, only 69 met the inclusion criteria. The most important exclusion criteria were factors that could influence body composition or energy metabolism, including pathologies such as infectious diseases, severe chronic diseases, eating disorders, medications, and intensive sporting activity the previous day. A detailed list of the exclusion criteria is presented in the supplementary material.
      The study subjects included 69 normal-weight, healthy participants (women: n = 36, men: n = 33; age: 18–80 years; BMI: 18.5–25 kg/m2). For subjects aged 65 and above, the BMI upper limit was increased to ≤27 kg/m2. Seca 704 scales (Seca, Germany) and a Seca 222 stadiometer (Seca, Germany) were used to measure body weight and height.
      Participants were instructed to have a night's rest of at least seven hours and to be sober for the test. They presented themselves for examination on an empty stomach and an empty bladder.

      2.3 Skinfold measurement

      Skinfolds were measured under standardized conditions [
      • Hume P.
      • Marfell-Jones M.
      The importance of accurate site location for skinfold measurement.
      ], and at the same time, at the same ambient temperature, with the same equipment, and always by the same investigator. Measurements were made on the right side of the body, with the subjects standing, and at previously marked points on the thighs, abdomen, chest, triceps, and back. The marking ensured that both the caliper and the USG measurements were made at exactly the same locations (Fig. 1). The exact measuring sites were selected and found according to Eston and Reilly [
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ].
      Fig. 1
      Fig. 1Measuring points of skinfold measurements (our representation). Ventral: [
      • Kuo F.C.
      • Lu C.H.
      • Wu L.W.
      • Kao T.W.
      • Su S.C.
      • Liu J.S.
      • et al.
      Comparison of 7-site skinfold measurement and dual-energy X-ray absorptiometry for estimating body fat percentage and regional adiposity in Taiwanese diabetic patients.
      ] Chest Fold, oblique skinfold raised along the borderline of the musculus pectoralis major between the anterior axillary fold and the nipple. Females: measurement is taken at one third of the distance between anterior axillary fold and nipple. Males: measurement is taken at one half of the distance between anterior axillary fold and nipple [
      • Sengeis M.
      • Müller W.
      • Störchle P.
      • Führhapter-Rieger A.
      Body weight and subcutaneous fat patterning in elite judokas.
      ,
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ] Abdominal Fold, horizontal fold raised 3 cm lateral and 1 cm inferior to the umbilicus [
      • Yeşil E.
      • Köse B.
      • Özdemir M.
      Is Body Adiposity Index a Better and Easily Applicable Measure for Determination of Body Fat?.
      ,
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ] Thigh Fold, vertical skinfold raised on the anterior aspect of the thigh midway between the inguinal crease and the proximal border of the patella [
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ]; Dorsal: [
      • Ezzati M.
      Worldwide trends in body-mass index, underweight, overweight, and obesity from 1975 to 2016: a pooled analysis of 2416 population-based measurement studies in 128.9 million children, adolescents, and adults.
      ] Back/Subscapular Fold, oblique skinfold raised 1 cm below the inferior angle of the scapula at approximately 45° to the horizontal plane following the natural cleavage lines of the skin [,
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ] Triceps Fold, vertical skinfold raised on the posterior aspect of the musculus triceps, exactly halfway between the olecranon process and the acromion process when the hand is supinated [
      • Eston R.
      • Reilly T.
      Kinanthropometry and exercise physiology - laboratory manual.
      ].
      A Holtain Tanner/Whitehouse Skinfold Caliper (Holtain Model 610ND), an established and widely used, high-quality measuring device, was used. The measurement was carried out five times subsequently, and the investigator waited until the skinfold was smoothed between each measurement. We have followed the ACSM's Guidelines for Exercise Testing and Prescription [
      • Riebe D.
      • Ehrman J.K.
      • Liguori G.
      • Magal M.
      ACSM’s Guidelines for Exercise testing and prescription.
      ] and duplicated each of the five measurements if they were not within 1 mm for skinfolds under 20 mm, and within 2 mm for skinfolds above 20 mm. From this, a mean value was calculated for use in the statistical evaluation.
      A high-end USG device, Philips EPIQ 7 (SIDEM. SpA, Italy), with its high-frequency L18-5 MHz broadband linear transducer preset to high resolution, was used for the measurement of subcutaneous fat thickness between the epidermis (epidermis included) to the upper edge of the muscle fascia. Two measurements, longitudinal and transversal, were always performed by the same experienced examiner, who also did the skinfold measurements by caliper and then the mean was calculated. Fat thickness was always evaluated in the middle of the scan.
      Both the longitudinal and transverse planes were examined, specifically for the detection and elimination of possible artifacts.
      The positions measured using the USG device were the same as those measured using the caliper. To find the same spot, the positions were marked with a pen. To compare the USG measurements with the caliper measurements, the former was multiplied by a factor of two. When using USG, the skin thickness and subcutaneous fat tissue are measured without generating a skinfold, with the USG probe placed on flat skin without any pressure. In contrast to caliper measurements, there is only one layer of skin thickness and subcutaneous fat tissue measured using the USG method. Whereas while performing caliper measurements, the skin is pushed together to form a fold, and consequently, each layer is measured twice (Fig. 2, Fig. 3, Fig. 4, Fig. 5, Fig. 6).
      Fig. 2
      Fig. 2Example of subcutaneous fat thickness measured using USG in two planes: Chest.
      Fig. 3
      Fig. 3Example of subcutaneous fat thickness measured using USG in two planes: Abdomen.
      Fig. 4
      Fig. 4Example of subcutaneous fat thickness measured using USG in two planes: Thigh.
      Fig. 5
      Fig. 5Example of subcutaneous fat thickness measured using USG in two planes: Back/Subscapular.
      Fig. 6
      Fig. 6Example of subcutaneous fat thickness measured using USG in two planes: Triceps.

      2.4 Bioelectric impedance analysis

      Although the purpose of this study is to compare skinfold measurements made using calipers and USG, a determination of FFM and FM via bioelectric impedance analysis (BIA) was included in the examination of the subjects. The measurements were conducted by the same trained individual throughout the study. No physical exertion or activity was allowed during the 24 hours preceding the examination, and no food was allowed for at least 12 hours before the examination, though drinking water was allowed. The subjects were examined at the same time (8 am), lying on their backs and completely relaxed, for at least 20 minutes. The room temperature was 24 °C and the study participants were awake and lightly dressed. Before beginning the measurements, the subjects had warm extremities with normal skin circulation. The device used for these measurements was a Nutribox analyzer (Data Input GmbH, Pöcking, Germany), a 50 kHz single frequency meter with four electrodes. The output variables obtained from the Nutribox BIA system are phase angle [°], body water [l], FFM (body cell mass [BCM] + extracellular mass [ECM]), [kg], ECM [kg], BCM [kg], ECM/BCM index, and amount of BCM in FFM [%], FM [kg], FM [%], vector diagram. Evaluation and interpretation were based on a large German population BIA database with data on more than 200,000 adults [
      • Bosy-Westphal A.
      • Danielzik S.
      • Dörhöfer R.-P.
      • Piccoli A.
      • Müller M.J.
      Patterns of bioelectrical impedance vector distribution by body mass index and age: implications for body-composition analysis.
      ].

      2.5 Statistics

      R (version 3.5.1) and R-Studio (version 1.2.1335) were used for the statistical calculation and creation of graphics. To describe the distributions of the quantitative data collected, we calculated the mean value and standard deviation. To compare the centers of distribution, we used the paired t-test.
      All data sets were checked for normal distribution using the Shapiro–Francia test, a Cullen and Frey graph, and a Q-Q plot with a 95% confidence interval (CI). It follows that all the data can be considered normally distributed. In this report, unless otherwise stated, we used the P-value and effect size (Cohen's d with the following interpretation: small = 0.2, medium = 0.5, large = 0.8) from the paired t-test. We set the significance level at 5% (0.05).
      A large part of the statistical evaluations concern correlation analyses, in which we calculated the Pearson's correlation coefficient and Lin's concordance correlation coefficient based on the variables in our data. In addition to correlation plots, Bland–Altman plots were used to interpret the comparative data on the methods being compared. We performed an analysis of agreement for all skinfolds examined, independent of the sex of the participants.
      For the agreement analysis, we did not use the common accurate prediction rule, which employs the percentage rates of the measured values [
      • Bartolucci A.
      • Singh K.
      • Bae S.
      Introduction to statistical analysis of laboratory data.
      ]. This was partly because the different spans of the triceps and thigh skinfold thicknesses between men and women—and partly because the varying thicknesses of the different skinfolds themselves—would have led to non-constant evaluation criteria. Because the precision of repeated measurements using the Harpenden calipers—with the arms open from 20 mm to 25 mm—varies by about 1 mm [
      • Léger L.A.
      • Lambert J.
      • Martin P.
      Validity of Plastic Skinfold Caliper Measurements.
      ], we used this value (i.e., 1 mm) for the agreement analysis of the caliper measurements with respect to the USG measurements. We therefore consider a caliper measurement accurate if the difference from the USG measurement is less than 1 mm.

      3. Results

      A summary of the anthropometric data on the study population is given in Table 1.
      Table 1Anthropometric data.
      ParameterGenderMeanSDMedianMin - Q25 - Q75 - Max
      Age (years)Women49.418.650.0[18.0 - 30.8 - 65.5 - 80.0]
      Men49.117.252.0[18.0 - 35.0 - 62.0 - 77.0]
      Height (cm)Women165.74.8165.3[155.5 - 162.0 - 169.0 - 179.8]
      Men177.85.0177.8[168.5 - 175.0 - 181.0 - 191.0]
      Body mass (kg)Women60.97.761.2[47.7 - 55.9 - 65.5 - 74.9]
      Men73.06.873.2[53.6 - 68.6 - 76.9 - 91.4]
      Fat mass (kg)Women16.44.715.1[8.6 - 13.0 - 20.2 - 26.3]
      Men12.53.812.3[4.9 - 9.9 - 14.8 - 19.5]
      Fat-free mass (kg)Women44.53.743.5[37.3 - 42.3 - 46.2 - 53.9]
      Men60.65.161.1[46.0 - 58.0 - 62.9 - 73.4]
      BMI (kg/m2)Women22.12.522.0[18.5 - 20.2 - 24.5 - 26.9]
      Men23.11.923.2[18.5 - 21.7 - 24.7 - 26.5]
      Waist (cm)Women77.99.577.0[61.0 - 72.5 - 86.0 - 93.0]
      Men83.46.484.0[70.0 - 78.0 - 89.0 - 95.0]
      Hip (cm)Women93.06.992.0[74.0 - 89.8 - 97.3 - 106.0]
      Men92.43.893.0[81.0 - 90.0 - 95.0 - 100.0]
      Waist-to-Hip RatioWomen0.840.070.83[0.67 - 0.80 - 0.88 - 0.97]
      Men0.900.050.92[0.80 - 0.86 - 0.95 - 1.04]
      Fat mass and fat-free mass were determined via BIA measurement (Nutribox analyzer, Data Input GmbH, Pöcking, Germany).
      BMI (body mass index), Max (maximum value), Min (minimum value), SD (standard deviation), Q25 (25th percentile), Q75 (75th percentile).
      For all skinfolds, there was a high correlation between the USG and caliper measurements. For women, the highest correlation was for the skinfold at the back (Pearson r = 0.97, with P < 0.0001; Lin's concordance correlation coefficient = 0.97), and for men, the skinfold at the thigh (Pearson r = 0.97, with P < 0.0001; Lin's concordance correlation coefficient = 0.94). For both sexes, the lowest correlation was found for the chest skinfold (women: Pearson r = 0.87, with P < 0.0001, and Lin's concordance correlation coefficient = 0.86; men: Pearson r = 0.86, with P < 0.0001, and Lin's concordance correlation coefficient = 0.86).
      In our data, women had skinfolds on the triceps and thighs that were almost twice as thick as similar skinfolds on men (Table 2).
      Table 2Skinfolds (USG and caliper measurements) on women and men.
      Mean

      (mm)
      SD

      (mm)
      Median

      (mm)
      Min - Q25 - Q75 - Max

      (mm)
      Corr

      r
      Exact p

      (t-test)
      Effect size

      Cohen's d
      Agreement

      (%)
      Female (n = 36)
      Triceps: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      19.46.019.7[10.0 - 14.0 - 22.7 - 32.4]0.930.8360.0222.2
      Triceps: Calipers19.56.517.6[9.0 - 14.5 - 24.1 - 33.6]
      Back: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      17.27.715.9[6.8 - 10.4 - 22.8 - 32.4]0.970.5530.0238.9
      Back: Calipers17.48.515.4[7.2 - 9.5 - 24.2 - 34.5]
      Chest: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      12.75.412.1[6.4 - 8.1 - 14.4 - 28.0]0.870.2100.1247.2
      Chest: Calipers12.15.011.2[3.0 - 8.9 - 14.9 - 25.4]
      Abdomen: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      22.68.522.7[8.8 - 15.4 - 29.8 - 39.8]0.910.0130.2033.3
      Abdomen: Calipers21.07.220.0[8.2 - 14.4 - 26.8 - 35.2]
      Thigh: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      25.07.823.7[12.0 - 18.8 - 27.4 - 44.6]0.940.0020.1922.2
      Thigh: Calipers26.57.826.0[12.0 - 21.4 - 29.5 - 46.4]
      Male (n = 33)
      Triceps: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      10.03.09.6[4.8 - 8.0 - 11.6 - 16.8]0.900.0080.2051.5
      Triceps: Calipers9.43.08.6[4.0 - 7.2 - 11.8 - 16.0]
      Back: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      14.05.312.4[7.2 - 10.0 - 17.2 - 27.8]0.930.4210.0451.5
      Back: Calipers14.25.312.0[6.0 - 10.4 - 18.2 - 27.0]
      Chest: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      11.14.710.2[5.0 - 7.6 - 14.0 - 28.4]0.860.3340.1130.3
      Chest: Calipers10.64.89.2[4.2 - 7.2 - 14.0 - 22.2]
      Abdomen: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      19.87.918.0[6.0 - 14.2 - 24.0 - 40.2]0.930.2530.0842.4
      Abdomen: Calipers19.26.918.0[5.8 - 14.0 - 23.6 - 36.2]
      Thigh: USG
      USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      11.33.710.8[6.0 - 8.6 - 13.6 - 18.2]0.970.0460.1263.6
      Thigh: Calipers11.84.611.6[5.0 - 7.6 - 15.0 - 20.8]
      Corr (Pearson product-moment correlation coefficient), Max (maximum value), Min (minimum value), Q25 (25th percentile), Q75 (75th percentile), SD (standard deviation).
      a USG measurement multiplied by a factor of 2, † P < 0.0001, P-value and effect size (Cohen's d), Accuracy (percent of measurements with a difference of ≤1 mm between USG and calipers).
      The caliper measurements of skinfolds on women were less accurate than those of skinfolds on men. For women, the measurements on the back and chest were the most accurate (despite somewhat lower correlation coefficients), whereas for men, it was the measurements on the triceps, back, and thighs (Fig. 7).
      Fig. 7
      Fig. 7Percent of caliper values with a difference of ≤1 mm in comparison to values obtained using USG (our representation).
      Independent of gender and the site of the skinfold measured, agreement decreased with increasing thickness. Agreement was lower in women than in men (Table 3).
      Table 3Accuracy of measurements based on skinfold thickness.
      Skinfold thickness

      Independent of localization
      nMean difference

      USG − calipers (mm)
      Agreement

      (%)
      Independent of gender
       <20 mm242−0.0646.7
       ≥20 mm1030.5225.2
      Women
       <20 mm970.0342.3
       ≥20 mm830.1021.7
      Men
       <20 mm145−0.1049.0
       ≥20 mm202.340.0
      n: number of measured skinfolds without repetition (Σn = 5 skinfolds × 69 participants = 345), Accuracy: percent of measurements with a difference of ≤1mm between USG and caliper measurements.
      As presented in Table 2, in women, there was a significant difference in the mean value of measurements made via USG and those obtained using calipers for both the abdominal skinfold (P = 0.013, paired t-test) and the thigh skinfold (P = 0.002, paired t-test). We found the highest d values for the abdominal skinfold (Cohen's d = 0.20) and the thigh skinfold (Cohen's = 0.19), but the effect strength was small.
      For the men, there was a significant difference in the mean value of the USG and caliper skinfold measurements of the triceps skinfold (exact P = 0.008, paired t-test; effect size d = 0.20). The same applies to the thigh skinfold (exact P = 0.046, paired t-test; effect size d = 0.12).
      For a comparison of the two methods, the evaluation via the Bland–Altman plot is more robust than a significance test because all measurements are compared in pairs. A significant systematic error (mean offset with a 95% CI in the Bland–Altman plot) was found in the data on females for the abdominal fold and the thigh fold. A minor systematic error was found in the data on males for the triceps fold. The variability in the lines of agreement interval (LoA) (mean ± 1.96 SD, 95% of data are in this area) was lowest among the male subjects for the triceps (upper LoA = 3.3 mm, lower LoA = −2 mm), the back (upper LoA = 3.5 mm, lower LoA = −4 mm), and thigh fold (upper LoA = 2.2 mm, lower LoA = −3.2 mm); and among the female subjects, for the triceps (upper LoA = 4.6 mm, lower LoA = −4.8 mm), the back (upper LoA = 3.9 mm, lower LoA = −4.4 mm), and the chest folds (upper LoA = 5.8 mm, lower LoA = −4.6 mm). We found the highest variability among both males and females for the abdominal fold (female: upper LoA = 8.6 mm, lower LoA = −5.5 mm; male: upper LoA = 6.5 mm, lower LoA = −5.3 mm). With the exception of the thigh fold, trends were hardly or not at all present in men (values of caliper measurements became too high with increasing skinfold thicknesses) (Fig. 8).
      Fig. 8
      Fig. 8Bland–Altman plots (created using R [version 3.5.1] or R-Studio [version 1.2.1335]).

      4. Discussion

      This study confirms that there is good agreement between caliper and USG skinfold measurements. Some of the differences between the caliper and USG methods are attributable to the measurement position, gender, and skinfold thickness.
      There was a high correlation between the USG and caliper measurements for all skinfolds. The interrater reliability did not play a confounding role in our examination because both measurements were always performed by the same examiner, who also had sufficient experience in the correct use of the USG equipment. A good correlation was also confirmed in several recent studies [
      • Selkow N.M.
      • Pietrosimone B.G.
      • Saliba S.A.
      Subcutaneous Thigh Fat Assessment: A Comparison of Skinfold Calipers and Ultrasound Imaging.
      ,
      • Gomes A.C.
      • Landers G.J.
      • Binnie M.J.
      • Goods P.S.
      • Fulton S.K.
      • Ackland T.R.
      Body composition assessment in athletes: Comparison of a novel ultrasound technique to traditional skinfold measures and criterion DXA measure.
      ], although a lower correlation was found in other studies [
      • Müller W.
      • Horn M.
      • Fürhapter-Rieger A.
      • Kainz P.
      • Kröpfl J.M.
      • Maughan R.J.
      • et al.
      Body composition in sport: a comparison of a novel ultrasound imaging technique to measure subcutaneous fat tissue compared with skinfold measurement.
      ]. In our research, these high correlation coefficients could be highly indicative of multicollinearity or may be due to the small sample size.
      There was a significant difference in the mean of the USG and caliper measurements for the abdominal and thigh folds on women, and for the triceps fold on men. There were larger differences in the mean value for thicker skinfolds (≥20 mm), such as the abdominal and thigh folds in women; an observation also made by Selkow et al. [
      • Selkow N.M.
      • Pietrosimone B.G.
      • Saliba S.A.
      Subcutaneous Thigh Fat Assessment: A Comparison of Skinfold Calipers and Ultrasound Imaging.
      ]. The agreement also decreased with increasing skinfold thickness, such that the best agreement was achieved with skinfolds less than 10 mm thick. Cataldo et al. [
      • Cataldo M.G.
      • Brancato D.
      • Brancato G.
      • Verga S.
      • Buscemi S.
      • Licata G.
      Correlation between skinfold thickness and ultrasonography in the study of subcutaneous adipose tissue in females.
      ] described a good agreement between the two methods only for a skinfold thickness below 20 mm [
      • Cataldo M.G.
      • Brancato D.
      • Brancato G.
      • Verga S.
      • Buscemi S.
      • Licata G.
      Correlation between skinfold thickness and ultrasonography in the study of subcutaneous adipose tissue in females.
      ]. The thicker a skinfold, the more difficult it is to measure using calipers. There was a clear gender difference in this respect, but this could be due to a bias caused by the lower number of thick skin folds in the male gender.
      To compare the two methods, however, it is not sufficient to simply compare the central tendencies of the distributions. In the Bland–Altman plot, we searched for systematic errors (bias) and trends and analyzed the variability. Clinically relevant systematic errors were found only for the thicker skinfolds, such as skinfolds on the abdomen and thighs of the female subjects.
      We detected a trend in the thigh folds of men, with larger differences in the measurements at higher skinfold thicknesses. This trend was not observed in women. In contrast to other skinfolds, the thickness of the skinfolds on the thigh is generally overestimated using calipers, a point also noted by Selkow et al. [
      • Selkow N.M.
      • Pietrosimone B.G.
      • Saliba S.A.
      Subcutaneous Thigh Fat Assessment: A Comparison of Skinfold Calipers and Ultrasound Imaging.
      ]. On the thigh, the USG measurements can be especially useful as it is common and reliable [
      • Mechelli F.
      • Arendt-Nielsen L.
      • Stokes M.
      • Agyapong-Badu S.
      Validity of Ultrasound Imaging Versus Magnetic Resonance Imaging for Measuring Anterior Thigh Muscle, Subcutaneous Fat, and Fascia Thickness.
      ]. The thicknesses of skinfolds on the abdomen of women and those on the triceps of men, however, were underestimated when measuring with calipers. The variability in the Bland–Altman plot was greater for all skinfolds in the data on females than in the data on males, and it was highest for the skinfolds on the abdomen for both sexes. These gender-specific differences may be due to differences in the consistency of the tissue. In contrast to soft tissue, taut tissue is more resistant to pressure and is more difficult to form into a measurable skinfold. McRae [
      • McRae M.P.
      Male and female differences in variability with estimating body fat composition using skinfold calipers.
      ] described a difference in tissue compressibility between men and women, while Ward et al. [
      • Ward R.
      • Rempel R.
      • Anderson G.S.
      Modeling Dynamic Skinfold Compression.
      ] described a greater elasticity and lower viscosity of the skinfolds on females [
      • Ward R.
      • Rempel R.
      • Anderson G.S.
      Modeling Dynamic Skinfold Compression.
      ], most likely due to the different anatomy and biology of the fat tissue [
      • Chang E.
      • Varghese M.
      • Singer K.
      • Building M.P.
      • Arbor A.
      Gender and Sex Differences in Adipose Tissue.
      ]. In the data on males, the skinfolds on the triceps, back, and thigh had the best scores in the Bland–Altman analysis. In the data on females, the skinfolds on the triceps, back, and chest had the best scores. Nevertheless, it is important to point out the wide range of the LoA and that clinical acceptance must be assessed on a case-by-case basis. For the skinfolds on the abdomen and thigh, we certainly have clinically unacceptably wide LoA.
      In the case of USG measurements it would be obvious to exclude the skin from the measurement when measuring the subcutaneous fat tissue thickness. Since in our case we are making a comparison with the caliper skinfold measurements, the skin must be included in the measurement.
      With regard to muscle fascia, the USG also offers the option of including or excluding it in the measurement [
      • Mechelli F.
      • Arendt-Nielsen L.
      • Stokes M.
      • Agyapong-Badu S.
      Validity of Ultrasound Imaging Versus Magnetic Resonance Imaging for Measuring Anterior Thigh Muscle, Subcutaneous Fat, and Fascia Thickness.
      ]. In our study participants, the delineation of the outer layer of the muscle fascia from the subcutaneous fat tissue was usually good and therefore we excluded the muscle fascia from the USG measurement.

      4.1 Limitations of the study

      The wide range of participants' ages is certainly a limiting factor in this study. Unfortunately, age could not be eliminated as a confounder due to the limited number of participants. Nevertheless, the study strives to establish a balance in gender. Another limitation of the study is that the participants are normal-weight individuals. Therefore, the results cannot be extrapolated to the general population. Due to insufficient availability of resources, the use of reference methods such as CT scans and MRI to measure subcutaneous fat tissue were not employed. Potential sources of error in USG measurements should not be ignored. For instance, due to the non-standardized contact pressure of the USG probe, subcutaneous fatty tissue might be compressed to varied extents, and furthermore, it is challenging to delimit the subcutaneous fatty tissue from the muscle fascia, particularly if transducers with low resolution are used [
      • Störchle P.
      • Müller W.
      • Sengeis M.
      • Ahammer H.
      • Fürhapter-Rieger A.
      • Bachl N.
      • et al.
      Standardized Ultrasound Measurement of Subcutaneous Fat Patterning: High Reliability and Accuracy in Groups Ranging from Lean to Obese.
      ].
      Regardless of the examination results, we should not forget that calipers have a higher ease of use and availability when compared to USG and are therefore more prevalently used in the field. Nowadays USG is easily available in every hospital, but not all general practitioners and coaches have USG equipment. The development of measurement technology is progressing rapidly; thus, it is possible that newer USG probes that work along with smartphones will catch up to the superior practicability of calipers.

      5. Conclusion

      The thicker the skinfold, the greater the difference between the caliper and USG measurements. However, compared to the USG measurements, the reliability of the caliper measurements also depended on the site of the measured skinfolds.
      USG measurements differs from caliper measurements especially in the skinfolds on the abdomen and thigh. Best comparable sites in our collective are the skinfolds on triceps, back and chest.

      Credit author statement

      Hannes Nösslinger: Conceptualization, Methodology, Formal Analysis, Investigation, Data Curation, Writing (Original Draft).
      Ewald Mair: Conceptualization, Methodology, Formal Analysis, Investigation, Data Curation, Writing (Original Draft).
      Hermann Toplak: Supervision.
      Marlies Hörmann-Wallner: Writing (Review & Editing), Supervision.
      This research paper was written as part of a Master's degree thesis for the Department of Applied Nutritional Medicine at the Medical University of Graz and the FH Joanneum University of Applied Sciences, Graz. The study was supervised by Dr. Marlies Hörmann-Wallner ([email protected]) and Prof. Hermann Toplak ([email protected]).

      Funding

      This research did not receive any specific grants from funding agencies in the public, commercial, or not-for-profit sectors.

      Conflicts of interest

      None of the authors has a conflict of interest to declare.

      Acknowledgments

      We thank all participants in the study and our supervisors who contributed to making our study a success.

      Appendix A. Supplementary data

      The following is the Supplementary data to this article:

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