The authors would like to acknowledge the editorial reviewers as well as the two anonymous reviewers. Their suggestions and critiques were valid, much appreciated and vastly improved the original manuscript.

The protocol herein adheres to the North Carolina State University&#39;s ethics guidelines for human research.
1. Machine Preparing
NOTE: The following protocol can be broadly applied to any whole body, clinical DXA and BMD scanner.
<ol>
	<li>Perform calibration once daily prior to scanning any individuals to ensure quality control. After calibration prompts appear upon start-up of the systems&#39; software, scan a lumbar spine phantom of known density to ensure c...

The authors declare no competing financial interests.

The results presented in this paper are illustrative of the applicability of BMD metrics in forensic contexts. As Figure 6 and Figure 8 show, the scanning position of living individuals for clinical BMD scans is reproducible with skeletal remains, but care must be taken to ensure proper positioning. This is especially critical for the hip examination where identifying the midline of the femoral neck require the proper angle of the femur and overestimation of BMD...

The objective of forensic anthropological analyses relies on the practitioner&#39;s understanding of bone as a complex tissue with multiple units and variation. Bone is a hierarchical, composite tissue with both organic and inorganic components organized into a matrix of collagen and carbonated apatite1,2,3,4. The inorganic component, or bone mineral is organized in a nanocrystalline structure to provide stiffness and framework for the organic portion1,2,5. The mineral aspect comprises approximately 65% of bone by weight and its&#39; mass is influenced by both genetic and environmental factors1,2,4,6. Because bone mineral occupies a three-dimensional space, it can be measured as bone mineral density (BMD), or a function of the mass and the volume occupied7. The bulk density of bone mineral varies with age from birth into adulthood8,9,10,11,12 and has been used extensively in clinical settings as an indicator of osteoporosis and fracture risk4,13,14,15,16,17,18. Dual-energy X-ray absorptiometry (DXA) has been a widespread tool for the assessment of bone health since its introduction in 1987, particularly scans performed in the lumbar spine and hip regions11,13,19. Validation of DXA scans has been shown as the gold standard when investigating changes in BMD13,19,20,21,22,23. Subsequently, the World Health Organization (WHO) has created normative standards including t- and z-score definitions for juvenile and adult lumbar spine (L1-L4) and hips as these are the regions easily captured volumetrically11,13,19,24.
The increasing reliance on forensic anthropology in medicolegal casework has encouraged the investigation of novel techniques to better assess skeletal remains in a variety of circumstances. Among these potential techniques is the application of DXA scans to assess BMD as an indicator of bone quality in cases involving fatal starvation and neglect in juveniles25,26, identification of metabolic bone diseases, and estimating survivability of skeletal elements in taphonomic research7,27.
In the 2015 U.S. Department of Health and Human Services Child Maltreatment Report, 75.3% of the reported child abuse cases were some form of neglect with ~1,670 fatalities resulting from fatal starvation and neglect in 49 states28. Most juvenile victims of neglect fail to show signs of external physical abuse, but failure-to-thrive is seen in all cases29,30. Failure-to-thrive is defined as the inadequate nutrition intake to support growth and development. These can have different factors, one of which is neglect resulting from nutritional deprivation25,31 (see Ross and Abel32 for a more comprehensive review). Deliberate starvation that results in the death of a child or infant is much rarer and considered as the most extreme form of maltreatment25,33,34. These nutritional deficiencies have a significant effect on bone growth, particularly longitudinal growth in children as an immediate consequence of malnutrition35. Skeletal growth and mineralization primarily depend on vitamin D and calcium, and their supplementation has been linked to increased BMD25,35,36.
It is exceedingly difficult to identify or prosecute these cases even following a complete autopsy31,37,38 and special consideration to methods employed must be used. Thus, in cases where fatal starvation or malnutrition is suspected, a multidisciplinary approach is needed particularly in cases involving remains in advanced states of decomposition26. When skeletal remains are involved, bone densitometry is a useful tool in conjunction with other skeletal indicators such as dental development, measurement of the pars basilaris of the skull, and long bone lengths26. Without using the skeletal indicators mentioned above for infants and juveniles, it would not be possible to discern if low BMD is the result of an inherent metabolic disorder, malnutrition, or taphonomic process. Another concern is the estimation of body size (weight and stature) in infant or juvenile skeletal remains. Most normative data sets require information about height or weight for comparison purposes as bone growth in children is size and age dependent12. When the remains being assessed are unidentified, estimation methods should be employed. For infants under one, normative DXA data is age matched only. In juveniles over the age of 1, Ruff39 or Cowgill40 are recommended for estimating body size in skeletal remains as they are based on the Denver Growth Study sample including ages 1 - 1739,40. When age and body size are estimated, confidence intervals vary and comparison of the mean to the Center for Disease Control (CDC) produced growth curves41 should be included in the report as well as the confidence interval for the estimated body size. It is important to note that in most cases, information regarding ancestry and sex cannot be determined from juvenile skeletal remains prior to puberty, which is particularly important for adolescents as ancestry and sex are known to significantly impact BMD in adults. In these circumstances, the DXA method may not be applicable. In identified cases, biological information regarding ancestry, sex, and body size, should be obtained prior to analysis.
Bone densitometry in pediatrics has increased with the development of normative data42,43 with DXA being the most widely available technique44. Malnourished children show significantly lower levels in BMD than healthy children with mineralization correlated with severity of malnutrition45. DXA scans of the lumbar spine and hips are the most appropriate areas to assess for juveniles according to The American College of Radiology46. Reproducibility has been shown for spine, whole hip, and whole body in children throughout the growth period47. However, the lumbar spine is preferred as it is primarily composed of trabecular bone, which is more sensitive to metabolic changes during growth and has been found to be more precise than whole hip assessments25,47,48. Using DXA scans is common in pediatric assessment. However, since DXA is two-dimensional, it does not capture true volume and produces a BMD based on bone area13. In children, this is an important distinction as body and bone size vary within and between age groups in children12. Most normative data available is for comparison with DXA measurements, but care should be exercised to choose an appropriate reference population (see Binkovitz and Henwood13 for a list of commonly used DXA normative databases).
Following the scan, a z-score is calculated using an age-matched and population specific reference sample. Z-scores are more appropriate for juveniles since t-scores compare the measured BMD to a young adult sample12. A z-score between -2 to 2 indicates normal BMD for chronological age while any score below -2 indicates low BMD for chronological age49. The -2 to 2 range for both the t- and z-score represent up to two standard deviations from the mean. Plainly, if a measured BMD score is within two standard deviations above or below their reference population mean, they are considered clinically normal.
The reliance on morphological variation for the forensic anthropologist comes from many sources. One of which is the skeletal variation that arises from disease processes, including metabolic bone disorders50. The ability to identify specific disorders in skeletal remains has a two-fold advantage: 1) adding information to the biological profile making it more robust and 2) identifying if fractures are pathological or the result of inflicted trauma. There are a variety of metabolic bone disorders51,52,53, but the most relevant for BMD measures of contemporary remains is osteoporosis. Osteoporosis develops when the rate of trabecular bone loss is greater than the rate of cortical bone loss with a net loss in bone density53,54,55. Trabecular bone loss is correlated to an increased risk of fracture, especially in bones that have greater trabecular bone content (e.g., the os coxa)4,55.
Numerous studies on osteoporosis and bone mineral density in skeletal remains have been conducted on archaeological assemblages using both DXA56,57,58,59 and other methods60,61,62. However, when assessing osteoporosis in the adult skeleton from archaeological contexts, practitioners disregard that diagnosing osteoporosis clinically requires the mean of a younger reference sample contemporaneous with the individuals being assessed55,63,64. This is not an issue in forensic anthropology contexts since individuals are age- and sex- matched to modern populations with developed reference samples for both the hip and the lumbar spine, although changes in BMD through diagenesis should be considered for forensic remains. However, taphonomy is the likely factor affecting the ability to obtain legitimate BMD measures from archaeological samples. This is a consideration in forensic contexts as well, where remains recovered from burial conditions with potential postmortem intervals beyond a few months. While still of forensic interest, sufficient doubt could be raised for any BMD scores obtained from remains found in these circumstances.
Osteoporosis is clinically assessed using t-scores of BMD measures that are derived from the individuals&#8217; BMD measures in the hip or lumbar spine relative to a young adult reference sample using DXA65,66,67,68. This reference sample can be employed for identifying the occurrence of osteoporosis in the skeleton. In forensic contexts, this is useful for two reasons: 1) differentiating between fractures related to abuse-inflicted trauma in the elderly and those from increased bone fragility in osteoporotic individuals69, and 2) as a possible personal identification feature50.
Bone density has long been considered an indicator that reflects the activity and nutrition of an animal70,71. More recently it has been noted that bone density, as an intrinsic property of bone, affects its survivability during taphonomic processes7.&#160; A consequence of decomposition is the differential survivability of skeletal elements (i.e., discrete, anatomically complete units of the skeleton) and bone density can be used as a predictor of survivability, or bone strength7,70,71,72,73,74,75. This is important in forensic contexts as well as archaeological and paleontological environments in that it affects the practitioners&#8217; ability to adequately employ methods to estimate a biological profile (or age, sex, stature, and ancestry) if only certain skeletal elements are represented.
Bulk density (bone density with pore space included in the measurement) is the appropriate measurement in this situation, considering it is precisely the porous structure of bone that influences its susceptibility to taphonomic processes7. Many methods of assessing bone density have been employed including single-beam photon densitometry27,75, computed tomography76,77,78, photodensitometry72,79, and DXA80,81,82. DXA scans may be preferable to other methods as it is relatively inexpensive, whole body scans can be performed, and individual skeletal elements can be assessed separately or together during analysis. Using BMD scans before and after taphonomic research studies provides useful information on bone survivability resulting from different taphonomic factors and environments82.
This paper outlines a protocol for obtaining DXA scans of skeletal remains. The method employs the common, clinical positioning of individuals when performing lumbar spine and hip scans. This allows practitioners to compare the skeletal remains with the appropriate normative standards. The protocol outlined is applicable to both juvenile and adult remains with limitations discussed later.

<table border="0" cellpadding="0" cellspacing="0" width="869">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">QDR Discovery 4500W system</td>
			<td style="width:71px;">Hologic</td>
			<td style="width:119px;">Discovery W</td>
			<td style="width:463px;">All inclusive DXA whole body scanner that includes APEX software for visualization and analysis of scans. Incorporates FRAX reference data developed by WHO to provide both t- and z- scores.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">APEX 3.2</td>
			<td style="width:71px;">Hologic</td>
			<td style="width:119px;">APEX</td>
			<td>Software used by the DXA PC connected to the bone desitometer (QDR Discovery 4500W system) to acquire the BMD data and analyze results.</td>
		</tr>
	</tbody>
</table>

scanning skeletal remains for bone mineral density in forensic contexts

The purpose of this paper is to introduce a promising, novel method to aid in the assessment of bone quality in forensically relevant skeletal remains. BMD is an important component of bone&#39;s nutritional status and in skeletal remains of both juveniles and adults, and it can provide information about bone quality. For adults remains, it can provide information on pathological conditions or when bone insufficiency may have occurred. In juveniles, it provides a useful metric to elucidate cases of fatal starvation or neglect, which are generally difficult to identify. This paper provides a protocol for the anatomical orientation and analysis of skeletal remains for scanning via dual-energy X-ray absorptiometry (DXA). Three case studies are presented to illustrate when DXA scans can be informative to the forensic practitioner. The first case study presents an individual with observed longitudinal fractures in the weight bearing bones and DXA is used to assess bone insufficiency. BMD is found to be normal suggesting another etiology for the fracture pattern present. The second case study employed DXA to investigate suspected chronic malnutrition. The BMD results are consistent with results from long bone lengths and suggest the juvenile had suffered from chronic malnutrition. The final case study provides an example where fatal starvation in a fourteen-month infant is suspected, which supports autopsy findings of fatal starvation. DXA scans showed low bone mineral density for chronological age and is substantiated by traditional assessments of infant health. However, when dealing with skeletal remains taphonomic alterations should be considered before applying this method.

The methodology proposed here is commonly used in living patients and consideration of its novelty to deceased individuals should be noted. Figure 6 and Figure 8 present the results of an AP lumbar spine and left hip scan, respectively. The individual assessed in these scans is a deceased white, female, 31 years of age that is housed at the Forensic Analysis Laboratory of North Carolina State University. This individual had a tot...

Watch this Scientific Journal Video about Scanning Skeletal Remains for Bone Mineral Density in Forensic Contexts at JoVE.com

Scanning Skeletal Remains for Bone Mineral Density in Forensic Contexts

Bone mineral density (BMD) is an important factor in understanding nutritional intake. For human skeletal remains, it is a useful metric to assess quality of life in both juveniles and adults, particularly in fatal starvation and neglect cases. This paper provides guidelines for scanning human skeletal remains for forensic purposes.

The purpose of this paper is to introduce a promising, novel method to aid in the assessment of bone quality in forensically relevant skeletal remains. ...

The overall goal with this procedure is to provide information about bone quality in forensically relevant remains. This method can help answer pertinent questions for forensic anthropologists such as non-destructive assessments of juvenile fatal starvations and neglect cases. This method can also differentiate between elder abuse and pathologies, as well as distinguishing taphonomic artifacts from pathology or trauma.The main advantage of this technique is that it is reproducible and provides a quantitative assessment that can be substantiated with clinical standards. To begin this procedure, prepare the machine as outlined in the text protocol. Next, create a new patient profile for each new individual to be scanned to maintain chain of custody and to ensure scans are correctly associated.If the individual being scanned is unidentified, establish the biological profile prior to scanning. Enter the demographic information into the patient profile, including the estimated stature if the actual stature is unknown. For anterior-posterior lumbar spine scans, select Perform Scan, choose Patient, select Scan Type, AP Lumbar Spine, and then Next.Then select a container that is at least as large as the articulated segment of L1 through L4.Fill the bottom of the container with rice, which will act as a soft tissue proxy. Place the segment in anatomical position into the rice with approximately 0.7 centimeters between each vertebral body. Make sure that the superior and inferior articular facets are articulated, but that the vertebral bodies are not in contact with one another.Next, center the scanning table. Place the container on the table so that L1 is oriented toward the head of the scanning table and L4 is placed one centimeter superior to the intersecting cross hairs. The vertical laser line should be bisecting the vertebrate bodies of all four vertebrae.After this, cover the exposed bone with rice. Following the scan, an Exit Exam prompt box will appear. Select Analyze Scan.In the Scan Analysis window, select Results. If the vertebrae were not properly separated, select Vertebral Lines to make minor adjustments. Then collect the results graph for visualization of the individual relative to the reference population.To perform a left or right hip scan, first select Perform Exam, choose Patient, select Scan Type, Left Hip or Right Hip, and then Next. Select an open container that is at least as large as the articulated os coxa and femur being scanned. Then fill the bottom of the container with rice.Place the ox coxa into the rice with the acetabulum and obturator foramen facing laterally and the pubic bone oriented medially. Position the ischial tuberosity beneath the femoral head as it articulates with the acetabulum. Next, place the femurs such that the femoral head is in the acetabulum with the greater trochanter and femoral head in line parallel to the scanning table.Make sure that the femoral shaft is medially rotated with the lateral condyle rotated medially and slightly higher than the medial condyle. Then center the scanning table. Move the scanning arm and table until the laser cross hairs are oriented with the center directly above the subtrochanteric area of the femur.The laser's vertical line should bisect the top half of the femoral shaft. After this, cover the remaining visible portion of the femoral acetabular joint with rice. Select Start Scan.An Exit Exam prompt will appear after the scan is completed. Select Analyze Scan. Next, select the Bone Map tool to add or delete areas that are not part of the femoral neck and trochanteric region.Select the Neck tool and reposition the midline to realign adjustments directly on the scan. In the Scan Analysis window, select Results. For adults, compare the scan to re reference data for the femoral neck, trochanteric region, and intertrochanteric region.In this study, skeletal remains are scanned for use in forensic context. Representative scans show that the scanning positions typically used for living individuals for clinical BMD scans can be reproduced with skeletal remains. The first case studies of a 40-year old male who exhibited a rare fracture series.A DXA scan of the left hip is obtained to determine if these fractures were the results of fracture fragility or simply a post-mortem artifact from natural drying processes. The determined total BMD is 1.299 grams per square centimeter with the corresponding T-score of 1.8, indicating that bone insufficiency was not the cause. The second case study is a 13-year old girl with a suspected history of long-term abuse, recovered from a clandestine grave.The total BMD is 0.660 grams per square centimeter with the Z-score of 2.2. The Z-score is consistent with the low BMD for chronological age, supporting other methods suggesting chronic malnutrition. The third and final case study is a 14-month old infant with starvation as the suspected cause of death.The total BMD of the lumbar spine is measured at 0.190 grams per square centimeter, while the age-matched reference group's average is approximately 0.399 grams per square centimeter. The Z-score is calculated to be 5.225 with the relevant age-matched mean. These low values suggest that the infant is below the third percentile of spine BMD for 12-month olds.Once mastered, this technique can be done in one hour if it is performed properly. While attempting this procedure, it's important to remember that anatomical orientation is the primary determinate for a successful scan. After watching this video, you should have a good understanding of how to adapt clinical BMD to forensic cases.

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17.3K Views.  North Carolina State University. The overall goal with this procedure is to provide information about bone quality in forensically relevant remains. This method can help answer pertinent questions for forensic anthropologists such as non-destructive assessments of juvenile fatal starvations and neglect cases. This method can also differentiate between elder abuse and pathologies, as well as distinguishing taphonomic artifacts from pathology or trauma.The main advantage of this technique is that it is reproducible and ...