Live hands-on demonstration of the iNSiGHT, DXA imaging system for body composition analysis. During this live demonstration, we showed the full imaging and measurement capabilities of the iNSiGHT system. These measurements included: Bone mineral density in g/cm2 Bone mineral content in g Bone area in cm2 Tissue area in cm2 Fat tissue as percentage and weight in % and g Lean tissue as percentage and weight in % and g Total weight g Additionally, specific bone length measurements can be drawn on the 2D x-ray image The iNSiGHT is a fully functional DXA (DEXA, dual energy x-ray absorptiometry) system, designed specifically for preclinical research applications. The DXA technology provides quantification of body composition, such as bone mineral density, and measures of lean and fat mass.

1. Live Virtual
Demonstration
iNSiGHT
Preclinical DXA system for in
vivo body composition and
bone mineral density
measurements

2. Tonya Coulthard, MSc
Manager, Imaging Division
Scintica Instrumentation
Val Fajardo, PhD
Canadian Research Chair – Tissue Remodeling and Plasticity
Assistant Professor
Brock University

3. WWW.SCINTICA.COM
Topics of Discussion
• What is DXA and How Does it Work
• iNSiGHT System Overview
• Live Virtual Demonstration – *animal change normal mice*
• Acquiring Live Data, and data analysis review
• Previously acquired data – discussion with Dr. Val Fajardo
• Looking forward to next live demonstration on June 9, 2021
• Muscular dystrophy model – D2 mdx
• Q&A Session
3

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Audience Poll

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What is DXA and How Does it Work
• Dual Energy X-Ray Absorptiometry
• Acquired Images
• Available Measurements
• Comparison to Other Techniques
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Dual Energy X-Ray Absorptiometry (DXA or DEXA)
• DXA is used to assess body composition
• Different tissues in the body have varying mass attenuation
coefficients (µn)
• Two different x-ray beams, having different energies are
generated at the source, passing through the body, hitting
the x-ray detector
• Equations are used to determine which type of tissue each
pixel on the image represents
• Bone
• Soft tissue – fat mass or lean tissue mass
Figure from Luo, Yunhua. 2017. Chapter 3 – Bone Imaging for
Osteoporosis Assessment
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Acquired Images
Images acquired using the iNSiGHT DXA system 7
X-Ray Attenuation
Image
Bone Mineral Density
Image
Color Image
Fat (Orange) and Lean (Green) Mass

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Available Measurements
8
Parameter
Unit of
Measure
Description
(available on whole animal, or from each ROI)
BMC g
Bone Mineral Contents (Bone Mass)
BMC = bone density x bone area
Fat g Fat mass
Fat Ratio % Fat Ratio = Fat/Total Mass
Lean g Fat free mass
Lean Ratio % Lean Ratio = Lean/Total Mass
Total Mass g Total Mass = Fat + Lean + Bone
BMD g/cm2 Bone Mineral Density
Bone Area cm2 Bone Area in Image
Tissue Area cm2 Tissue Area in Image
Images acquired using the iNSiGHT DXA system

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Comparison to Other Techniques
9
DXA NMR (Echo MRI)
Measurements
Available
• Fat Mass
• Lean Mass
• Bone Mineral Content
• Fat Mass
• Lean Mass
• Water Content
Scan Time • 25 seconds
• 80 seconds
• 30 seconds without water
content
Advantages
• Bone/lean/fat and weight
measurements
• Images acquired
• High degree of accuracy
(CV < 1%, R2 > 0.9)
• Water content available
• No anesthesia required
Disadvantages • Requires anesthesia
• No bone or weight
information
• No images available
DXA
NMR

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iNSiGHT System Overview
• System Components
• System Features and Benefits
• Technical Specification
• Analysis Software
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iNSiGHT: System Components
• X-Ray Benchtop Cabinet
• Self-shielded
• Integrated anesthesia
• Standard electrical connection
• Software Workstation
• Windows 10 operating system
• Offline Analysis Software
• License based for offline analysis

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iNSiGHT: System Features and Benefits
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• Longitudinal studies
• Non-invasive data acquisition allows studying changes in body composition
over time
• Easy data acquisition
• No preparation steps other anesthesia required
• Low dose radiation
• Minimizing effects on animals during each imaging session
• Fast scan times (~25 seconds/scan)
• Allows for high throughput studies
• High-resolution images (100µm)
• Study changes in body composition on small animals, incl. mice, rats, etc.
• Wide scan area (16.5 x 25.5 cm)
• Allows for a variety of animal models to be imaged, from 10 ~ 500g

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iNSiGHT: Technical Specifications
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Specification Details
Scan Method Cone Beam
Animal Size 10 ~ 500g
Scan Time 25 seconds
Image Area (maximum) 16.5 x 25.5cm
Pixel Size
100µm at 1.2X
31µm at 4X
Operating System Windows 10, 64bit
Dimensions (WxDxH) 66 x 61 x 113 cm
Power 110/240VAC, 50/60Hz, 200VA
Operating Temp 10 ~ 40oC

14. Hands On Demonstration
of iNSiGHT DXA System

15. iNSiGHT Small Animal DXA
Coefficient of variation + low-dose lithium and body composition
Val Fajardo, May 19, 2021

16. Main uses
 Body composition (fat and lean mass) – obesity related studies,
muscular dystrophy (June 9)
 Couple with metabolic cage data
 Bone structure
 Bone mineral density and content – real and simulated microgravity, and the
effects of GSK3 inhibition (i.e., lithium) on osteogenic signaling
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17. CVs – female mouse
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18. CVs – male mouse
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19. Placement CV – male moved 6x
 Male mouse, measured 6x where the mouse was repositioned before
each scan
19
Number of values
Mean
Std. Deviation
Std. Error of Mean
Coefficient of variation
BMD (g/cm2)
6
0.06550
0.001049
0.0004282
1.601%
BMC (g)
6
0.4985
0.01540
0.006286
3.089%
Bone Area (cm2)
6
7.624
0.1570
0.06411
2.060%
Tissue Area (cm2)
6
18.37
0.3475
0.1419
1.891%
Fat (%)
6
15.60
0.1285
0.05245
0.8234%
Fat (g)
6
4.630
0.05465
0.02231
1.180%
Lean %
6
83.00
0.1595
0.06512
0.1922%
Lean(g)
6
25.04
0.1133
0.04626
0.4525%
Total Weight(g)
6
30.17
0.1593
0.06501
0.5279%

20. Male rat (3-4 months old)
20
Non-repositioned CV
RepositionedCV
Number of values
Mean
Std. Deviation
Std. Error of Mean
Coefficient of variation
BMD (g/cm2)
7
0.1799
0.0003780
0.0001429
0.2101%
BMC (g)
7
5.463
0.01793
0.006777
0.3282%
Bone Area (cm2)
7
30.36
0.1376
0.05199
0.4531%
Tissue Area (cm2)
7
108.2
0.2582
0.09759
0.2386%
Fat (%)
7
11.62
0.1153
0.04360
0.9929%
Fat (g)
7
33.53
0.3294
0.1245
0.9825%
Lean %
7
86.74
0.1110
0.04197
0.1280%
Lean(g)
7
255.1
0.3781
0.1429
0.1482%
Total Weight(g)
7
294.1
0.1301
0.04916
0.04423%
Number of values
Mean
Std. Deviation
Std. Error of Mean
Coefficient of variation
BMD (g/cm2)
7
0.1769
0.002035
0.0007693
1.151%
BMC (g)
7
5.340
0.07211
0.02725
1.350%
Bone Area (cm2)
7
30.18
0.1727
0.06529
0.5723%
Tissue Area (cm2)
7
107.4
0.6508
0.2460
0.6062%
Fat (%)
7
11.61
0.1119
0.04228
0.9634%
Fat (g)
7
33.71
0.2540
0.09599
0.7535%
Lean %
7
86.79
0.1206
0.04559
0.1390%
Lean(g)
7
256.6
1.251
0.4728
0.4875%
Total Weight(g)
7
295.6
1.118
0.4226
0.3782%

21. Low-dose lithium and body composition
 Project in collaboration with Dr. Rebecca
MacPherson (Associate Professor, Dept.
Health Sciences)
 Stimulate energy expenditure to combat
combat obesity
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22. Low-dose lithium ↑ energy expenditure
 10 mg/kg/day (serum concentration of 0.02 mM) for 6-12 weeks
 Dose shown to reduce high-fat diet induced weight gain (Choi et al., 2010Vascular Pharmacology, 53: 264-272)
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Geromella et al., unpublished data

23. Does this translate to a change in body
composition?
23
Geromella et al., unpublished data
Will re-assess at 12 weeks of feeding. *non-invasive longitudinal studies

24. What happens when we adjust energy
expenditure to fat free mass?
24
Meunier et al., 2005 Eur J Clinical Nutrition

25. Low-dose lithium still ↑ energy expenditure
 Even after normalizing to FFM.
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Geromella et al., unpublished data
D
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fat-free
mass/min)
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26. June 9 OnlineWebinar
 Duchenne muscular dystrophy and body composition using the D2
mdx mouse model
 Lean mass and bone structure (BMD and BMC)
26

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Audience Poll

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Q&A Session
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INFO@SCINTICA.COM
Please enter your questions in
the Q&A section
Tonya Coulthard,
MSc
Manager, Imaging
Division,
Scintica
Val Fajardo, PhD
Canadian Research Chair –
Tissue Remodeling and Plasticity
Assistant Professor,
Brock University

29. Globally linking scientists with
precision tools for research
through expertise in science,
engineering and support

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Thank You
WWW.SCINTICA.COM
INFO@SCINTICA.COM

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iNSiGHT: Analysis Software
31
Parameter
Unit of
Measure
Description
(available on whole animal, or from each ROI)
BMC g Bone Mineral Contents (Bone Mass)
BMC = bone density x bone area
Fat g Fat mass
Fat Ratio % Fat Ratio = Fat/Total Mass
Lean g Fat free mass
Lean Ratio % Lean Ratio = Lean/Total Mass
Total Mass g Total Mass = Fat + Lean + Bone
BMD g/cm2 Bone Mineral Density
Bone Area cm2 Bone Area in Image
Tissue Area cm2 Tissue Area in Image
The image can be magnified to help
draw specific ROIs, or to make length
measurements on any bone

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iNSiGHT: Analysis Software
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• Up to 7 ROIs can be drawn to get specific
information
• An exclusion ROI (blue) can be drawn
and applied to all calculations
• Length measurements can be completed
on any bone

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iNSiGHT: Analysis Software
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• Results from a longitudinal study can be
displayed over time
• Both trendlines and graphical
representation of data is available
• All data can be exported in .csv format
• Images can be exported in standard
formats

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Key Research Applications
• Example Images
• Research Areas of Interest
• Key Studies
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Example Images - Mouse
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X-Ray Attenuation Image Bone Mineral Density Image
Color Image
Fat (Orange) and Lean (Green) Mass

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Example Images - Rat
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X-Ray Attenuation Image Bone Mineral Density Image
Color Image
Fat (Orange) and Lean (Green) Mass

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Example Images - Other
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Research Areas of Interest
• DXA, resulting bone mineral density
and body composition data can be
used in a wide variety of research
areas
• The iNSiGHT system can be used to
assess the progression or regression
of disease in response to a
therapeutic regime
• Bone Mineral Density:
• Metabolic bone diseases, such as
osteoporosis, etc.
• Arthritis
• Musculoskeletal diseases
• Drug safety and toxicology
• Lean/Fat Mass:
• Metabolic disorders, such as diabetes,
obesity, etc.
• Musculoskeletal diseases
• Drug safety and toxicology
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Key Studies – Body Composition Analysis
• Body composition was analyzed in both
mice and rats over time
• X-Ray images were used to visualize
skeletal and organ structures
• Bone Segmentation images were used to
check posture and confirm results
• Color Composition images were used to
monitor lean and fat mass over time
39
2019 KALAS International Symposium

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Key Studies – Obesity Study in Rats
• This study showed the iNSiGHT system could be
used to study changes in body composition over
time
• Rats fed normal and high fat diets over an 8-week study
• Accuracy of DXA weight results was assessed by
comparison to electronic scale measurements
• Precision was assessed by measuring the coefficient
of variation (CV) by making repeated measurements
at week 8
40
Yeu, Jungyun et al. Laboratory Animal Research (2019) 35:2

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Key Studies – Obesity Study in Rats
• Precision was assessed by measuring the coefficient
of variation (CV) by making repeated measurements
at week 8
• Accuracy of DXA weight results was assessed by
comparison to electronic scale measurements
41
Yeu, Jungyun et al. Laboratory Animal Research (2019) 35:2
• Change in total body weight (TBW), total body fat
weight (TBFW), and total body lean weight (TBLW)
is shown over time in high fat diet and normal diet
• Difference between diets is shown in the bar graphs
over time

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Key Studies – DXA vs NRM (Echo MRI)
• Comparison of body composition analysis results
between DXA and NMR (echo MRI), verifying both
precision and accuracy
• Results were compared between modalities and to
autopsy results
• Authors found a higher level of precision and
accuracy in the DXA measurements compared to
NMR (echo MRI), along with additional bone
mineral density measurements available using DXA
and not with NMR
42
Baek, Kyung-Wan et al. Journal of Bone Metabolism. (2020) 27:4

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Key Studies – DXA vs NMR (Echo MRI)
• Accuracy of results between systems was compared by
measuring Fat Mass
• No difference was seen between euthanized and
anesthetized animals using DXA, a difference was
noted when using NMR
• Body Weight and Bone Mineral Content were only
available on DXA and showed very strong correlation with
the reference measured on electronic scale
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Baek, Kyung-Wan et al. Journal of Bone Metabolism. (2020) 27:4
• Precision was assessed for all
available measurements on each
modality
• Bone mineral content, body
weight, and femur length
measurements are not available
using NMR
• RE results are from euthanized
animals, RA results are from
anesthetized animals,
repositioned between each
measurement