**2. Principles of micro-CT**

This chapter covers the basic principles of micro-CT. *Ex vivo* specimens are typically placed on a rotating stage between the X-ray source and the microscope objective, which is followed by a detector (Fig. 1). For high resolution imaging, the sample size must be reduced to a minimum. The dimensions should not exceed 500-1000 times the resolution limit required. In large samples, the X-rays must penetrate more material, which results in a lower photon count and increased exposure time.

Certain devices operate with geometrical magnification only, in which the resolution increases with the distance between the sample and the detector. Unfortunately, increased geometrical magnification can result in blurriness, depending on the X-ray source spot size.

© 2012 Tonar et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2012 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**Figure 1.** Configuration of a micro-CT scanner with an *ex vivo* sample rotating within a stationary X-ray system (redrawn and modified according to Jorgensen et al., 1998).

More sophisticated devices use microscope objectives for increased magnification and resolution. For optimal settings of the micro-CT scan, the spot size (SS) and the awaited pixel size (PS) are the parameters that set the source distance (SD) and detector distance (DD). When the SD and DD are smaller, higher photon counts can be achieved, thus reducing the time costs. The settings should fulfill equation 1; see also Fig. 2:

$$PS \ge SS \cdot \frac{DD}{SD} \tag{1}$$

Correlating Micro-CT Imaging with Quantitative Histology 175

details smaller than 1 µm, 24 hours or more might be necessary, whereas an overview scan with a pixel size of approximately 10 µm can be achieved within an hour. Sample drifting might be an issue during a long scanning time. From the reconstructed images, the objects of

Compared with standard human CT devices, which offer a resolution limit of approximately 0.4 mm, the micro-CT introduced a promising modality. However, the clinical use of this method is limited by its higher radiation exposure and longer scanning times. It is used either to visualize individual fine functional and anatomical structures of *ex vivo* human or animal organs (e.g., liver lobules or bone trabeculae) or for whole-body imaging of small animals (Schambach et al., 2010). *In vivo* micro-CT systems are based on a rotating system of X-ray tube and detectors. The construction of these devices is the same as in human CT, except that their dimensions are adapted to small animals (Bag, 2010). The minimal space resolution of *in vivo* micro-CT is from 100 to 30 µm. The imaging of living animals must be faster than in *ex vivo* micro-CT. It is enabled by, among other factors, the use of flat panel detectors that allow us to acquire an abundance of thin sections during one rotation. The examination is also limited by the necessity of using a radiation dose that does not harm the tested animal. In *ex vivo* micro-CT, the X-ray source and the detectors are stationary, and it is possible to adjust the distance between the X-ray tube and the detectors (based on the size of the examined object) to improve the spatial resolution and minimize artifacts. Moreover, in *ex-vivo* micro-CT, time resolution is not important; thus, the examination may take a very long time (hours), and any amount of radiation may be used. This technique enables us to acquire much higher spatial resolution than *in vivo* micro-CT

Most organs have already been analyzed with micro-CT, including bones (Peyrin, 2011), heart and blood vessels (Schambach et al., 2010), lungs, kidney, liver, and cerebral structures (Schambach et al., 2010). Micro-CT devices can be used for the characterization of bone or vascular microarchitecture (Peyrin, 2011; Burghardt et al., 2011; Missbach-Guentner et al., 2011). This method also allows the precise detection of the margins of tumors and their vascularity (Ma et al., 2011; Missbach-Guentner et al., 2011). Tissue composition (e.g., bone mineralization) can be directly linked to 3-D tissue morphometry (Burghardt et al., 2011). Thus, 3-D micro-CT analysis becomes a method of choice for describing the spatial complexity of organ segmentation and the relationships between morphological and functional units (e.g., hepatic lobules and portal acini) (Schladitz, 2011). Micro-CT can also link the imaging of anatomical structures with functional and molecular imaging, e.g., tissue and organ perfusion, the flow rate of exocrine secretions within parenchymatous organs and

For the examination of soft tissues and vessels, contrast solutions are necessary. In *in vivo* imaging, such as in human medicine, standard iodinated contrast media or intravascular blood-pool contrast agents are used (e.g., contrast material covered by a polyethylene glycol

glands (Marsen et al., 2006), or heart movements (Badea et al., 2005).

**3. Current applications of micro-CT in biomechanics and medicine** 

interest are visualized, thresholded, traced and analyzed.

(30 to 1 µm) (Zagorchev et al., 2010).

**Figure 2.** For optimal settings of the micro-CT scan, the spot size (SS) and the awaited pixel size (PS) are the parameters that set the source distance (SD) and detector distance (DD) (redrawn and modified according to Roth et al., 2010).

The source SS is strongly dependent on the power of the X-ray tube used. In today's machines, the source SS ranges between 1-10 µm. For high resolution scanning of biological samples, it is advisable to operate with a low accelerating voltage (the typical range for the Xradia XCT 400 (Xradia, Pleasanton, CA, USA) is 20–60 kV used with a power of 4 W, at which the tube yields the lowest SS). The scanning time depends on the magnification and resolution required. For details smaller than 1 µm, 24 hours or more might be necessary, whereas an overview scan with a pixel size of approximately 10 µm can be achieved within an hour. Sample drifting might be an issue during a long scanning time. From the reconstructed images, the objects of interest are visualized, thresholded, traced and analyzed.
