The aim of this study is to determine the variation in Hounsfield values with single and multi-slice methods using in-house software on fan-beam computed tomography (FCT), linear accelerator (linac) cone-beam computed tomography (CBCT), and Icon-CBCT datasets acquired using Gammex and advanced electron density (AED) phantoms. The AED phantom was scanned on a Toshiba computed tomography (CT) scanner, five linac-based CBCT X-ray volumetric imaging systems, and Leksell Gamma Knife Icon. The variation between single and multi-slice methods was assessed by comparing scans acquired using Gammex and AED phantoms. The variation in Hounsfield units (HUs) between seven different clinical protocols was assessed using the AED phantom. A CIRS Model 605 Radiosurgery Head Phantom (TED) phantom was scanned on all three imaging systems to assess the target dosimetric changes due to HU variation. An in-house software was developed in MATLAB to assess the HU statistics and the trend along the longitudinal axis. The FCT dataset showed a minimal variation (central slice ± 3 HU) in HU values along the long axis. A similar trend was also observed between the studied clinical protocols acquired on FCT. Variation among multiple linac CBCTs was insignificant. In the case of the water insert, a maximum HU variation of -7.23 ± 68.67 was observed for Linac 1 towards the inferior end of the phantom. All five linacs appeared to have a similar trend in terms of HU variation from the proximal to the distal end of the phantom, with a few outliers for Linac 5. Among three imaging modalities, the maximum variation was observed in gamma knife CBCTs, whereas FCT showed no appreciable deviation from the central value. In terms of dosimetric comparison, the mean dose in CT and Linac CBCT scans differed by <0.5 Gy, whereas at least a 1 Gy difference was observed between CT and gamma knife CBCT. This study shows a minimal variation with FCT between single, volume-based, and multislice methods, and hence the current approach of determining the CT-electron density curve based on a single-slice method would be sufficient for producing a HU calibrations curve for treatment planning. However, CBCTs acquired on linac, and in particular, gamma knife systems, show noticeable variations along the long axis, which is likely to affect the dose calculations performed on CBCTs. It is highly recommended to assess the Hounsfield values on multiple slices before using the HU curve for dose calculations.