KEYWORDS: Bacteria, Luminescence, Diagnostics, Single photon emission computed tomography, In vivo imaging, In vitro testing, Quantitative analysis, Bladder, Tissues, Analytical research
There is a need for the development of diagnostic and analytical models in experimental infection models. We performed in vivo cell-tracking of S. aureus functionalized in vitro with a hybrid antimicrobial peptide tracer 99mTc-UBI29−41-Cy5, containing both a fluorescent and radioactive moiety. To create an invasive infection in mice, viable 99mTc-UBI29−41-Cy5 functionalized bacteria were inoculated in a thigh muscle. Thereafter, the mice were imaged using SPECT and fluorescence imaging modalities at various intervals for a 28h time period. In addition, biodistribution studies were performed at all intervals for quantitative analysis of the colonization and dissemination of the bacteria.
SPECT and fluorescence imaging in mice revealed clear uptake of the tracer in the thigh muscle localization, decreasing over time from 52%ID/g at 4h to 44%ID/g (15% decrease) at 28h p.i. There was little uptake of the tracer in the urinary bladder only at 2-4h p.i.;. Since viable bacteria S. aureus were cultured in the urine samples obtained from the infected mice at all time-points it seems that this reduction is the result of bacterial dissemination. For the other tissues, no substantial accumulation of radioactivity or fluorescence was noticed.
This non-GMO approach of imaging bacteria allowed us to accurately map the distribution of the labeled bacteria in a non-invasive manner. Given the versatility of the approach we are confident that this will pave the way for the development of diagnostic and analytical options in fundamental and translational research on experimental infection models.
Fluorescence guidance facilitates real-time intraoperative visualization of the tissue of interest. However, due to attenuation, the application of fluorescence guidance is restricted to superficial lesions. To overcome this shortcoming, we have previously applied three-dimensional surgical navigation to position the fluorescence camera in reach of the superficial fluorescent signal. Unfortunately, in open surgery, the near-infrared (NIR) optical tracking system (OTS) used for navigation also induced an interference during NIR fluorescence imaging. In an attempt to support future implementation of navigated fluorescence cameras, different aspects of this interference were characterized and solutions were sought after. Two commercial fluorescence cameras for open surgery were studied in (surgical) phantom and human tissue setups using two different NIR OTSs and one OTS simulating light-emitting diode setup. Following the outcome of these measurements, OTS settings were optimized. Measurements indicated the OTS interference was caused by: (1) spectral overlap between the OTS light and camera, (2) OTS light intensity, (3) OTS duty cycle, (4) OTS frequency, (5) fluorescence camera frequency, and (6) fluorescence camera sensitivity. By optimizing points 2 to 4, navigation of fluorescence cameras during open surgery could be facilitated. Optimization of the OTS and camera compatibility can be used to support navigated fluorescence guidance concepts.
Robot-assisted laparoscopic surgery is becoming an established technique for prostatectomy and is increasingly being explored for other types of cancer. Linking intraoperative imaging techniques, such as fluorescence guidance, with the three-dimensional insights provided by preoperative imaging remains a challenge. Navigation technologies may provide a solution, especially when directly linked to both the robotic setup and the fluorescence laparoscope. We evaluated the feasibility of such a setup. Preoperative single-photon emission computed tomography/X-ray computed tomography (SPECT/CT) or intraoperative freehand SPECT (fhSPECT) scans were used to navigate an optically tracked robot-integrated fluorescence laparoscope via an augmented reality overlay in the laparoscopic video feed. The navigation accuracy was evaluated in soft tissue phantoms, followed by studies in a human-like torso phantom. Navigation accuracies found for SPECT/CT-based navigation were 2.25 mm (coronal) and 2.08 mm (sagittal). For fhSPECT-based navigation, these were 1.92 mm (coronal) and 2.83 mm (sagittal). All errors remained below the <1-cm detection limit for fluorescence imaging, allowing refinement of the navigation process using fluorescence findings. The phantom experiments performed suggest that SPECT-based navigation of the robot-integrated fluorescence laparoscope is feasible and may aid fluorescence-guided surgery procedures.
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