NIRFC imaging of a biliary excreted NIR probe facilitates intra-operative visualization of the bile ducts in real-time and high definition and allows for rapid identification of bile duct anatomy and pathology. Clinically, identification of free bile in the abdominal cavity confirms bile duct transection or perforation. However, during conventional laparoscopic cholecystectomy it is difficult to distinguish a small amount of extra-luminal bile from a mixture of blood and transudations in the surgical field [25
]. Moreover, unintentional small bile duct ligature may occur and is usually not detected immediately. These two situations are responsible for the majority of intra-operative bile duct injuries [27
X-ray cholangiography is a standard clinical imaging technique used during cholecystectomy surgeries, the benefits of which have been well researched and demonstrated [28
]. Its routine intra-laparoscopic use is still debated by some and the decision to perform IOCG depends on each surgeon’s experience and local facilities [29
]. Most commonly, intra-laparoscopic cholangiography is used when bile duct injuries are suspected.
In the clinical setting, a video laparoscopic imaging system could potentially provide multiple channels integrating white light and fluorescent light sources and several such systems have already been described [32
]. Images could be fused in real-time and thus provide the surgeon with additional information regarding anatomy and bile leaks. Theoretically, fluorescent intra-operative cholangiography could be done easily by injecting the fluorescent contrast agent and switching the camera from color to infrared lighting. However, intra-operative fluorescent imaging using shorter wavelength light [17
] has shown lower achievable TBRs due to absorbance of endogenous fluorochromes such as hemoglobin, melanin, and water.
Imaging hepatic excretion of indocyanine green (ICG) could be used as an alternative method to increase fluorescence [19
]. However, because ICG highlights both vasculature and biliary anatomy, it could result in difficulties distinguishing them intra-laparoscopically. Moreover, ICG is also dependent on cardiac and hepatic function. Thus, the use of the presented lipophilic indocyanine-based NIR fluorescent agent designed for hepatobiliary excretion imaging without fluorescence during the vascular time allows for clear differentiation of vessels from bile ducts.
The fluorescent agent used in the current experiment has absorption/emission peaks at 675/692 nm, but more importantly, it has ideal pharmacokinetic properties, which enhance extra-hepatic bile duct imaging with little background from the portal hepatis. An additional problem, common with fluorescent imaging, is the low penetration depth of light through tissue matter. However, considering that the thickness of a normal human bile duct wall and wall thickness in a biliary obstruction situation is 0.6 ± 0.3 mm and 0.8 ± 0.5 mm, respectively [26
], the low penetration depth would be unlikely to pose a problem and our strategy would therefore be suitable for laparoscopic cholecystectomy [35
]. The penetration of NIR light is dependent on tissue absorption, scatterings, auto fluorescence and wavelengths/illumination [36
]. The higher the wavelength, the better the signal-to-noise ratio. In a reflectance mode such as the one used here, maximum penetration depths are usually ~ 5–8 mm, before scattering degrades image quality [37
]. In tomographic mode however, penetration depths increase considerably to several centimeters [38
]. It is thus not inconceivable to devise hand held tomographic scanner units similar to intra-operative ultra-sounds probes [39
In high definition and real-time, we observed the NIR probe moving from the hepatic parenchyma towards the biliary ducts, as early as three minutes after intravenous administration. From 5 to 60 minutes post-injection, a bright fluorescent signal was seen inside the biliary ducts with low hepatic background signal. Intra-operative imaging facilitated identification of the dilation/stricture area in a bile duct stenosis model. Similarly, perforation of the common bile duct was easily located at the site where the bile and the probe leaked into the abdominal cavity. Synthetic beads were used to model bile stones and were readily identified within the lumen of the common bile duct, suggesting easy intra-operative identification and localization of choledocholithiasis.
Since the probe reaches the biliary system within minutes, a patient could be injected with the agent following anesthesia induction or during placement of the laparoscopic instruments. This would allow the probe to reach far within the extra-hepatic biliary system at the beginning of the surgery exploration. NIRFC could be used as a routine procedure, as has been suggested for intra-operative X-ray cholangiography. NIRFC could also be possibly used in patients falling into high-risk categories for development of complications or in those undergoing a second operation to repair suspected iatrogenic injuries. Alternatively, since the signal is bright at five minutes post-injection, the probe could be injected intra-operatively at any point or re-injected multiple times to assess for suspected biliary injury. In addition, post-operative patients necessitating drain placement could be injected with the fluorescent agent and have drain contents examined at the bedside with a portable NIR source.