Quantification of Cy-5 siRNA signal in the intra-vital multi-photon microscopy images.
Transgenic mice with Tie2- green fluorescent protein (GFP) are used as a model to study the kinetic distribution of the Cy5-siRNA delivered by lipid nanoparticles (LNP) into the liver. After the mouse is injected with the LNP, it undergoes a procedure of intra-vital multi-photon microscopy imaging over a period of two hours, during which the process for the nanoparticle to diffuse into the hepatocytes from the vasculature system is monitored. Since the images are obtained in-vivo, the quantification of Cy5 kinetics suffers from the moving field of view (FOV). A method is proposed to register the sequence of images through template matching. Based on the semi-automatic segmentations of the vessels in the common FOV, the registered images are segmented into three regions of interest (ROI) in which the Cy5 signals are quantified. Computation of the percentage signal strength in the ROIs over time allows for the analysis of the diffusion of Cy5-siRNA into the hepatocytes, and helps demonstrate the effectiveness of the Cy5-siRNA delivery vehicle.
Informatics IT, Merck Research Labs IT, Rahway, NJ 07065, USA.
Antong
A
Chen
Belma
B
Dogdas
Saurin
S
Mehta
Kathleen
K
Haskell
Bruce
B
Ng
Ed
E
Keough
Bonnie
B
Howell
D Adam
DA
Meacham
Amy G
AG
Aslamkhan
Joseph
J
Davide
Matthew
M
Stanton
Ansuman
A
Bagchi
Laura
L
Sepp-Lorenzino
Weikang
W
Tao
D000818:Animals; D002232:Carbocyanines / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D021621:Imaging, Three-Dimensional; D051379:Mice; D008822:Mice, Transgenic; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D034741:RNA, Small Interfering / Q000378:metabolism*; D012815:Signal Processing, Computer-Assisted
D002232:Carbocyanines; D034741:RNA, Small Interfering; C085321:cyanine dye 5; D049452:Green Fluorescent Proteins