Highly sensitive detection of cancer cells using femtosecond dual-wavelength near-IR two-photon imaging

Abstract

We describe novel imaging protocols that allow detection of small cancer cell colonies deep inside tissue phantoms with high sensitivity and specificity. We compare fluorescence excited in Styryl-9M molecules by femtosecond pulses at near IR wavelengths, where Styryl-9M shows the largest dependence of the two-photon absorption (2PA) cross section on the local environment. We show that by calculating the normalized ratio of the two-photon excited fluorescence (2PEF) intensity at 1200 nm and 1100 nm excitation wavelengths we can achieve high sensitivity and specificity for determining the location of cancer cells surrounded by normal cells. The 2PEF results showed a positive correlation with the levels of MDR1 proteins expressed by the cells, and, for high MDR1 expressors, as few as ten cancer cells could be detected. Similar high sensitivity is also demonstrated for tumor colonies induced in mouse external ears. This technique could be useful in early cancer detection, and, perhaps, also in monitoring dormant cancer deposits.

A machine learning based approach for quantitative evaluation of cell migration in Transwell assays based on deformation characteristics

“…Quantitative phase image techniques, such as spatial light interference microscopy 14 and digital holographic microscopy, 15 are used to acquire images of the cell phase to characterize the refractive index, because the refractive index is the intrinsic property of a cell. Due to the coupling of the cell thickness information with the refractive index in the phase value, some decoupling methods, including the dual wavelength technique and confocal technique, have been used to obtain the refractive index to investigate the morphology changes of cells 16,17 In addition, the synergy between microfluidics and cell refractometers have been developed to acquire the refractive index. 18 However, it is difficult to identify migrated cells, because of the similar refractive index between migrated cells and non-migrated cells.…”

Photoisomerization of Linear and Stacked Isomers of a Charged Styryl Dye: A Tandem Ion Mobility Study

“…Styrylpyridines, the aza-analogues of stilbene, have been thoroughly investigated theoretically − and experimentally due to their potential to propel molecular machines, drive spin changes in transition organometallic complexes, , and act as DNA staining agents . More complex styryls, including styryl 7 and styryl 9M (S9M), which have two or more isomerizable bonds, have been proposed as efficient fluorescent probes for bioimaging, biosensing, and medical diagnosis applications. ,− The attraction of S9M for various biotechnological applications stems partly from its high two-photon absorption cross-section in the near-infrared, enabling its use as a hyperspectral fluorescent probe. − The potential applications for styryl dyes have prompted studies of their photoexcitation, fluorescence and photoisomerization dynamics in solution. − Many styryl dyes exhibit strong solvatochromism and pH-dependent photoabsorption …”

Optimizing ultrafast illumination for multiphoton-excited fluorescence imaging

“…We evaluate the maximum achievable shot noise limited rate of imaging depending on the two-photon absorption (2PA) and three-photon absorption (3PA) cross section values of the fluorophores and other key parameters such as average laser power, sample damage threshold etc. Some applications such as early cancer detection [ 12 ] and high throughput screening [ 13 ] require much larger samples, that cannot be addressed by conventional coaxial illumination setups. For this purpose, we model a wide field-of-view (FOV) imaging system, where the illumination- and the fluorescence detection beam paths are separated, and where the illumination uses a ~kHz pulse rate amplified femtosecond laser.…”