Novel 3D Liquid Cell Culture Method for Anchorage-independent Cell Growth, Cell Imaging and Automated Drug Screening

Abstract Cells grown in three-dimensional (3D) cultures are more likely to have native cell-cell and cell-matrix interactions than in 2D cultures that impose mechanical constraints to cells. However, most 3D cultures utilise gel matrix which, while serving as a scaffold, limits application due to its solid and opaque nature and inconsistency in cell exposure to exogenous signals. In 3D culture without gel matrix, cells tend to adhere to each other and form clumps with necrotic zone at the centre, making them unsuitable for analyses. Here we report that addition of low-molecular-weight agar named LA717 to culture media allows cells to grow as dispersed clonal spheroids in 3D. LA717 maintains cells dispersed and settled to the bottom of the medium while keeping the medium clear with little additional viscosity, making it suitable for microscopic observation. Importantly, cancer spheroids formed in LA717-containing medium show higher sensitivity to anti-cancer drugs such as Trametinib and MK-2206 that are not as effective in 2D. Because of the small and consistent size of spheroids, cell viability and drug toxicity are readily detectable in automated imaging analysis. These results demonstrate that LA717 offers a novel 3D culture system with great in vivo reflection and practicality

Mineral oil certified reference materials for the determination of polychlorinated biphenyls from the National Metrology Institute of Japan (NMIJ)

Four mineral oil certified reference materials (CRMs), NMIJ CRM 7902-a, CRM 7903-a, CRM 7904-a, and CRM 7905-a, have been issued by the National Metrology Institute of Japan, which is part of the National Institute of Advanced Industrial Science and Technology (NMIJ/AIST), for the determination of polychlorinated biphenyls (PCBs). The raw materials for the CRMs were an insulation oil (CRM 7902-a and CRM 7903-a) and a fuel oil (CRM7904-a and CRM 7905-a). A solution of PCB3, PCB8, and technical PCB products, comprising four types of Kaneclor, was added to the oil matrices. The total PCB concentrations in the PCB-fortified oils (CRM 7902-a and CRM 7904-a) are approximately 6 mg kg−1. In addition, the mineral oils which were not fortified with PCBs were also distributed as CRMs (CRM 7903-a and CRM 7905-a). Characterization of these CRMs was conducted by the NMIJ/AIST, where the mineral oils and the PCB solution were analyzed using multiple analytical methods such as dimethylsulfoxide extraction, normal-phase liquid chromatography, gel permeation chromatography, reversed-phase liquid chromatography, and chromatography using sulfoxide-bonded silica; and/or various capillary columns for gas chromatography, and two ionization modes for mass spectrometry. The target compounds in the mineral oils and those in the PCB solution were determined by one of the primary methods of measurement, isotope dilution–mass spectrometry (ID-MS). Certified values have been provided for 11 PCB congeners (PCB3, 8, 28, 52, 101, 118, 138, 153, 180, 194, and 206) in the CRMs. These CRMs have information values for PCB homologue concentrations determined by using a Japanese official method for determination of PCBs in wastes and densities determined with an oscillational density meter. Because oil samples having arbitrary PCB concentrations between respective property values of the PCB-fortified and nonfortified CRMs can be prepared by gravimetric mixing of the CRM pairs, these CRMs can be used for validation of PCB analyses using various instruments which have different sensitivities

Additional file 2: Figure S1. of The prognostic value of serum CA 19-9 for patients with advanced lung adenocarcinoma

Scatter-plot of serum CA 19-9 and CYFRA 21-1 levels. (JPG 394 kb