scholarly journals A viability study of 3D tumor spheroids after their mass-density characterization via an innovative flow-based biophysical method

2021 ◽  
Vol 4 (s1) ◽  
Author(s):  
Francesco Bacchi ◽  
Azzurra Sargenti ◽  
Francesco Musmeci ◽  
Simone Pasqua ◽  
Simone Bonetti ◽  
...  
Keyword(s):  
Mass Density ◽  
Cell Models ◽  
Tumor Spheroids ◽  
Viability Study ◽  
Simple Measurement ◽  
Biophysical Method

The simple measurement of mass density, size and weight of sphere-like 3D cell models has been recently enabled by a specifically conceived flow-based method. Here we demonstrate that such technique also allows the post-analysis collection of live 3D tumor spheroids, without compromising their viability.

scholarly journals A Reliable Flow-Based Method for the Accurate Measure of Mass Density, Size and Weight of Live 3D Tumor Spheroids

Micromachines ◽  
2020 ◽  
Vol 11 (5) ◽  
pp. 465 ◽  
Author(s):  
Domenico Andrea Cristaldi ◽  
Azzurra Sargenti ◽  
Simone Bonetti ◽  
Francesco Musmeci ◽  
Cecilia Delprete ◽  
...  
Keyword(s):  
Single Cell Analysis ◽  
Mass Density ◽  
3D Culture ◽  
Physical Method ◽  
3D Models ◽  
Terminal Velocity ◽  
Tumor Spheroids ◽  
Culture Models ◽  
Technical Solutions ◽  
Free Falling

Gathering precise information on mass density, size and weight of cells or cell aggregates, is crucial for applications in many biomedical fields with a specific focus on cancer research. Although few technical solutions have been presented for single-cell analysis, literature does not cover this aspect for 3D models such as spheroids. Since the research interest on such samples is notably rising, here we describe a flow-apparatus, and the associated physical method and operative protocol for the accurate measurements of mass density, size and weight. The technique is based on the detection of the terminal velocity of a free-falling sample into a specifically conceived analysis flow-channel. Moreover, in order to demonstrate the accuracy and precision of the presented flow-device, analyses were initially carried out on standardized polystyrene beads. Finally, to display the application of the proposed system for biological samples, mass density, size and weight of live SW620 tumor spheroids were analyzed. The combined measurements of such parameters can represent a step toward a deeper understanding of 3D culture models.

Mass Determination by Direct Measurement of Electron Scattering

1975 ◽  
Vol 33 ◽  
pp. 240-241
Author(s):  
M. K. Lamvik ◽  
A. V. Crewe
Keyword(s):  
Cross Section ◽  
Electron Scattering ◽  
Mass Density ◽  
Variable Mass ◽  
Scattering Cross Section ◽  
Mass Determination ◽  
Number Density ◽  
Cross Sectional ◽  
Thin Slice ◽  
Scattering Cross

If a molecule or atom of material has molecular weight A, the number density of such units is given by n=Nρ/A, where N is Avogadro's number and ρ is the mass density of the material. The amount of scattering from each unit can be written by assigning an imaginary cross-sectional area σ to each unit. If the current I0 is incident on a thin slice of material of thickness z and the current I remains unscattered, then the scattering cross-section σ is defined by I=IOnσz. For a specimen that is not thin, the definition must be applied to each imaginary thin slice and the result I/I0 =exp(-nσz) is obtained by integrating over the whole thickness. It is useful to separate the variable mass-thickness w=ρz from the other factors to yield I/I0 =exp(-sw), where s=Nσ/A is the scattering cross-section per unit mass.

Image formation analysis of thick biological specimens using three-dimensional power-spectra of through focus series

1994 ◽  
Vol 52 ◽  
pp. 124-125
Author(s):  
Karen F. Han
Keyword(s):  
Inelastic Scattering ◽  
Imaging Techniques ◽  
Mass Density ◽  
Relative Proportion ◽  
Three Dimensional ◽  
Power Spectra ◽  
Image Formation ◽  
Energy Filtering ◽  
Ewald Sphere ◽  
Nonlinear Imaging

The primary focus in our laboratory is the study of higher order chromatin structure using three dimensional electron microscope tomography. Three dimensional tomography involves the deconstruction of an object by combining multiple projection views of the object at different tilt angles, image intensities are not always accurate representations of the projected object mass density, due to the effects of electron-specimen interactions and microscope lens aberrations. Therefore, an understanding of the mechanism of image formation is important for interpreting the images. The image formation for thick biological specimens has been analyzed by using both energy filtering and Ewald sphere constructions. Surprisingly, there is a significant amount of coherent transfer for our thick specimens. The relative amount of coherent transfer is correlated with the relative proportion of elastically scattered electrons using electron energy loss spectoscopy and imaging techniques.Electron-specimen interactions include single and multiple, elastic and inelastic scattering. Multiple and inelastic scattering events give rise to nonlinear imaging effects which complicates the interpretation of collected images.

Novel epoxy/anhydride alternatives for biological electron microscopy: Physical and performance characteristis of embed 812 and LX 112 in combination with NSA/NMA/DMAE

1989 ◽  
Vol 47 ◽  
pp. 1000-1001
Author(s):  
Joe A. Mascorro ◽  
Gerald S. Kirby
Keyword(s):  
Electron Microscopy ◽  
Flow Rate ◽  
Succinic Anhydride ◽  
Volume Flow Rate ◽  
Mass Density ◽  
Uranyl Acetate ◽  
Volume Flow ◽  
Base Resin ◽  
And Performance ◽  
Acid Anhydrides

Embedding media based upon an epoxy resin of choice and the acid anhydrides dodecenyl succinic anhydride (DDSA), nadic methyl anhydride (NMA), and catalyzed by the tertiary amine 2,4,6-Tri(dimethylaminomethyl) phenol (DMP-30) are widely used in biological electron microscopy. These media possess a viscosity character that can impair tissue infiltration, particularly if original Epon 812 is utilized as the base resin. Other resins that are considerably less viscous than Epon 812 now are available as replacements. Likewise, nonenyl succinic anhydride (NSA) and dimethylaminoethanol (DMAE) are more fluid than their counterparts DDSA and DMP- 30 commonly used in earlier formulations. This work utilizes novel epoxy and anhydride combinations in order to produce embedding media with desirable flow rate and viscosity parameters that, in turn, would allow the medium to optimally infiltrate tissues. Specifically, embeding media based on EmBed 812 or LX 112 with NSA (in place of DDSA) and DMAE (replacing DMP-30), with NMA remaining constant, are formulated and offered as alternatives for routine biological work.Individual epoxy resins (Table I) or complete embedding media (Tables II-III) were tested for flow rate and viscosity. The novel media were further examined for their ability to infilftrate tissues, polymerize, sectioning and staining character, as well as strength and stability to the electron beam and column vacuum. For physical comparisons, a volume (9 ml) of either resin or media was aspirated into a capillary viscocimeter oriented vertically. The material was then allowed to flow out freely under the influence of gravity and the flow time necessary for the volume to exit was recored (Col B,C; Tables). In addition, the volume flow rate (ml flowing/second; Col D, Tables) was measured. Viscosity (n) could then be determined by using the Hagen-Poiseville relation for laminar flow, n = c.p/Q, where c = a geometric constant from an instrument calibration with water, p = mass density, and Q = volume flow rate. Mass weight and density of the materials were determined as well (Col F,G; Tables). Infiltration schedules utilized were short (1/2 hr 1:1, 3 hrs full resin), intermediate (1/2 hr 1:1, 6 hrs full resin) , or long (1/2 hr 1:1, 6 hrs full resin) in total time. Polymerization schedules ranging from 15 hrs (overnight) through 24, 36, or 48 hrs were tested. Sections demonstrating gold interference colors were collected on unsupported 200- 300 mesh grids and stained sequentially with uranyl acetate and lead citrate.

Structural Studies on the Eukaryotic Ribosome

1990 ◽  
Vol 48 (1) ◽  
pp. 256-257
Author(s):  
Adriana Verschoor ◽  
Ronald Milligan ◽  
Suman Srivastava ◽  
Joachim Frank
Keyword(s):  
Chick Embryo ◽  
3D Reconstruction ◽  
Single Particle ◽  
Mass Density ◽  
Particle Surface ◽  
Uranyl Acetate ◽  
Electron Microscopic ◽  
Eukaryotic Ribosome ◽  
Negative Stain ◽  
Reconstruction Methods

We have studied the eukaryotic ribosome from two vertebrate species (rabbit reticulocyte and chick embryo ribosomes) in several different electron microscopic preparations (Fig. 1a-d), and we have applied image processing methods to two of the types of images. Reticulocyte ribosomes were examined in both negative stain (0.5% uranyl acetate, in a double-carbon preparation) and frozen hydrated preparation as single-particle specimens. In addition, chick embryo ribosomes in tetrameric and crystalline assemblies in frozen hydrated preparation have been examined. 2D averaging, multivariate statistical analysis, and classification methods have been applied to the negatively stained single-particle micrographs and the frozen hydrated tetramer micrographs to obtain statistically well defined projection images of the ribosome (Fig. 2a,c). 3D reconstruction methods, the random conical reconstruction scheme and weighted back projection, were applied to the negative-stain data, and several closely related reconstructions were obtained. The principal 3D reconstruction (Fig. 2b), which has a resolution of 3.7 nm according to the differential phase residual criterion, can be compared to the images of individual ribosomes in a 2D tetramer average (Fig. 2c) at a similar resolution, and a good agreement of the general morphology and of many of the characteristic features is seen.Both data sets show the ribosome in roughly the same ’view’ or orientation, with respect to the adsorptive surface in the electron microscopic preparation, as judged by the agreement in both the projected form and the distribution of characteristic density features. The negative-stain reconstruction reveals details of the ribosome morphology; the 2D frozen-hydrated average provides projection information on the native mass-density distribution within the structure. The 40S subunit appears to have an elongate core of higher density, while the 60S subunit shows a more complex pattern of dense features, comprising a rather globular core, locally extending close to the particle surface.

Intronic miRNAs of Apoptosis Genes as Indicators of Cell Death

2019 ◽  
Vol 35 (6) ◽  
pp. 108-113
Author(s):  
J.A. Makarova ◽  
A.A. Poloznikov
Keyword(s):  
Breast Cancer ◽  
Cancer Cell Line ◽  
Ministry Of Education ◽  
Apoptosis Induction ◽  
Cell Models ◽  
Process Stage ◽  
Mature Micrornas ◽  
The Russian Federation ◽  
Apoptosis Level ◽  
Mcf 7

A method to assess the apoptosis level in cell models based on the analysis of the expression of micRNAs located in introns of apoptosis genes has been developed. Bioinformation analysis identified 536 genes associated with apoptosis; 30 of them contained 38 pre-microRNAs encoding 41 mature microRNAs. A significant change in the expression of hsa-miR-1244 and hsa-miR-4479 in response to apoptosis induction in the MCF-7 breast cancer cell line was revealed. A correlation was also found between the expression level of these miRNAs and the size of the primary tumor (process stage) in patients with breast cancer. apoptosis, microRNA, MCF7, breast cancer This work was supported by the Ministry of Education and Science of the Russian Federation (Project no. RFMEFI61618X0092).

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