Dataset Information

3D printed biaxial stretcher compatible with live fluorescence microscopy.

ABSTRACT: Mechanical characterization and tensile testing of biological samples is important when determining the material properties of a tissue; however, performing tensile testing and tissue stretching while monitoring cellular changes via fluorescence microscopy is often challenging. Additionally, commercially available cell/tissue stretchers are often expensive, hard to customize, and limited in their fluorescence imaging compatibility. We have developed a 3D printed Open source Biaxial Stretcher (OBS) to be a low-cost stage top mountable biaxial stretching system for use with live cell fluorescence microscopy in both upright and inverted microscope configurations. Our OBS takes advantage of readily available open source desktop 3D printer hardware and software to deliver a fully motorized high precision (10 ± 0.5 μm movement accuracy) low cost biaxial stretching device capable of 4.5 cm of XY travel with a touch screen control panel, and an integrated heated platform with sample bath to maintain cell and tissue viability. Further, we designed a series of tissue mounts and clamps to accommodate varying samples from synthetic materials to biological tissue. By creating a low-profile design, we can directly mount the stretcher onto a microscope stage, and through coordinated biaxial stretching we maintain a constant field of view facilitating real-time sample tracking and time-lapse fluorescence imaging.

SUBMITTER: Shiwarski DJ

PROVIDER: S-EPMC8794355 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:One therapeutical alternative in the treatment of functional disorders is the use of printed oral splints. The mechanical properties of these materials are highly essential to their clinical effectiveness, and their performance may vary depending on factors such as cleaning, post-polymerization, or their orientation during construction. The objective of this in vitro investigation is to evaluate the effectiveness of the selected materials in terms of their biaxial flexural strength in relation to the criteria listed above. Splint materials were used in the printing of 720 discs. The printing process was carried out in different orientations in relation to the building platform. Either an automatic or manual cleaning process was performed on the samples. For post-polymerization, either an LED or Xenon light was utilized. A piston-on-three-ball test was used to measure the biaxial flexural strength (BFS) of the materials after they were stored in water for either 24 h or 60 days. The homogeneity of the data was controlled by employing the Levene method, and the differences between the groups were analyzed using the ANOVA and Bonferroni methods. After being stored for twenty-four hours, the mean BFS ranged anywhere from 79 MPa to 157 MPa. Following a period of sixty hours, the BFS exhibited a substantial drop and revealed values that ranged from 72 to 127 MPa. There was no significant difference that could be identified between the materials or between the various cleaning processes. The results of post-polymerization showed that the LED light produced higher means than the Xenon light did. In terms of position, the mean values varied greatly, with 0°'s mean value being 101 MPa, 45°'s mean value being 102 MPa, and 90°'s mean value being 115 MPa. The use of a build orientation of 90° and post-polymerization with LED light resulted in significantly increased biaxial flexural strength. According to this study, this design should be implemented in order to ensure that splint materials have the highest possible strength.

Project description:Background:Live-cell fluorescence microscopy (LCFM) is a powerful tool used to investigate cellular dynamics in real time. However, the capacity to simultaneously measure DNA content in cells being tracked over time remains challenged by dye-associated toxicities. The ability to measure DNA content in single cells by means of LCFM would allow cellular stage and ploidy to be coupled with a variety of imaging directed analyses. Here we describe a widely applicable nontoxic approach for measuring DNA content in live cells by fluorescence microscopy. This method relies on introducing a live-cell membrane-permeant DNA fluorophore, such as Hoechst 33342, into the culture medium of cells at the end of any live-cell imaging experiment and measuring each cell's integrated nuclear fluorescence to quantify DNA content. Importantly, our method overcomes the toxicity and induction of DNA damage typically caused by live-cell dyes through strategic timing of adding the dye to the cultures; allowing unperturbed cells to be imaged for any interval of time before quantifying their DNA content. We assess the performance of our method empirically and discuss adaptations that can be implemented using this technique. Results:Presented in conjunction with cells expressing a histone 2B-GFP fusion protein (H2B-GFP), we demonstrated how this method enabled chromosomal segregation errors to be tracked in cells as they progressed through cellular division that were later identified as either diploid or polyploid. We also describe and provide an automated Matlab-derived algorithm that measures the integrated nuclear fluorescence in each cell and subsequently plots these measurements into a cell cycle histogram for each frame imaged. The algorithm's accurate assessment of DNA content was validated by parallel flow cytometric studies. Conclusions:This method allows the examination of single-cell dynamics to be correlated with cellular stage and ploidy in a high-throughput fashion. The approach is suitable for any standard epifluorescence microscope equipped with a stable illumination source and either a stage-top incubator or an enclosed live-cell incubation chamber. Collectively, we anticipate that this method will allow high-resolution microscopic analysis of cellular processes involving cell cycle progression, such as checkpoint activation, DNA replication, and cellular division.

Dataset Information

3D printed biaxial stretcher compatible with live fluorescence microscopy.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets