There are a number of apoptosis analysis methods that use imaging. When focusing on nuclei, such phenomena as chromatin aggregation, nuclei condensation and DNA cutting,etc. are observed along with the progress of apoptosis, which are recognizable from microscopic images. The following is an example of the analysis of nuclear transformation by using the images acquired by CV7000 and 「Nuclear Morphology」function of the analysis software.
Images were acquired using multiple object lenses and analyzed by the analysis software. As a result, changes in the nuclear areas due to apoptosis were observed with any of the objective lenses(Fig. 1). In the created doseresponse curves of three factors (Fig. 2), both the 4X and 40X lenses gave almost the same results. In addition, bar charts of individual cell distribution in the wells with staurosporine concentrations of 0μM and 10μM (Fig. 3) and pie charts (Fig. 4) were created using Spotfire®, both of which clearly indicated apoptosis. In this way, the 「Nuclear Morphology」function of the analysis software enables multilateral analysis of changes in nuclear morphology, and stable analysis of images acquired by as low as 4X magnification object lenses, too.
Fig. 1: Images captured by CV7000, and their recognition results
Based on the nuclear area per cell, live cells and cells with apoptosis are recognized and colored in green and red, respectively.
It is clearly shown that almost all nuclei are fragmented due to apoptosis in the wells with 10μM staurosporine.
- HeLa cells were cultured in 96-well plates with 10,000 cells/well for 24 hours.
- Add Staurosporine, fix with formaldehyde four hours later, and stain nuclei with Hoechst33342.
- CV7000 observation conditions are shown below: (wavelength: 405nm)
Magnification Exposure time (405nm) Number of images (per well) Number of recognized cells (well average) 4x 500 msec 1 3,452 cells 10x 250 msec 4 3,028 cells 20x 250 msec 9 1,243 cells 40x 250 msec 16 493 cells
Analyze captured images using the 「Nuclear Morphology」function of the analysis software in its advanced mode.
• Extract nuclear areas from the images to recognize the nuclei with less than 100μm2 area as micronuclei fragmented due to apoptosis.
• Calculate the area and average fluorescence intensity of the nuclei and fragmented micronuclei, respectively, as well as the number of micronuclei.
Fig. 2: Staurosporine dose-response curves: Imaged with (a)4x or (b)40x lens
Fig.3 Cell number distribution graphs (40X)
Fig. 4 (a): Pie-chart : Ratio of cells at various staurosporine concentrations. (Magnification: 40x)
The ratio of cells with nuclear fluorescence intensity of 800 or more are colored in green.
Fig. 4 (b): Pie-chart: Ratios of cells at various staurosporine concentrations. (Magnification: 40x)
The ration of cells with one or more fragmented nuclear granules are colored in green.
Our Social Medias
We post our information to the following SNSs. Please follow us.
|Follow us||Share our application|
|@Yokogawa_LS||Share on Twitter|
|Yokogawa Life Science||Share on Facebook|
|Yokogawa Life Science||Share on LinkedIn|
Yokogawa's Official Social Media Account List
ilgili Ürünler & Çözümler
Benchtop CQ1 Confocal System
The CellVoyager CQ1 provides the highest quality confocal images and extended live-cell imaging in a space-saving benchtop design.
CellPathfinder is designed for our HCA systems, CQ1 and the CellVoyager series. From beginners to experts, the analysis software lets you quantify subtle physiological changes and even label-free samples with various graph options.
CV8000 High-Throughput System
CellVoyager CV8000 is the most advanced high-content screening system. The improved built-in incubator lets you analyze extended live cell responses. With its expandability, 4 cameras, 5 lasers and an optional built-in pipettor, the system permits increasingly complex assay development and high-content screening.
High Content Analysis CellVoyager
Our high-content analysis (HCA) systems utilize powerful software to address a wide range of research applications from basic science to complex compound screening.
Yokogawa’s high content analysis systems and dual spinning disk confocal technologies provide high-speed and high-resolution live cell imaging, enabling leading-edge research around the world.