Designed for specific uses (e.g., gradients or gel matrices)
All-in-one chambers reduce the time and number of experimental steps in cell-based assays. The ibidi μ-Slides, μ-Dishes and μ-Plates are designed as all-in-one chambers, so that all of the experimental steps, from cell cultivation to microscopic imaging, can be done in one chamber.
The open-well format of the µ-Slide 2 Well | 4 Well | 8 Well and the μ-Dish 35 mm, high allows for the use of standard immunofluorescence protocols. After fixation and staining, the sample can be observed through the coverslip bottom using high resolution microscopy. There is no need for an additional glass coverslip.
Channel formats like the μ-Slide VI 0.4 are ideal for an exact exchange of small amounts of reagents. These μ-Slides reduce the number of experimental steps needed in immunofluorescence assays. The channel format supports homogeneity of cell distribution and phase contrast microscopy.
Order a free sample to test ibidi’s µ-Slides, µ-Dishes, and µ-Plates with your experiments.
ibidi µ-Slides, µ-Dishes, and µ-Plates are Designed as All-in-One Chambers.
Inverted microscopes are popular for live cell imaging, because:
Cells sink to the bottom and onto the coverslip for adherence
Sample access from the top, e.g., for liquid exchange or micropipettes
No contact between objective and sample: sterile working conditions are possible
Homogeneous Cell Distribution, as a Result of Geometry
Cell densities in open wells are very dependent on handling during cell seeding. Unlike in the open wells, cell densities in channels neither vary with the position inside the slide, nor with the handling and treatment during and after cell seeding.
To demonstrate the influence of slide geometry in cell distribution, cells were cultivated in various formats. Cell layers were visualized macroscopically, using crystal violet staining, and additionally by using fluorescence and phase contrast microscopy.
The macroscopic photographs (shown here) illustrate that cells cultured in open wells, such as the µ-Slide 8 Well, formed characteristic patterns. One common pattern that was found was that some of the cells congregated to the edges of the well. Normally, fewer cells attach to that area, while in the middle of the well the cell density will reach its maximum. In contrast, when cultured in a channel format (such as the ibidi μ-Slide VI 0.4) the cell distribution was always homogeneous. These macroscopically derived results were confirmed by phase contrast and fluorescence microscopy.
Inhomogeneous cell distribution in open wells
Homogeneous cell distribution in cell culture channels
Phase Contrast in Channel Slides is Superior to Small Open Wells
Unstained samples, such as bacteria or living cell cultures, absorb practically no light. This makes them barely, or not at all, visible in brightfield, even in a well-aligned microscope. Poor light absorption results in extremely small differences in the intensity distribution in the image. In phase contrast, small phase delays that come from slightly different refractive indices in different materials are converted into intensity changes over the entire image.
A pre-condition for good phase contrast, is to use the correct Köhler illumination without any distortion or unwanted diffraction.
The meniscus formation in open well chambers at the air-water-interphase is a normal and unavoidable effect disturbing phase contrast microscopy.
96 well plate or small open well: Strong influence of meniscus, low contrast near the edges
Channel or parallel plates: No meniscus, good phase contrast over the entire area
Phase contrast microscopy in channel slides (e.g., the ibidi µ-Slide VI 0.4) is much better than in small open wells, especially near the edges. The diffraction, due to the meniscus, disarranges the correct alignment of the phase ring and phase plate inside the optical pathway.
Condensation on the lid of the culture vessel is another annoying disturbing effect.
In the ibidi channel slides, condensation inside the optical pathway is intrinsically impossible. The example below shows this result after the sample is removed from the incubator.
The Ph+ Feature: Excellent Meniscus-Free Phase Contrast Microscopy
The Ph+ Feature Diminishes the Meniscus and Increases the Area of Well-Contrasted Cells
The illustration on the right shows the perturbing effect of a meniscus. Light is refracted on the air-water-interface, leading to poor contrast in microscopy. Only the small center part exhibits satisfying phase contrast.
Working with the Ph+ Well diminishes the meniscus and increases the area of nicely contrasted cells. This nice contrast is due to the parallel beam path that is created by the plate.
Standard Well Versus Ph+ Well: No Meniscus Effect in Ph+ Well