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FAQ: Live Cell Imaging Using the ibidi Heating and Gas Incubation System

Is the ibidi Heating and Gas Incubation System compatible with every type of microscope?

The ibidi Heating and Gas Incubation System is available in several different versions that can be used with a variety of microscopic systems.
The ibidi Heating and Gas Incubation System, Universal Fit, is compatible with any inverted microscope that has a stage or a frame for multi-well plates, such as 96-well plates. The outer dimensions of this ibidi Heating System and a 96-well plate are identical.

The ibidi Heating System, Multi-Well Plates, K-Frame is compatible with microscopic stages with a K-Frame opening (160 x 110 mm). The compatible stages are listed in the table below:

Compatibility to Microscopes

Nikon --- Ti-S-E, Ti-S-ER, TI2-S-SE-E, and TI2-S-SS-E Stages
ASI MS 2000 Series
PZ 2000 Series
---
Leica K-Frame Opening (160x110mm) ---
LUDL BioPrecision 2 Inverted
Biopoint 2 Series
---
Märzhäuser Scan IM Series ---
Olympus K-Frame Opening (160x110mm) ---
Physik Instrumente M-687 PILine ---
PRIOR H107
ES107
---
Thorlabs MLS203-1/MLS203-2 ---
Zeiss Scanning Stage 130 x 85 mot. P
K-Frame Opening (160x110mm)
---


The ibidi Heating System, Multi-Well Plates for Nikon Ti-S-E and Ti-S-ER is compatible with Nikon Ti-S-E, Ti-S-ER, Ti2-S-SE-E, and Ti2-S-SS-E motorized stages.

Is it possible to incubate multi-well plates using the ibidi Heating System?

ibidi has developed two different Heating Systems for multi-well plates. Each of them is compatible with a distinct microscopic stage format. The ibidi Heating System, Multi-Well Plates, K-Frame fits into microscope stages with a K-Frame opening (160 x 110 mm). The ibidi Heating System, Multi-Well Plates for Nikon Ti-S-E and Ti-S-ER is specifically compatible with the Nikon Ti-S-E, Ti-S-ER, Ti2-S-SE-E, and Ti2-S-SS-E motorized stages, which hold multi-well plates. Both systems provide temperature control for samples held in multi-well plates.

If you would like further information on adapting your current Heating System, please contact us.

Does the Heated Lid of the ibidi Heating and Gas Incubation System affect the resolution?

No, the Heated Lid does not affect the resolution. When working with transmitted light there is no reported effect on the optical pathway, because the Heated Lid in the ibidi Heating and Gas Incubation System is made of glass. Thus, the resolution remains unchanged. In theory, there is a small percentage of intensity loss during light transmission, but this is not an issue for transmitted light.

When working with fluorescence microscopy, the optical pathway does not pass the Heated Lid, therefore it does not influence the resolution.

Can the ibidi Heating System be combined with gas incubation?

The ibidi Heating Systems are designed for use in combination with the ibidi Gas Incubation System. For this purpose, there are two openings in the Heated Lid: one for the gas inlet and the other for the humidity sensor. This allows not only temperature, but also the CO2 concentration and the humidity to be controlled. All ibidi temperature chambers are upgradeable for gas incubation.

Is oil-immersion microscopy possible with the ibidi Heating System, Universal Fit?

Yes, oil immersion microscopy is possible with the ibidi Heating System, Universal Fit due to the bottom opening—there is no obstacle between the sample and the objective lens. However, there could be restrictions for long travel length or when working with multiple samples in the ibidi Heating System, Universal Fit, for 4 µ-Slides. Please contact us for further details.

Is oil immersion microscopy possible with the ibidi Heating System, Multi-Well Plates?

No, the ibidi Heating System Multi-Well Plates is not compatible with oil immersion, because the Heated Glass Bottom is inserted between the bottom of the multi-well plate and the objective lens. Therefore, it is not possible to put oil onto the objective lens in this setup.

Is high-resolution microscopy (40x, 60x, 100x) possible with the ibidi Heating System, Multi-Well Plates?

In principle, high-resolution microscopy is possible, depending on the working distance of the individual objective. The 0.7 mm thick Heated Glass Bottom creates an additional distance of 0.7 mm between the bottom of the sample and the objective.

Examples:

  • When working with coverslip bottom well plates (e.g., ibidi µ-Plate 96 Well), a minimum working distance of ~1 mm is needed.  
  • When working with standard well plates with a thick plastic bottom, a minimum working distance of ~2 mm is needed.

Oil or water immersion objectives are not compatible with the ibidi Heating System, Multi-Well Plates.

Can I be sure that the temperature indicated on the controller is the same as in the immediate vicinity of the cells?

Not 100%. Technically, it is not possible to place a sensor directly into the cell culture medium. The sensors are placed on the surface of the Heated Lid/Heated Glass Bottom and inside the Heated Plate, respectively. Therefore, the indicated temperature on the controller represents the sensor temperature.

To define the temperature in your experimental sample, you first have to adjust the temperature settings on the controller using an unsterile test sample. For this purpose, we provide µ-Slides or µ-Dishes with a perforated lid. Once the settings are defined for the test sample, the same settings on the controller can be used for the experiment.

We recommend adjusting the temperature each time you change anything in your setup. Many factors, such as the microscope model, the format of the cell culture vessel, the amount of medium, or the current room temperature can influence the whole temperature distribution.

Which immersion oils are compatible with ibidi products?

For ibidi Glass Bottom products:
When using ibidi Glass Bottom products with oil immersion objectives, there is no known incompatibility with any immersion oil on the market. All types of immersion oils can be used.

For ibidi Polymer Coverslip (ibiTreat, uncoated, and protein-coated):
When using oil immersion objectives with the ibidi Polymer Coverslip, use only the immersion oils specified in the table below. The use of any oil that is not listed in the table could damage the ibidi Polymer Coverslip. The resulting leakage may harm objectives and microscope components. All immersion oils that are not listed in the table below should be considered non-compatible.

Company Product Ordering Number Lot Number Test Date
ibidi ibidi Immersion Oil 50101 16-12-27 01/2017
Zeiss Immersol 518F 444960 160706 01/2017
Zeiss Immersol W 2010 444969 101122 04/2012
Leica Immersion Liquid 11513859 n.a. 03/2011
Cargille Type A 16482 100592 01/2017
Cargille Type HF 16245 92192 01/2017
Olympus Silicone Immersion Oil SIL300CS-30CC N4190800 01/2017
Carl Roth Immersion oil X899.1 414220338 01/2017

Which hardware autofocus systems are compatible with the ibidi µ-Dishes and µ-Slides?

Due to the standard refractive index of nD=1.52, the ibidi Polymer Coverslip bottom is considered to be compatible with all hardware auto-focus systems that work with near IR LED or laser light. The following hardware auto focus systems were extensively tested by ibidi:

Company System Test Date Result
Leica Adaptive Focus Control (AFC) 2010 02/2017 No incompatibilities with ibidi labware products
Nikon Perfect Focus System 3 (PFS3) 2012 05/2015 No incompatibilities with ibidi labware products


Note: Channel µ-Slides might give two optical interface signals—one from the coverslip bottom and one from the ceiling of the channel.

Is the ibidi Heating System compatible with hardware autofocus systems such as the Nikon Perfect Focus System, the Zeiss Definite Focus, the Leica Adaptive Focus Control, or the Olympus Zero Drift?

The ibidi Heating System, Universal Fit is compatible with hardware autofocus systems. There is no obstacle between the bottom of the sample and the objective, so any hardware autofocus system can be used.

ibidi Heating System, Multi-Well Plates: We cannot guarantee that hardware autofocus systems function due to the extra glass plate between the sample bottom and the objective. Please contact our technical support for more information.

For a stable focus let the whole system equilibrate in temperature for at least half an hour.

Which objectives are compatible with the ibidi Heating System, Multi-Well Plates?

In principle, all dry objectives are compatible. Due to the 0.7 mm thick Glass Bottom, the working distance must be at least 1 mm when working with coverslip bottom plates (e.g., ibidi µ-Plate 96 Well) or at least 2 mm when working with standard thick bottom plates. Additionally, the compatibility depends on the parfocal length and the limitations of the vertical travel range of the objective.

Oil or water immersion objectives are not compatible with the ibidi Heating System, Multi-Well Plates.

If you are not sure which ibidi Heating System fits your setup, please contact us for further details.