- When leaving the system for the next user: sign out from Windows.
- Full shutdown:
- Shut down the computer.
- Swith off all laser keys (4).
- Power off the power strips in reverse order (3, 2, 1).
## Hardware Autofocus
The hardware autofocus permits focus stabilization under non-bleaching conditions. It uses backreflection of grid pattern generated with a weak LED (850 nm) to measure the distance between the objective and the cover glass and to keep it constant. An aqueous medium (with a refractive index close to that of water) is required. It cannot be used with glycerol-based or hardening mounting media.
The hardware autofocus permits focus stabilization under non-bleaching conditions. It uses backreflection of grid pattern generated with a weak LED (850 nm) to measure the distance between the objective and the cover glass and to keep it constant. An aqueous medium (with a refractive index close to that of water) is required. It cannot be used with glycerol-based or hardening mounting media.
Objects above the cover glass/medium interface are not monitored. If objects of interest can change their focus position (e.g., cells rounding up during mitosis), the software autofocus should be used.
Objects above the cover glass/medium interface are not monitored. If objects of interest can change their focus position (e.g., cells rounding up during mitosis), the software autofocus should be used.
### A. Continuous mode (constant offset)
#### A. Continuous mode (constant offset)
This mode is convenient to keep the focus at a contant distance above the cover glass while moving the XY stage across large areas of the sample.
This mode is convenient to keep the focus at a contant distance above the cover glass while moving the XY stage across large areas of the sample.
1. Click the button "HW-AF Dialog" in the toolbar on the right side of the application. Alternatively, the dialog can be opened with the menu command Configure → Autofocus.
1. Click the button "HW-AF Dialog" in the toolbar on the right side of the application. Alternatively, the dialog can be opened with the menu command Configure → Autofocus.
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To use a different offset, turn off the autofocus, focus manually on a different position and turn it on again. For convenience, the toolbar also contains "HW-AF ON" and "OFF" buttons. The button "AutoFocus NOW" has no function (it works only with devices from other manufacturers).
To use a different offset, turn off the autofocus, focus manually on a different position and turn it on again. For convenience, the toolbar also contains "HW-AF ON" and "OFF" buttons. The button "AutoFocus NOW" has no function (it works only with devices from other manufacturers).
### B. Stage positions with multiple offsets
#### B. Stage positions with multiple offsets
The following procedure describes the setup of a multiposition time series with multiple offsets (i.e., positions may have different z-coordinates above the cover glass).
The following procedure describes the setup of a multiposition time series with multiple offsets (i.e., positions may have different z-coordinates above the cover glass).
1. In the Time lapse tab, check "Time-lapse series" and the "Autofocus" option. Click the "Config" button right of the "Autofocus" option
1. In the Time lapse tab, check "Time-lapse series" and the "Autofocus" option. Click the "Config" button right of the "Autofocus" option
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@@ -195,6 +179,24 @@ The following procedure describes the setup of a multiposition time series with
