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Measure the gap between the boundary surface of the

cover glass and the position of the observation target

(Z-axis direction) using the method below.



This step is not required for 2D-STORM.

(1)

Step 1:

When the dialog shown on the right is displayed,

select the imaging laser (647 nm). Set the focus to

the boundary between the cover glass and the

specimen by using the PFS offset controller. (Adjust

the power to about 50% to 100% by using the

slider.)

Set the focus to the boundary surface of the cover

glass.

When setting the focus, move the XY stage and view, as

boundary marks, the dye molecules that are

non-specifically adsorbed onto the boundary surface of

the cover glass to identify the boundary surface.

After setting the focus, click [OK]. The XY stage

automatically returns to its original position.

(2)

Step 2:

When the dialog shown on the right is displayed, set

the focus to the target imaging position by using the

PFS offset controller.

After setting the focus, click [OK]. The measured

gap (Z-axis direction) is automatically saved and

used in analysis.

When acquisition is started, the dialog box shown on the

right is displayed. Do not perform any operation using this

dialog box until image acquisition has been completed.

Clicking [Events...], [Pause], or [Refocus] prevents the

acquisition of appropriate images for STORM analysis.

To halt the acquisition, click [Finish] or [Abort]. (If [Finish] is

clicked, the acquired images are saved. Be careful

because, if [Abort] is clicked, the acquired images will be

discarded.)

Chapter 2

STORM Microscopy Operation

7-(1)

Boundary surface of

cover glass

7-(2)

Imaging target surface

Cover glass