Grid Preparation

Clipping Autogrids

Prepare the following utensils prior to clipping:

1. blue clipping station with lid

2. c clips

3. c clip rings

4. 6x clipping tools

5. tweezers

6. auto grid tweezers

7. nano cap

8. dewar with liquid nitrogen

9. warm up heater to 80C

Prepare the clipping station:

1. add the c clip rings

2. add the auto grid boxes

3. screw in the lids of the auto grid boxes

Load the clipping tool with c clip:

1. With a pair of tweezers, take up a c clip from its center.

2. Gently insert it into the clipping tool.

3. Push the sides of the c clip to the clipping tool wall.

4. Gently push the clipping tool onto the filter paper .

5. Check top of the clipping tool to locate the c clip.

Close the clipping station and fill with liquid N2 so that your grid boxes and sample is covered.

Warning

Do not fill liquid N2 above the sample, as this might cause the grid squares to break.

Once the clipping station is cool, do the following:

1. Transfer your grid boxes from the ethan container or from your storage puck.

2. Open your grid boxes.

3. Place a c clip ring with flat side down onto the clipping site.

4. With a pair of tweezers, gently take out a grid from the storage grid box and transfer in onto your c clip ring.

5. With the auto grid tweezers, rotate the golden part of the clipping station.

6. Use the loaded clipping tool and place it on top of your grid.

7. Gently push the clipping tool until you hear a gentle clicking sound.

8. Remove the clipping tool and check if the c clip is still on the tool or on the grid.

9. To double check if the c clip is tight you can rotate the autogrid with the auto grid tweezers.

10. Insert the clipped autogrid into the auto grid box, starting from position 1. C clips face towards the front of the box.

Loading of clipped Grids into the Cassette

1. Cool down styrofoam box including the metal device for grid boxes and the autoloader cassette

2. Insert and cool down cassette (silver plate faces front)

3. Cool down tools

4. Once the metal device is cold, transfer auto grid box with clipped grids into the metal device

5. Unscrew the lid of the auto grid box

6. Use the cartidge tool, insert fully into the grid box and take out grid, hold grid firmly

7. Gently Insert clipped grid into the cassete between the springs

8. The small c clip faces towards the front of the cassette

9. Hold the grid firmly with the tool, until it is really inserted in the cassette (otherwise it is very difficult to recover the grid)

For visual illustration wacth the following video:

Preparing Nanocap

1. Gently pour liquid N2 into the nanocup, try not to spill liquid N2 on the sides

2. Firmly press lid for about 10s, wait

3. Top up with liquid N2, press lid again

4. The nanocap is ready when their is no bubbling of liquid N2 anymore

For visual illustration watch the following video:

Unloading Existing Cassette

First, remove the cassette of the previous user from the microscope. Follow the instructions below.

Loading of Nanocap into the microscope

1. Take cold nanocap, remove lid, turn yellow label towards you.

2. Gently insert nanocap into plastic insert of the autoloader, don’t use force, but hold nanocap firmly.

3. Press the dock button at the Talos computer screen.

4. Wait for 1-2 minutes until the old cassette unloads, you will see a message at the screen or via the autoloader tab in the user interface.

5. Remove cassette with cassette holding tool and insert into liquid N2, remove old grids.

For visual illustration watch the following video:

Loading of Cassette with clipped grids

1. insert cassette with clipped grids into the nanocap, insert the cassete from the right hand side.

2. place the nanocap to your left (with the yellow label facing towards you).

3. use the cassette holding tool and hold the cassette from the back.

4. insert the cassette into the nano cap, with the front silver metal plate going in first.

5. load the nanocap into the microscope, follow the instructions above