How to prepare RNA Nano chip for Agilent 2100 Bioanalyzer?

Before we start

Step 1: put the samples in ice if you haven’t already done so.

Step 2: get the reagents and ladder. The reagents are stored in the refrigerator in room 0213B, which is right across the GeneChip lab.

NOTE: some reagents will react with light, so don’t take them out unless you are using them.

Step 3: Let the reagents equilibrate to room temperature for 30 minutes.

Prepare the Gel

Step 4: Allocate 550μl of RNA 6000 Nano gel matrix (red) into a spin filter.

Step 5: Centrifuge it at 1500g (rcf) for 10 minutes.

Step 6: Remove the filter and label the tube “Gel” and the date of production. Use this within the next four weeks.

Prepare the Gel-Dye Mix

Step 6: Allocate 65μl of gel into an RNase-free microfuge tube.

Step 7: Vortex RNA 6000 Nano dye concentrate (blue) for 10 seconds.

Step 8: Add 1 μl of RNA 6000 Nano dye concentrate (blue) to the microfuge tube.

Step 9: Vortex the solution until the dye is no longer visible. (the gel and the dye is evenly mixed.)

Step 10: Centrifuge the tube at 13000 g (rcf) for 10 minutes at room temperature. Label the tube “Gel-Dye” and use it within one day.

Loading the Gel-Dye Mix

Step 11: Put a new RNA Nano chip on the Chip Priming Station (the little device with a syringe on the top), and adjust the syringe so that the plunger is at about 1 ml position.

Step 12: Pipette 9.0μl gel-dye mix in the well marked G (a white G on a black circle, this is the well in the third row of the last column).

Step 13: Close the Priming Station and press the plunger until it is held by the clip.

Step 14: Wait for 30 seconds and then release the clip.

Step 15: Pipette 9.0μl gel-dye in the two other “G” wells (wells marked with a black “G”).

Load the marker

Step 16: Pipette 5μl of RNA 6000 Nano Marker (green) in all the remaining wells, including the ladder well.

Step 17: if there are wells that you are not going to use, pipette another 1μl into those wells so that there is total of 6μl solution in the well.

Loading the Ladder and Samples

Step 18: Pipette 1μl of ladder into the well marked by a small ladder.

Step 19: Pipette 1μl of sample in each well that you plan to use.

Step 20: Vortex the chip at 2400 rpm for 1 minute.

Step 21: The preparation of the RNA Nano chip is completed. Run the chip within 5 minutes

Note: The above information is based on the information provided by Agilent Technology, modified by James Zhou