Using the Fluidics Station

We highly recommended that a member of staff is trained in the use of the fluidics station and is given day to day responsibility for the running and maintenance of the fluidics station and GeneArray scanner. This will minimize errors and equipment failures.

The GeneChip 400 fluidics station will perform the following actions:

1. Wash the array repeatedly with a non- and then stringent solution at selected temperatures to reduce the extent of non-specific hybridization of targets to probes.

2. Draw the solutions with R-phycoerythrin streptavidin and biotinylated anti-streptavidin antibody from the "sample" vial into the cartridge and mix it by alternately draining and filling the cartridge at a selected temperature.

3. Fill the cartridge with wash solution ready for scanning.

However, you will still need to manually change the "sample" vial to the first SAPE solution, to staining solution, and to second SAPE solution according to the prompts on the workstation.

Follow the detailed instructions in the accompanying manual to set up your experiments and to prepare the machine for use.

You will need to:

1. Make up the nonstringent and stringent wash buffer solutions and the 2X staining solution as detailed in Subheadings 2.14.1. to 2.14.5.

2. Prepare the SAPE staining solution. In a separate 1.5-mL Microfuge tube for each array to be washed and stained, make SAPE staining solution the as follows:

2X stain buffer 600 ^L

Analytical-grade water 540 ^L

Acetylated bovine serum albumin (50 mg/mL) 48 ^L

Final volume 1200 ^L

3. Split the SAPE solution into two equal aliquots of 600 |L and label the tubes 1 and 3. Store shielded from light at 4°C.

4. Prepare the antibody solution in a separate 1.5-mL Microfuge tube for each array, as follows:

6. When the "fluidics" procedures have finished, hold each array to a light to see if any large bubbles that could interfere with the laser scanning are present.

7. If no large bubbles are present shield the arrays from light and store at 4°C.

8. If large bubbles are present, either manually vent the array and top up with nonstringent wash buffer (solution A) or return the cartridge to its module in the fluidics station for it to be automatically drained and refilled with solution A (see Note 62).

9. If you have a second batch of arrays ready for processing, you will need to clean the fluidics station (as detailed in the manual), before proceeding as for the first batch of arrays.

10. Once all your arrays have been processed, follow the procedure in the manual for closing down the fluidics station.

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