title: NADC MAP 70mer ver 1.0
status: Public on Jul 29 2016
contact.city: Saint Paul
contact.first: Elise
contact.last: Lamont
contact.email: lamo0062@umn.edu
contact.state: MN
contact.address: 1971 Commonwealth Avenue, Veterinary Science, Room 205
contact.country: USA
contact.institute: University of Minnesota
organism: Mycobacterium avium subsp. paratuberculosis K-10
description: MAP_oligo_array_v1_updated.gal
manufacture_protocol: This is a protocol that works with the MicroGrid Compact array spotter from Apogent Discoveries.
  Protocol uses poly-L-lysine coated slides from Fischer Thermo Scientific.
  * Check the room humidity using the software. Ideally, want the chamber humidity around 45%. So if the room is above 55%, you might want to turn on the dehumidifier in the small room and close the door.
  * Open the TAS application. The GRID file is pre set. Load this GRID file.
  * Check to see if the reservoir tank needs filling. There is an indicator between the “fill” and “drain” buttons on the toolbar.
  * Fill the two flat white reservoir trays with distilled water. The black one in the chamber is the main wash station. The reservoir will fill this one.
  * The pins are stored in a white container that should be carefully removed away by pulling the pin assembly out sideways.
  * Secure the pin assembly into the clamp above the sonicator bath and turn it on and let it run for 5 minutes. Then pull it out of the sonicator bath and spray compressed air on the pins. Put it back in the bath for another 5 minutes. No need to dry before putting it into the machine. Always degas the water prior to sonication.
  * Within the application, click the green “Go” button on the menu bar. Now the software will prompt you as to what to do. Its initial instruction is to load the pin tool.
  * Pull out the pin assembly and put it into the machine. The holder is spring loaded so push it up into the hole and twist a quarter turn. Once seated the empty pin holders should be toward the front.
  * Put on gloves and open the sealed new packages of slides and place those on the slide tray in the machine.
  * The first 8 slots on the top tray are reserved for the preblotting slides. The instrument is programmed to place 5 preblots on these slides before going over and doing the array slides to be used in experimental studies.
  * The spotting buffer is 6 x SSC and 3M betaine. That is a 2x stock.
  * Load slides from the back first then work toward the front. Be very careful as you get closer to the pin assembly. The slides should be placed in the hitches on the tray so that they are flush with the lower left tab. This makes the array location on each slide very consistent and helps when scanning.
  * Once the top tray is completely loaded, close the lid, turn on the vacuum using the software and open the lid to check the seal on the vacuum by trying to move one of the slides. PUT THE FIRST SOURCE PLATE ON now as well, since you will not be prompted to do this later.
  * Close the lid and click the okay button to make the second tray move forward. Load that tray with all remaining slides. Remember to date any excess slides you do store after removal from package and use those in the next run, if unexpired.
  * For the MAP K-10 array, there are 15 source plates sealed and double bagged to prevent evaporation.
  * Remove the tape of the first plate and seat it in the silver holder with the “A1” well in the lower left corner. Now you should be ready to click “ok” and you have about 20 minutes before needing to switch out the source plates.
  * Machine is programmed as follows: wash #1, wash #2, black tray wash, dries, dips into source plate, pre-spots 5 times, then spots on slides. Does duplicate, then triplicate for each slide on the trays.
  * For QC, check the initial spots shortly after the run begins. Shine a flashlight at an angle to see the spots. If they are not perfectly aligned, or you think there is a piece of lint that might be ruining the spotting, pull out either a single pin or the entire assembly and look at it under a dissecting microscope to find the problem. Then re-sonicate the problem pin or the entire assembly and try again. Check spotting frequently throughout the print run.
  * When each source plate is completed, which takes about 20-22 minutes, then the computer indicates to put in the next source plate. Place it into the arrayer with the A1 well in the lower left corner, as before. Close the lid and click “OK” in the dialog box on the computer.
  * After the last source plate is completed, the software prompts you to perform a series of clean-up steps.
  * One thing you will be prompted to do is remove the printed slides. The first 8 preblotted slides should be disposed of. Collect the remaining slides, score the top left corner with a diamond pen, and place them in the box the manufacturer provided for storage.
  * The pin tool can be removed for the sonication, air blowing and sonication treatment again. This time, give it a good air blow-drying at the very end before putting them back into the white container.
  * In addition, empty the pins baths (flat white trays) and rinse them out and put them in the chamber upside down.