Procedure Beckman Coulter ProteomeLabTM PA 800 Steps for sample testing using the SDS MW Method Prepared By: Rafael Hernandez, Jan 2013
Procedure: Logging in Turning on instrument PA800 Refer to Preconditioning SDS procedure. Logging into windows Refer to Preconditioning SDS procedure Logging into the PA800 software (32 karat 8.0 software) Refer to Preconditioning SDS procedure. Choose the SDS MW icon
Procedure: Logging into SDS MW On the 32 karat software window (see next slide), log in by typing the following: Username: PA800 Password: pa800 Project: SDS MW (select from scroll down menu) If you are not in the Direct Control window, do the following: select from the top menu, Control Select Direct Control Select View (see next slide)
Procedure: Turning on and Calibrating PDA Detector Turn the lamp on by clicking on the rainbow icon located on the Direct Control view. Follow the instruction on the lamp control window (see next slide) to turn it on. Calibrate detector by clicking on Calibrate on the PDA Detector Parameters. It will take about 2 minutes for calibration to complete. If calibration fails, make sure that the lamp remains on for at least 20 minutes before repeating the calibration.
Check Buffer Trays used during preconditioning and sample testing Go to the Direct Control View. Click on Home. Trays-rack will move around until it position itself in buffer s BI:A1 and BO:A1. Then, click on Load Open the lid of the instrument, and position the buffer s as indicated in the next two slides (layout of buffer trays and s). The first slide lay out is for 6 injections or cycles. If planning to do more than 6 cycles, an additional row of buffer and gel s is needed for every 6 cycles. The second slide lay out is for 24 injections or cycles. Close the lid. Ensure that is completely close otherwise the trays will not move to the home position (TI:A1 and TO:A1). It may be necessary to push down the lid and the front cover where the capillary cartridge is located.
Layout of the Buffer Trays/Vials during preconditioning and for a CE run of 6 injections (cycles) max Inlet Buffer (BI) Tray on the left Outlet Buffer (BO) Tray on the right H2O Empty Gel NaOH HCl H2O H2O Gel A B C D E F A B C D E F
Layout of the Buffer Trays/Vials during preconditioning and for a CE run of 24 injections (cycles) max Inlet Buffer (BI) Tray on the left Outlet Buffer (BO) Tray on the right H2O H2O H2O Empty Gel NaOH HCl H2O H2O Gel H2O Empty Gel NaOH HCl H2O H2O Gel H2O Empty Gel NaOH HCl H2O H2O Gel H2O Empty Gel NaOH HCl H2O H2O Gel A B C D E F A B C D E F
Procedure: Performing Pre-conditioning Perform preconditioning before CE sample testing. For preconditioning refer to Preconditioning SDS_procedure.
Procedure: Performing CE sample testing using the SDS MW method Prepare sample(s) as indicated in SDS-MW Prep Procedure. For a 24 sample injection or cycle, set the sample s into the Inlet Tray as indicated in the next slide from position A1 (sample 1) to C8 (sample24). The Inlet Tray is located in the back of the instrument sample tray. For loading sample s into the instrument follow Preconditioning SDS procedure section called Check Buffer Trays used during preconditioning.
Layout of sample s for CE sample run Inlet (IS) Tray located on the left buffer/sample compartment 8 8 16 24 7 7 15 23 6 6 14 22 5 5 13 21 4 4 12 20 3 3 11 19 2 2 10 18 1 1 9 17 A B C D E F
Procedure: Performing CE sample testing using the SDS MW method Write a sequence. In the direct control view, go to file, then sequence, then open, then SDS20.seq, and click on Open (see next three slides). The sequence will open, and must be saved as a different file (ex. SDSMW_date_operator initials)
Procedure: Performing CE sample testing using the SDS MW method Write a Sequence (cont d). On the Sequence page, the headings of the chart are the following: Run # indicates the sample injection or cycle for preconditioning. In a 24-sample or cycle format, Run # 1, 8, 15 and 22 are set up for preconditioning the capillary every six sample injections. Ensure that the Method associated with preconditioning is 20cmPreConditioning.met. Run # 2-7, 9-14, 16-21, and 23-28 are set up for samples and SDS-MW analysis. Ensure that the Method associated with these injections is SDS20Separation.met.
Procedure: Performing CE sample testing using the SDS MW method Write a Sequence (cont d). The Inject Inlet heading indicates the position of the sample for that run. For example, for Run # 2, the nomenclature SI:A1 means sample inlet tray, position A1. Make the necessary changes on the chart that will match the position of each of your sample s. The Inject Outlet heading indicates the outlet of the capillary during that cycle. For example, for Run # 2, the nomenclature BO:C1 means buffer outlet tray, position C1. Make the necessary changes on the chart that will match the position of each of your s.
Procedure: Performing CE sample testing using the SDS MW method The next eight slides contain the program graphical screen for each step during the set up of a sequence.
Sequence Page
Procedure: Performing CE sample testing using the SDS MW method Once the Sequence is set up, instrument is ready for the CE run. Go to the sequence page, top menu and select Control. Under Control select Sequence run. A Run Sequence window will appear, then click on Start.