ACME DIAGNOSTICS STANDARD OPERATING PROCEDURE

ACME DIAGNOSTICS STANDARD OPERATING PROCEDURE Procedure Number: II.01.200a Effective Date: 09/14/99 Review and Update: 09/2003 TITLE: ADVIA 120 ROUTINE MAINTENANCE PURPOSE AND PRINCIPLE: The ADVIA 120 Hematology System is a fully automated diagnostic instrument with a throughput of 120 samples per hour (CBC/diff). The analyzer uses whole blood samples to provide Complete Blood Count, differential count, reticulocyte count and body fluid count. PROCEDURE: 1. Assure that instrument, computer, and printer are all on. 2. Perform START UP by following Starting each Shift procedure. Go to Help. Click Quick Guide then Starting each Shift. Follow the procedures for Emptying the waste container, Emptying the overflow bottle, Checking reagents, and Obtaining the background count. 2.1 Procedure for Emptying the waste container: a. Make sure that the analyzer is not sampling. b. Cancel all the Auto Cycles, Auto Wash, Auto Rinse, Auto Standby, and Auto Startup. c. Disconnect the level-sensor switch (1). d. On the waste removal assembly, turn the mode selector knob (2) from NORMAL to EMPTY. e. The waste in the container should start to empty. It will take between two and five minutes to completely empty the container. When you see air bubbles in the discharge line (3), the container is empty. II.01.200a Advia 120 Maintenance 9/2003 Page 1 of 9

f. Once the container is empty, reconnect the level-sensor switch (1) and turn the mode selector knob (2) back to the NORMAL setting. g. Reset any auto cycles that you cancelled in step 2. 2.2 Procedure for Emptying the Overflow bottle: Visually check the fluid level in the small, overflow bottle located to the right of the RBC sample and sheath pumps. If it has any liquid in it, empty the bottle, then clean the vent lines and chambers. 2.3. Procedure for Cleaning the Vent Lines and Chambers: Note: Clean one vent line or chamber at a time and analyzer mode should be Ready to Run. a. Remove the tubing from the overflow bottle that leads to the vent of the Pathway or chamber that you want to clean. Or, if there is no overflow tube installed, attach a 12-inch piece of tubing to the vent. b. Put the free end of the tube into a beaker of 25% solution of household bleach and water. EZ KLEEN MUST be used to clean the perox vacuum shuttle chamber and vent line! c. Open the valve that will create a vacuum to aspirate the bleach solution from the beaker through the vent line and chamber to the main waste line at the bottom of the UFC block.. To open a valve, on the Utilities menu, click the Exerciser tab. Click Valves from the list on the left. Click the appropriate valves. The appropriate valves are: Baso chamber open V13 Perox vacuum shuttle chamber open V18 and V17 (USE EZ KLEEN ONLY) Retic chamber open V65 RBC chamber open V10 RBC/baso/retic vacuum shuttle chamber open V22 and V21 Hemoglobin chamber open V36 d. Allow alternate segments of household bleach solution and air to run through the line and the chamber until they are clean. e. Immerse the free end of the tube into a beaker of deionized water to rinse the line and chamber. f. Remove the tube from the beaker and close the valve. g. Exit the Exerciser. h. Replace the tube into the overflow bottle, or if it was not an overflow tube, remove it from the vent opening. 2.4. Procedure for checking reagents: a. Use the Startup tab or the Reagent Log tab to check the supply of all reagents except ADVIA 120 DEFOAMER. Visually check the supply of ADVIA 120 Defoamer. b. If you need to replace reagents, use the Reagent Log tab. II.01.200a Advia 120 Maintenance 9/2003 Page 2 of 9

2.5. Procedure for Obtaining the background counts: Run a background count cycle to obtain a BASO WBC background count, a platelet background count, and an HGB Trans (Hemoglobin baseline Transmission). a. On the Routine Operations menu, click Startup. b. Click Refresh. c. The background results are color coded. Green - Within range Red - Out of range If any result is out of range, click Refresh on the Startup tab to run another background count cycle. If any result is still unacceptable, perform a system wash. 3. Perform any scheduled maintenance that is due. Go to System Log then click on Service Log. Click Scheduled. Perform all maintenance that is checked. When done click on Update Log. Weekly or after 2000 samples 3.1 Procedure for Washing the RBC/baso/retic flowcell: a. Prepare 25% solution of household bleach and water. b. On the Utilities menu, click te Hydraulics tab. Click Flowcell Wash and select RBC/baso/retic flowcell. c. Hold the 25% bleach solution under the open-tube sample probe and click Start in the window. d. Do not push the aspiration paddle. The vacuum will aspirate the solution. e. Hold the beaker until the wash block starts to move down, approximately 80 seconds. f. The flowcell cleaning will continue approximately 2.5 minutes longer. 3.2 Procedure for Washing the Perox flowcell: g. Prepare a beaker with EZ Kleen. h. On the Utilities menu, click te Hydraulics tab. Click Flowcell Wash and select Perox flowcell. i. Hold the beaker ofez Kleen under the open-tube sample probe and click Start in the window. j. Do not push the aspiration paddle. The vacuum will aspirate the solution. k. Hold the beaker until the wash block starts to move down, approximately 80 seconds. l. The flowcell cleaning will continue approximately 2.5 minutes longer. 3.3 Procedure for cleaning shear-valve and aspiration pathways in the UFC Anaylzer mode should be Ready to Run. a. Prepare a beaker of bleach and another beaker with water. b. On the Utilities menu, click the Exerciser tab. Click Syringe Pumps on the left hand side. Click on the image of the valve II.01.200a Advia 120 Maintenance 9/2003 Page 3 of 9

under Selector valve to Automated Closed then to Manual Closed then to Open. c. Click the valves button on the left. Click valve V72 to close. Closed valve Opened Valve d. Open valves V1, V47, and V74. e. Hold a beaker of household bleach under the open-tube sample probe until 5 ml is aspirated. f. Repeat step e using 5ml of water. g. Close valve V74 and click V72 to open and make sure that V73 is closed. h. Repeat steps e and f. i. Close V74 and V72, and open V73. j. Repeat steps e and f. k. Close valves V1, V47, and V73. l. Click on the Analyzer Status tab to exit. 3.4 Procedure for Cleaning the Shear Valve Analyzer mode should be in Standby. a. Prepare beaker with bleach and fill a wash bottle with DI water. b. Place paper towels directly under the shear valve to prevent any fluid from dripping down into the analyzer. c. Remove the knurled nut by turning counterclockwise. d. Remove the compression spring. e. Remove the rotor by holding the shear valve with one hand and with the other hand rotate the rotor until it can be pulled forward and off the shaft. f. Remove the front shear face by gently rotating the front face until loose, then pull forward and remove. DO NOT use sharp objects or excessive force to remove the shear face. g. Place the front shear face in the beaker of bleach for 10 minutes, then rinse thoroughly with DI water. h. Rinse the rear shear face by using the DI water bottle. Be sure to use paper towels to catch the dripping water. DO NOT wipe either of the shear faces with paper towels. II.01.200a Advia 120 Maintenance 9/2003 Page 4 of 9

i. Put the shear valve back together. Be sure to align the front shear valve with the smaller loops at 9 and 11 o clock positions and the large loop at 5 o clock. 3.4 Inspect the centering collars for salt buildup. Clean if necessary. 3.4.1 Procedure to clean the Autosampler centering collar a. Turn off the analyzer b. Tilt the front cover down. c. Remove the sample line from the bottom of the needle base. d. Loosen the two thumb screws and tilt the autosampler assembly forward. e. Pull the spring-loaded knob and turn a ¼. f. Remove the centering collar by pulling it up and out. g. Place the red needle cover over the needle. h. Remove the three lines from the collar. i. Place the collar in a beaker filled with 25% bleach and soak for 5 minutes. j. Remove any remaining residue using a cotton swab then rinse with DI water. k. Use a stylet or heavy fishing line to clean the three ports and the center bore of the collar. l. Attach a piece of tubing to a syringe, then flush each port with DI water. m. Reconnect the three tubes to the collar. n. Remove the needle cover and replace the collar. Be careful not to bend the needle. Turn the spring loaded knob back to its original position. Carefully pull on the collar to check the collar is put in correctly. If the collar can be removed check the spring loaded knob. o. Reposition the autosampler assembly and that it is seated correctly over the guide pins, tighten the thumb screws. p. Reconnect the sample line to the base of the collar. q. Close the analyzer cover and turn the power on. 3.4.2 Procedure for cleaning the Manual closed-tube centering collar. a. Turn off the analyzer b. Open the front cover. II.01.200a Advia 120 Maintenance 9/2003 Page 5 of 9

c. Pull the centering collar up. d. Remove the tubing from the collar e. Place the red needle cover over the needle. f. Place the centering collar in a beaker filled with 25% bleach solution for 5 minutes. g. Remove any remaining residue with a cotton swab and rinse with DI water. h. Use a stylet or heavy fishing line to clean the port and center bore of the collar. i. Attach a piece of tubing with a syringe to the port and flush with DI water. j. Reconnect the tubing to the collar. k. Remove the red needle cover and replace the collar, being careful not to touch the needle. l. Close the analyzer cover and turn the power on. 3.5 Inspect the perox-cap vent hole for buildup and clean if necessary. 3.6 Procedure for Shutting down the system a. On the Routine Operations menu, Log OFF. c. Click Shut down NT. Wait 1-2 minutes while the software shuts down. d. When the message appears It is now safe to turn off your computer, turn the analyzer off then switch the computer off. e. Leave the system shut down for 10 minutes f. Turn the computer on. g. When the computer displays the Begin Logon message, press Ctrl, Alt, and Delete at the same time to log on to Windows NT. h. Enter the operator name and password in the Logon Information and click OK. i. Click on Advia 120 j. Turn on analyzer. Monthly or after 8,000 samples 3.7 Procedure for inspecting and cleaning the syringes and plungers. a. Turn the analyzer off. b. Turn the thumb wheel counterclockwise until the plunger is lowered about ½ inch. c. Disconnect the tubing from the fittings, then disconnect the fittings from the syringe head. d. Remove the nut and washer at the top of the syringe and slide the syringe down and remove. e. If inspecting and cleaning the 50 ul syringe, remove the small plastic bushing inside the syringe by using a paper clip to pop the bushing out. f. Remove the plunger from the syringe. II.01.200a Advia 120 Maintenance 9/2003 Page 6 of 9

g. Inspect the syringe. Clean with 25% bleach solution and rinse with water if dirty. h. Inspect the plunger. Clean with a kim-wipe and 25% bleach solution and rinse with water if dirty. i. If the plunger appears to be worn or seems to be loose when replacing in the syringe, replace the plunger. (See procedure 3.10 ) j. Wet the plunger with saline and insert into the barrel of the syringe. k. Replace the bushing into the syringe (50-ul only). l. Insert the bottom end of the syringe into the slot at the bottom of the carriage. m. Slide the syringe up and into the mounting hole in the frame with the input port facing left on the RBC/baso/retic channel and facing right on the perox channel. n. Install the washer and the nut. o. Reconnect the tubing to the syringe fittings. 3.8 Procedure for replacing the autosampler and manual sampler needle. a. Turn off the analyzer b. Remove the centering collar. c. Place the needle cover over the needle firmly and turn counterclockwise to loosen. d. Discard the needle along with its cover. e. Clean the needle base using a cotton swab soaked in DI water. f. Remove the clear plastic end from the cover of a new needle. g. Screw the needle clockwise into the needle base. h. Remove the red cover from the needle and save for next installation. 3.9 Procedure for replacing the sheath filters Analyzer mode should be in Ready to Run Note: The filters are not interchangeable. The perox sheath filter is green and the RBC/baso filter is clear. a. Pull the perox sheath filter located on the left side toward you to remove it from its mounting. b. Disconnect the reagent line attached to the barbed fitting located at the top of the filter. c. Disconnect the luer fitting from the connector at the output port located at the bottom of the filter. DO NOT disconnect the reagent line leading from the luer fitting. d. Replace the filter. e. Attach the luer fitting to the connector located at the bottom of the filter first, then connect the reagent line to the barbed fitting at the top of the filter f. Insert the filter back into the mounting clip. g. Repeat the procedure for the RBC/baso sheath filter on the right side of the instrument. II.01.200a Advia 120 Maintenance 9/2003 Page 7 of 9

h. Prime reagent lines 5 times. Every two months or after 16,000 samples 3.10 Procedure for replacing the 50-ul syringe plunger a. Follow procedure 3.7 to remove the plunger. b. Loosen the set screws on the metal bushing on the plunger stem using a hex wrench. c. Remove the plunger and discard. Keep the metal bushing. d. Insert the pin gauge into the top side of the syringe. e. Wet the plunger with saline and insert into the syringe barrel until it touches the pin gauge. f. Replace the small plastic bushing into the syringe. g. Replace the metal bushing onto the plunger stem with the narrow collar of the bushing toward the plunger. h. Push the metal bushing up against the syringe and tighten the set screws. i. Remove the pin gauge from the syringe. j. Reinstall the sample syringe. Every 6 months 3.11 Procedure to replace the 1000-ul syringe plungers. Note: 1000-ul syringe plungers do not have plastic bushings a. Follow procedure 3.10 3.12 Procedure for cleaning the air-circulation filter. Note: Facing the back of the instrument the air filter is located on the bottom right side. a. Slide the filter out of its frame. b. Use canned air to remove excess dust from the filter. c. Flush the filter with DI water. d. Allow it to air dry. e. Replace the filter 4. Run Quality Control materials. Follow Quality control procedure (procedure number II.01.050). 5. At the end of each shift, perform Ending Each Shift procedure. Go to Help menu then click on Quick Guide then Ending Each Shift. Follow procedures for Cleaning the perox chamber, Washing the RBC/Baso/Retic flowcel, Washing the system, Performing End of Day procedure and Logging Off. 5.1 Procedure for Washing the system: a. On the Utilities menu, click Hydraulic Functions. b. Click System Wash, select 3 or 5 for the number of cycles, then click start. 5.2 Procedure for performing the End of Day: a. On Customize menu, click System Setup, then click Tools Modify. II.01.200a Advia 120 Maintenance 9/2003 Page 8 of 9

b. When the tools list appears, double-click End of Day. c. Select the SID Reset check box to perform a SID RESET. d. At the end of the month, select the QC Close Out check box to transfer the Daily QC file into Cumulative QC file. Use the list box to select close out for all controls or a specific control. e. Select the Purge Database check box. f. Note values in Total Samples and to be purged fields. In the to be purged field, type the number of record, if any, you want deleted from the All complete file. g. Click Ok to confirm and start. 5.3 Procedure for Logging Off: a. On the Routine Operations menu, click Log On/Off. b. Click Log Off. REAGENTS AND EQUIPMENT: 1. CBC Timepac (catalog no. T01-3620-52) 2. Diff Timepac (catalog no. T01-3621-52) 3. AutoRetic (600 test) (catalog no. T01-3622-01) 4. DEFOAMER (catalog no. T01-3625-01) 5. EZ KLEEN (catalog no. T01-3624-54) 6. Sheath Rinse 20-L (catalog no. T01-3623-01) 7. Bayer Advia 120 instrument. REFERENCES: 1. ADVIA 120 Hematology System Workbook, Customer Training Manual, and Operator s Guide. REVIEW DATES AND SIGNATURES: Chief Medical Officer: Associate Chief Medical Officer: II.01.200a Advia 120 Maintenance 9/2003 Page 9 of 9