Bio-Rad Laboratories (>: D-10" Dual HbA,, Program 220-0201 Quick Guide Ce wo * This Quick Guide is for reference use only; for detailed information, see the Instructions For Use. * Consider any materials of human origin as infectious and handle them using typical biosafety procedures. + Wear personal protective equipment while handling all reagents and ‘samples and while operating the O-10 system. Sample Preparation Whole Blood Primer: ‘+ Reconstitute with 1 mL of DI water. * Allow to stand for 10-15 minutes; swirl gently to dissolve. * Stable for 1 day at 2-8 °C. HbA,, Calibrators: + 2 Calibrators (Level 1 and Level 2), ‘+ Reconstitute each vial with 7 mL of cold Calibrator Diluent. ‘= Allow to stand for 5-10 minutes; swirl gently to dissolve. * Stable for 30 days at 2-8 °C. Lyphochek® Controls: * Reconstitute each vial with 0.5 mL of Di water. * Allow to stand for 5-10 minutes; swirl gently to dissolve. * Stable for 7 days at 2-8 °C. * Dilute 1:300 prior to analysis (6 pL of control in 1.5 mL of WastvDiluent Solution). Liquichek™ Controls: ‘= After opening vial, stable for 14 days at 2-8 °C. ‘* Dilute 1:200 prior to analysis (6 pL of control in 1.0 mL of Wash’Diluent Solution). Whole blood samples: * Samples should be collected in vacuum collection tubes containing EDTA. * Stable for 4 days at 2-8 °C or 1 day at room temperature (15-30 °C). ‘= Allow sample tubes to reach room temperature (15-30 °C) prior to analysis. No sample preparation required. ‘If abnormal tube type or sample is less than 2 mL, then predilute 1:300 in a 1.5 mL sample vial (5 pL of sample in 1.5 mL of Wash/Diluent Solution) Prior to analysis. NOTE: The use of Monoject™ collection tubes can result in elevated total area. For more information, see the Instructions For Use.Bio-Rad Laboratories <> Floppy Diskette a pi 5 a fey o S is i 2 by 1, Goto the LOT INFO screen. 2. Press Update Kit 3, Insert the Update Kit floppy diskette in the A:drive ++ Follow the instructions on the screen to proceed with the Update kit Procedure + FRomove the Roppy diskette from the Arve once the procedure is completed. Installing Reagents 1. install new Elution Butfers and WashvDiluent Solution: + Place the system in Sleep state. = Remove the reagent bottles one at a time. = Do not touch the lines below the caps. + Do not wipe the lines. + Place each bottle in the + Alter opening the bottles, proper position on the reagent bottle tray. the reagents are stable for 30 days at 15-30 °C. D-10 only The second botle of Elution Buffer 1 is installed at 200 A,.injactions or 400 A,/FIA,. injections. [Link] to the LOT INFO/Butfer 1 screen. b Press the Volume box to display the Reset Buffer Volume screen. [Link] Reset. NOTE: Resetting the volume in other method. D-10 + Rack Loader If both bottles of Elution Buffer 1 are installed sit the volume is not required. 9, Pevorm a System Flush (MAINTAIN screen) if a diferent lot of reagents installed Switching Methods ‘one method automatically resets the volume in the imultaneously, manually resetting | Place the system in Sleep state. Go to the LOT INFO screen. Press the Method box to display the Select Method screen. Select the desired method. Press Exit, Press Yes to confirm the method change. Press Exit. ‘The selected method is indicated in the status bar. ‘There is no need to perform a system flush unless a diferent lot of reagents installed. Seroneons Priming a New Cartridge ‘A priming run is performed once per new cartridge and also following the decontamination procedure. Priming can be performed with elther method selected. 1. The system should be in Stand By state. 2. Pipet 1 ml of reconstituted Whole Blood Primer into a sample vial. 3. Place the sample vial nto a sample vial adapter labeled with a Primer barcode then place the adapter into sample rack postion 1. Ensure the adapter magnet ane barcode faces the back othe rack. The Primer must be the only sample in the rack: |-10. only nore The priming run must be performed as a separate run, 1, Insert the rack through the rack door. 2. Alter the rack is finished loading, go to the RUN screen. Ensure the sample ID “Prime” appears in the worklst.f not, type “Prime : sample ID for postion 1 using the keypad in the RUNVEdit soreen, Prose oul Press Start. ‘The entire priming sequence lasts 13 min. 6, Press Eject to remove the processed rack 0-10 + Rack Loader NOTE: Separate runs are not needed for priming and patient runs, 1. Insert the rack (with the back of the rack facing the back of the loader into carrier, sliding the rack all the way to the left. ) into the rack | 2, From the RUN screen, press Start. 3. The entire priming sequence lasts 13 min, 4, Remove the processed rack trom the rack carrier Calibration for Short (HbA. ) Program Calibration is performed after priming a new cartridge and as needed for troubleshooting. 1. The system should be in Stand By state. 2. Place the following in a sample rack: Tube ion | Adapter Label | Reagent 1 Calibrator 1 HbA,, Cali | 2 Calibrator 2 HbA, Calibrat i 3 AtcLow Control | HbA,. Control, | 4 tc High Control | HbA,. Control, } 5+10__|NA Patient Samples 3, Ensure the barcodes face the back of the rack. 4, Ensure the “Stop if calibration fails” checkbox is selected in the i SETTINGS/Alert Settings screen; if not, the run will continue after calibration failure, using the last acceptable slope and intercept, | D-10 only 1. Insert the rack through the rack door. 2. Alter the rack is finished loading, go to the RUN screen, 3 . Itnecessary, enter sample IDs for missing barcodes using the keypad in the RUN/Edit screen. Press Done. . Press Start. The calibration report is printed after testing is complete. The slope and intercept acceptable ranges are provided in the Calibrator/Diluent Set Insert, 6. Press Eject to remove the processed rack D-10 + Rack Loader 1. Insert the rack (with the back of the rack facing the back of the loader) into the rack carrier, sliding the rack all the way to the left. ] 2. From the RUN screen, press Start. S. The calibration report is printed after testing is complete. The slope and intercept acceptable ranges are provided in the Calibrator/Diluent Set Insert. ] Remove the processed rack from the rack carrier.Daily Maintenance Pre-Run Checklist Check that the correct Method (HbA1c) is installed. Check buffer/wash levels, lot numbers, and line positions. Check reagent onboard expiration dates. Check cartridge injection count and lot number. Check pump pressure with pump running (MAINTAIN screen): * Flow rate at 1.5 mL/min, 50% Butfer2. * Pump pressure should not «The expected pressure range using this _ fluctuate more than 5%, attidge is 15-75 kgfom®. Prime the lines if needed. Check for leaks during pressure check. Check external waste tank level. check printer paper supply. q LitT UTR SE Ta) a Ce 1ace the following in a sample rack: | bpoooG Tube position | Adapter Label Reagent 1 |AteLowGontrot | HDA,, Control, Level 1 (optional) 2 HbA,. Control, Level 2 (optional)* |__ston | Patient Samples * Contols should be included in the run atleast once per 24 hours, “Predilued patient samples are placed in non-barcoded sample vial adapters. 2, Ensure the barcodes face the back of the rack D-10 only 1, Insert the rack through the rack door. 2. After the rack is finished loading, go to the RUN screen. 3. I necessary, enter sample IDs for missing barcodes using the keypad in the RUNEdit screen. Press Done. 4, Press Start. 5. Press Eject to remove the processed rack. D-10+ Rack Loader 1. Insert the rack (with the back ofthe rack facing the back ofthe loader) into an available position (green LED it) on the rack carrier, sliding the rack all the way to the let 2, From the RUN screen, press Start 3, Additional racks of samples can be inserted into the rack carrier during the run, 4, Remove the processed racks from the rack carrer. NOTE: When the run is complete, the system remains in Stand By state fora set time period (30-90 min. Shutdown Timeout period, as defined in the SETTINGS/General screen); more samples can be run at this time. ‘fa run is not initiated before the Shutdown Timeout period expires, the system enters the Sleep state,Bio-Rad Laboratories <3 14/wOGLOBIN TESTING Acceptance Criteria tem Criteria * 1.0 million to 5.0 milion + Results should not be reported ifthe area is outside this range; the sample should be manually prediluted and rerun. + For some high total area, high HDA, samples (e.g., 15% oF 140 mmol/mol HbA,, with 5 M total area), the Ate peak may elute outside of the established retention time window. Predilute the sample to | approximately 2.5 M total area and rerun. | Quality Control Values should be in range NGSP: 3.8-18.5% + IFCC: 18-179 mmol/mol HDA,, reportable range | + Results outside of this range should not be reported. ‘Any sample with >15% or >140 mmol/mol HbA,. should be suspected of having a hemoglobin variant Total Area range [HOF | 310% does not interfere with HDA,, result Labile A,, (CATCICHD-1) _| 34% does not interfere with HDA,, result Garba yy) 5% coset are wth HOA, reat Hemoglobin S trait and Ctrait HbA,, result is reportable Ta peak appears in the Variant window, the HDA,, result should not be reported. Combined area of 260% should be suspected of having a Variant, S, and C windows] homozygous variant or variant-p-thalassemia phenotype; l the HbA,, result should not be reported. Variant windowCTS D-10™ Dual HbA,, Program 220-0201 Quick Guide UN tem Observation 0) Total Area 1.0 million to 5.0 million @ | Atc and Ao peaks °| A1c and Ao retention times: Correctly identified Consistently in range (refer to current Insert for retention time windows) Cartridge: F peak a [TATCICHD: % Within reportable range Baseline Properly constructed (ie., stable, not driting) ° e © | HbA, result ° ° ‘ic peak shape ‘Sharp and symmetrical 0 ony | 008} ons oa ora one oo) Sra ome Ase , cae ee ie a Tap ss ws Ie = =Jo Bio-Rad Ate - Be the difference BIO-RAD eee ©2104 Do ad Labo Arron sored pa Anat 03 os 2 09<0 140 sa 4s 874 20026401 December 2010Bio-Rad Laboratories (> HEMOGLOBIN T D-10™ HbA,, Program = C€ 220-0101 i Chromatography Training “ Reviewing chromatography is important to ensure accurate results. Please review this supplemental information. ypical Chromatography Please note these characteristics when reviewing chromatograms: @ A\c peak shape: Sharp and uniform (j.e., NOT broad, shouldered, or tailing) @ A0peak: Correctly identified. © Baseline: Starts at 0.02 on Y-axis; stable with no ramping. 008 003} 5 eo) Peakble «1D: RACKB: Peak Reine ‘Awa Are% Union OMS 08D 03 Alt 020 «6757705 Alb ons miss ams 12 F ou sat 30m 09 tao ter ain am te Qe Fan dm ms fealame [Gocczarion [oe _[rnalinaloe Atypical Chromatography The following are examples of atypical chromatograms. When atypical chromatography occurs, HbA,, results may be impacted. Please contact your regional Bio-Rad Technical Service office for assistance, Refer to the Customer Notification: Release of Cartridge Resin or examples of expected chromatography for the specific resin lot. feXettakc) (yee)ete Weare] Broad Aic Peak 0.08; 0.07: 0.06; 0.05; 0.04; ‘Ac peakis wide 0.08: 0.07- 0.06: Calculated peak area] (shaded) does not align with detector peak area 0.05: 0.04: 0.03; ‘Ate peak is wide 0.02 0:0 1:00 2.00 3:00Alo ‘AO Peak Misidentifies .04} & 003) Be Peg 0.02} | a0 1:00 200 | Peak table ID CAL? Pak Rime Height Areas Arca % Als 020 8148 37115 13 Alb O2t 7931 25598 09 Unnown 037-4573. :1897 07 F 046 624 4453316 LAleiCH-1 069 3553 18283Poor Baseline: et) Baselineen - aeneenean Taney Bio-Rad Laboratories (> HEMOGLOBIN TESTING VARIANT: II TURBO HbA,, Kit - 2.0 Quick Guide 270-2455EX ce Wo] [la) [exrorr ony This Quick Guide is for reference use only; for detailed information, see the Instructions For Use. + Consider any materials of human origin as infectious and handle them using typical biosafety procedures. + Wear personal protective equipment while handling all reagents and samples and while operating the VARIANT Il TURBO system. Reagent / Sample Preparation Whole Blood Primer: * Reconstitute each vial with 1 mL of DI water. * Allow to stand for 10 minutes; swirl gently to dissolve. * Stable for 1 day at 2-8 °C. Calibrators: * 2 Calibrators (Level 1 and Level 2). * Reconstitute each vial with 7 mL of cold Calibrator Diluent. * Allow to stand for 2 minutes; swirl gently to dissolve. * Stable for 24 hours at 2-8 °C; do not freeze. Lyphochek® Controls: + Reconstitute each vial with 0.5 mL of DI water. + Allow to stand for 5~10 minutes; swirl gently to dissolve. * Stable for 7 days at 2-8 °C. * Dilute 1:300 prior to analysis (6 pL of control in 1.5 mL of Wash/Diluent Solution). Liquichek™ Controls: * Alter opening vial, stable for 14 days at 2-8 °C. * Dilute 1:200 prior to analysis (6 pL of control in 1.0 mL of Wash/Diluent Solution). Whole blood samples: + Samples should be collected in vacuum collection tubes containing EDTA. * Stable for 7 days at 2-8 °C or 1 day at room temperature (15-30 °C). + No sample preparation required. * If abnormal tube type or height of sample is less than 25 mm, then predilute 1:300 (5 pL of sample in 1.5 mL of Wash/Diluent Solution) prior to analysis.Installing the Update Kit CD-ROM 1. Go to the Setup/Test screen. Verify that V2TURBO_A\tc is selected in the Select New Test drop-down list. 2. Insert the Update Kit CD-ROM and click Update Kit. In the Update Kit dialog box, select drive e:\. Select the V2TURBO_Atc file. 4. Click OK. Installing Reagents 1. Install new Elution Buffers and Wash/Diluent Solution: * Remove the reagent bottles one at a time. + Do not touch the lines below the caps. + Do not wipe the lines. » Place each bottle in the proper position on the reagent reservoir module. ‘At 15-30 °C, open bottles are stable for: Elution Buffer A = 30 days, Elution Buffer B = 90 days, and Wash/Diluent Solution = 60 days. 2. Manually enter new lot information and expiration dates in the Setup/Test screen or use Update Kit CD-ROM. 3. Perform a System Flush (Setup/Test/Reagents screen) if a different lot of reagent is installed. Installing a New Analytical Cartridge Priming is performed only for a new analytical cartridge. The prefilter must be replaced when a new analytical cartridge is installed. 1. Replace cartridge (install with arrow pointing up). 2. Go to the Maintain/Instruments screen. Select Do Startup Actions from the Execute Commands list. Click Start. 8. After the startup actions are completed, return the instrument to Ready state by clicking Return to READY state. 4. Place the following in a sample rack: Sample peaien (tase Type _| Reagent 1 | PRIMER PR | Whole Blood Primer (1 mL) 2 | PRIMER PR | Whole Blood Primer (1 mL) 3 | BLANK BL__| Diwater 4 | BLANK BL | DIwater 5 _| BLANK BL | Diwater 6 __|stop E 5, Ensure microvial adapter barcodes are facing the instrument. Place rack on the right side of the VSS conveyor belt. 6. Verify that the system is in Ready state. Go to the Run/Worklist screen and click Start/Stop to start the run. 7. Calibration is required after priming is completed.SS. cae sTE: An Automatic Priming option is available beginn Nor 5.1. This option allows you fo run racks eae He He controls, and patient samples within the same run as the pe operator intervention, To use this option, setup the Priming anna M racks exactly as specified inthis instruction manual (includes barcoded BLANK and STOP microvial adapters). Select the ‘ur Priming checkbox in the Werklst Control clalog box betoro staat CM ey elec} Calibration is performed after priming a new analytical ca 1. Goto the Setup/Sample Types/Calibrator sore Enable Delta Factor is selected. Verify that Stop Worklist i the “Action if outside limits” drop-down list, ‘Selected in 2. Place the following in a sample rack: en. Verify that Tube Sample position Adapter Label | SFMPI° | Reagent 1 [BLANK BL Prediluted sample/control (i mb 2 [Calibrator Level! | 1 | Calibrator Level 1 (1 mL) 3 | Calibrator Level2 | C2 | Calibrator Level 2 (1 mL) 4 |Control Level 1 LC |Lowac 5 |Control Level 2 HC High ac BION | P — |Patient Samples N+1 |Control Level 1 LC |Low QC (optionaly N+2 |Control Level 2 HC | High QC (optional) N+3 |STOP a |e 3. Ensure microvial adapter barcodes are facing the instrument. Place rack on the right side of the VSS conveyor belt. 4. Verify that the system is in Ready state. Go to the Run/Worklist screen and click StarvStop to start the run. 5. Review the Calibrator Averaging/Summary Report, verifying that the slope and intercept values are within range (ie., not flagged). Installing a New Prefilter Replace the prefilter at 500 injections. 1. The prefilter can be installed in either direction. Push the prefilter fir over the Stainless Steel Prefilter Adapter. 2. Place the PEEK Housing into the inlet cap with the arrow pointing int direction of flow (bottom to top). Hand-tighten it clockwise, ensuring not leak. Update the prefilter injection counter in CDM: 1. Goto the Setup/TestCartridges screen. 2. Change the In Use column entry from Yes to No for the used prefil new line is generated for the new prefilter. : 3. In the new prefilter line, enter “N/A” in the Lot # column and 500 in Inj. Limit column, nthe In Use column, select Yes. 2Daily Warm-Up Procedure This procedure must be performed only when using CDM software ver wit PeNe sion 4.03 or earlier; the Automatic Warming Up option can be used ith CDM software version 5.1 or later. Go to the Maintain/Instruments screen. Click Return to Active. In the dialog box, click No (do not perform automatic warm-up operations), Select Do Startup Actions from the Execute Commands list. Click Start. Alter the startup actions are completed, return the instrument to Ready state by clicking Return to READY state. aily Maintenance Pre-Run Checklist Ooooo oooo Check that the correct test (V2TURBO_A1c) stalled. Check buffer/wash levels, lot numbers, and line positions. Check cartridge injection count and lot number. Check prefilter injection count. Check pump pressure on both pumps (with pumps running): es cas ca ar nicuiearre nano. ree eee nes Prime the lines if needed Check for leaks during pressure check. Check waste container level. Check printer paper supply. Check piston/seal wash tubing for liquid. Routine Sample Run 1. Ds 3. Place the following in a sample rack: Tube Sample Position [Adapter Label | “777° |Reagent 1 BLANK BL Prediluted sample/control (1 mL) 2 |Controltevel1 | LC |Low ac 3 |ControlLevel2 | HC |High ac 4toN |— P | Patient Samples N+1_ |Control Level 1 Lc Low QC (optional) N+2 |ControlLevel2| HC |High QC (optional) N+3 |STOP oi Ensure microvial adapter barcodes are facing the instrument. Place rack on the right side of the VSS conveyor belt. Verify that the system is in Ready State. Go to the Run/Worklist screen and click Start/Stop to start the run. NOTE: When the run is complete, the system will perform an automatic wash and remain in Ready state for 30 minutes; more samples can be run at this tim After being idle for 30 minutes, the system goes to Inactive state. 2.Bio-Rad Laboratories {> HEMOGLOBIN TESTING Acceptance Criteria Item Criteria fe + 1.0 million to 3.5 million *+ Results should not be reported if the area is outside this range; the sample should be manually diluted and reanalyzed. Quality Control__| Values should be in range + NGSP:3.5-19.0% HA, + IFCC: 15-184 mmoVmol repoviable range | * Results outside ofthis range should not be reported. + Any sample with >15% or >140 mmol/mol HA, should be suspected of having a hemoglobin variant. Total Area range HOF <25% does not interfere with assay Labile A, (LAtc) __| Noiinterference Carbamylated * No interference hemoglobin (CHb) | + CHb elutes in the LA1c window * P3 peak <5% for hemoglobin variant samples (le., HbS-, HbC-, HbE-, and HbD-trait) + P3 peak <10% for non-variant samples P3 or PA peak + P4 peak <10% ‘+ Ifeither peak exceeds the cutoff, the HDA,. result should not be reported; a fresh sample should be obtained for analysis. Heterozygous hemoglobins HbA, result is reportable E,D,S,andC Combined area of 360% should be suspected of ao EKG having a homozygous variant or variant-6-thalassemia phenotype; the HbA, result should not be reported. “Unknown” peaks _ | No interference[eee ene a VARIANT™ II TURBO HbA, Kit - 2.0 Quick Guide 270-2455Ex | Result Review Item Observation | Total Area 1.0 million to 3.5 million | @| AicandAopeaks | Correctly identified 9 | AteandAo Consistently in range (refer to current retention times Cartridge Insertfor retention time windows) 0 | F peak <25% | P3andP4 peaks | <10% each | HbA, result Within reportable range | Baseline Properly constructed (i., stable, not drifing) © | Aic peak shape Sharp and symmetrical 2 The standardized HDA,, master equation forthe ° Ga Reporting | primary reporting unit appears in the Summary Report. FCC Pea name | mmatmot al z ae = 1 cer SBT = = = 1 Tae ss - = 3 a 5 TRE = = 1 Tar aE 73 = a = T525| —ToH2T| = = T755 rr re oP 3 ar Toa a TotalArea 3,195 611-<«—@| = HDAtC (NGSP) = 4.9 %<—@ 150: 128: 100: Ate 78: 50. 204 ihe al 00. + 7) 0.00 0.85 0.50 0.75 1.00 125 150 Time (min) Bio-Rad Atc + Be the difference iD Citoratoies © 2004 Bio-Rad Laboratories SOOO All rights reserved. December 2015 [Link] 6Bio-Rad Laboratories (> HEMOGLOBIN TESTING VARIANT™ Il TURBO ce HbA, Kit - 2.0 wp 270-2455 & 270-2455EX Bey Chromatography Training Reviewing chromatography is important to ensure accurate results. Please review this supplemental information. Typical Chromatography Please note these characteristics when reviewing chromatograms: @ Aic peak shape: Sharp and uniform (i.e., NOT broad, shouldered, or tailing). @ P3/P4 peak integration: A drop-line appears between the P3 and P4 peaks. © Ao/A2 peak integration: A drop-line appears between the ° ‘Ao and A2 (not identified) peaks. Baseline: Starts at 0.0 on Y-axis; stable with no ramping. 20.0 17.5. 15.04 12.5. 10.04 %A1c 75. 5.0 2.54 oe > 0.0 000 0.25 0.50 0.75 1,00 1.25 1.50 Time (min.)Atypical Chromatography The following are examples of atypical chromatograms. When atypical chromatography occurs, HbA... results may be impacted. Please contact your regional Bio-Rad Technical Service office for assistance. Resin Release Notes Refer to the Customer Notification: Release of Cartridge Resin for examples of expected chromatography for the specific resin lot. | Poor Aic Peak Shape: [steer WW asic CM aS 20.0 17.54 15.04 12.54 [Calculated peak area (shaded) does not align peak area 10.04 AIC 7.54 5.04 “Ae peakis wide} +—! 0.0- 0.00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.)Poor Aic Peak Shape: 3 BET Tats Wola 9 20.0. Ac .50 0.75 1.00 1.25 4 Time (min.) Poor P3/P4 Peak Resolution: 3 Ree aes ines 20.0. 17.5. 15.0. 12.5. 10.0: %AIC 75: 5.0. 25: 0.00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.)HbA2 drop-line not present after ‘Ao peak 0,00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.) Baseline sloping upward 0.00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.)Baseline at 0.02 V on Y-axis ear LEEEM Nee Ce ere CRORES 0.00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.) 0.00 0.25 0.50 0.78 1.00 1.25 1.50 Time (min.)%A1c Basoline starts below 0.0 on Y-axis and shifts 0.00 0.25 0.50 0.75 1.00 1.25 1.50 Time (min.) Bio-Rad Ac : Be the difference ene Borat (© 2004 Bio-Rad Laboratories 12003430revB All rights reserved. May 2016 www [Link] .