rinciples of measurement -Biomedical engineering 测量原理-生物医学工程.ppt

Principles and Technology SYSMEX EUROPE GMBH Principles and Technology SYSMEX EUROPE GMBH DC Detection Method The Pulse Sample Beaker lFirst Dilution 1:500 (WBC) lSecond Dilution 1:50.000 (RBC) Sample Beaker DB - 1 Dilutions for Semiautomatic Analyser Principles and Technology SYSMEX EUROPE GMBH DC Detection Method The Pulse WBC Dilution + 3 Drops Quicklyser for Lysing and HGB RBC Dilution Dilutions for Semiautomatic Analyser Principles and Technology SYSMEX EUROPE GMBH DC Detection Method The Pulse External Electrode ApertureInternal Electrode Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Transducer Vacuum Blood cells Resistance Constant Current (ca. 100 V) Internal ElectrodeExternal Electrode Aperture Sample Beaker Blood Suspension Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Transducer External Electrode ApertureInternal Electrode Vacuum Principles and Technology SYSMEX EUROPE GMBH DC Detection Method External Electrode Aperture Internal Electrode U = R x I The Pulse Principles and Technology SYSMEX EUROPE GMBH DC Detection Method U = R x I External Electrode ApertureInternal Electrode The Pulse Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Time sec Pulse Picture Volume fl Pulses / Volume Blood suspension Transducer Start-Sensor Stop-Sensor Manometer Counting Time (defined Volume) forerunafterrun Absolute Counting I Principles and Technology SYSMEX EUROPE GMBH Time sec Pulse Picture Volume fl Pulses / Volume Blood suspension Transducer Start-Sensor Stop-Sensor Manometer Counting Time (defined Volume) PrerunAfterrun DC Detection Method Absolute Counting I Principles and Technology SYSMEX EUROPE GMBH Time sec Pulse Picture Volume fl Pulses / Volume Blood suspension Transducer Start-Sensor Stop-Sensor Manometer Counting Time (defined Volume) PrerunAfterrun DC Detection Method Absolute Counting I Principles and Technology SYSMEX EUROPE GMBH Time sec Pulse Picture Volume fl Pulses / Volume Blood suspension Transducer Start-Sensor Stop-Sensor Manometer Counting Time (defined Volume) PrerunAfterrun DC Detection Method Absolute Counting I Principles and Technology SYSMEX EUROPE GMBH Time sec Pulse Picture Volume fl Pulses / Volume Blood suspension Transducer Start-Sensor Stop-Sensor Manometer Counting Time (defined Volume) PrerunAfterrrun DC Detection Method Absolute Counting I Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Transducer Sample Start Stop Defined Volume Vacuum Counter offan Display Sum of Pulses / Volume on Pulse Processing digital Absolute Counting Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Transducer Sample Start Stop Defined Volume Vacuum Counter offan Display Sum of Pulses / Volume on Pulse Processing digital Absolute Counting II Signal-Pulses Disturbing Pulses Signal-Pulses Discriminator lNot to be changed by User: Sample Volume Dilution Ratio Counted Volume Aperture Diameter lAutomatically checked by the instrument: Counting Time Condition of aperture during measurement (Noise-Control) lAdvantages: No calibration of counters Automatic check of aperture Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Sample Vacuum Reference Sample for Instrument Calibration Pulses per Time electrical Voltage Digital-Analog Converter Analog-Digital Converter (Digital-Voltmeter) Display: digital Volt (= Target Value of Reference Sample) Adjustment of Voltage to Target Value of Reference Sample Pulsprocessing analog Signal-Pulses Disturbing Pulse Signal-Pulses Discriminator Pulsprocessing digital lMeasurement of Particles per Time Period lElements of Uncertainty: Reference Sample State of the Aperture Vacuum Relative Counting Principles and Technology SYSMEX EUROPE GMBH DC Detection Method lMeasurement of particles in a defined volume lCalculation of cell- concentration with known Sample volume and defined Sample dilution lno calibration of counting values Measurement of particles per time period The count is received indirectly through a reference-sample. It is taken for granted that the sample and the reference-sample have the same characteristics and give the same result. Calibration of counting values in necessary Absolute Counting Relative Counting Comparison of Absolute and Relative Counting Principles and Technology SYSMEX EUROPE GMBH 12345678910 11 12 13 14 Pulse Height Volume (Time) DC Detection Method Principles and Technology SYSMEX EUROPE GMBH 1234567891011121314 Cumulative Size Distribution Curve Pulse Picture 10 20 30 1234567891011121314 410 0012 345 43 21 DC Detection Method Histogram Principles and Technology SYSMEX EUROPE GMBH 1234567891011121314 Histogram 410 0012 345 43 21 Cumulative Size Distribution Curve 10 20 30 1234567891011121314 410 0012 345 43 21 DC Detection Method Histogram Principles and Technology SYSMEX EUROPE GMBH Erythrocyte Histogram 25-75 fl 200-250 fl DC Detection Method Histogram Principles and Technology SYSMEX EUROPE GMBH Platelet Histogram 2-6 fl12-30 fl fixed at 12 fl DC Detection Method Histogram Principles and Technology SYSMEX EUROPE GMBH DC Detection Method Histogram Leukocyte-Histogram fixed at 300 fl30-60 fl LDT1T2UD LymphocytesNeutrophilesMonocytes Basophiles Eosinophiles Principles and Technology SYSMEX EUROPE GMBH Operation Sequences Histogram SV2 SV3 SV1 SV4 NC NO C NO NC C CNO NC CNC Diluent Inlet (A) (B) Ball Foat Open to the Air or Pressure Pump Waste (Vacuum) lAll solenoid valves are de-energized. lC (Common) and NO (Normally Open) solenoid valve ports are connected. lBall Float is in the down position. “READY”-Status Principles and Technology SYSMEX EUROPE GMBH Operation Sequences Histogram SV2 SV3 SV1 SV4 NC NO C NO NC C CNO NC CNC Diluent Inlet (A) (B) Ball Float Open to the Air Waste (Vacuum) COUNT KEY on lSV1 and SV 4 are energized. lC and NC solenoid valve ports are connected. lDiluent is aspirated through SV4, SV2, Transducer, SV1 and SV3. lThe Ball Float in the manometer is pushed upward. lThe inner portion of transducer and manometer is automatically rinsed to minimize carry-over effect. Principles and Technology SYSMEX EUROPE GMBH Operation Sequences Histogram Counting lSV1 and SV4 are de-energized and SV2 is energized. lThe diluted sample is aspirated through the aperture and the Ball Float moves downward. lWhen the Ball Float reaches level A, the counter starts counting. lWhen the Ball Float reaches level B, the counting stops and SV2 is de-energized. lThe volume of aspirated sample is the same as the manometer-volume between points A and B. SV2 SV3 SV1 SV4 NC NO C NO NC C CNO NC CNC Diluent Inlet (A) (B) Open to the Air Waste (Vacuum) Principles and Technology SYSMEX EUROPE GMBH Operation Sequences Histogram SV2 SV3 SV1 SV4 NC NO C NO NC C CNO NC CNC Diluent Inlet (A) (B) Ball Float Pressure Pump Waste (Vacuum) lSV1 and SV3 are energized. lPressure Pump is activated. lPressure is applied to the aperture to remove the clog. lThe same function occurs if the FLUSH key is pressed. Clog removal Principles and Technology SYSMEX EUROPE GMBH Operation Sequences Histogram SV2 SV3 SV1 SV4 NC NO C NO NC C CNO NC CNC Diluent Inlet (A) (B) Ball Float Open to the Air Waste (Vacuum) lWhen power switch is turned on, SV1 and SV4 are energized. lDiluent is aspirated through the transducer. lThe same function occurs if the FILL key is pressed. lThe diluent removes diluent and any blood sample remaining in the system. Diluent Fill Principles and Technology SYSMEX EUROPE GMBH Haemoglobin Histogram lHaemoglobin is the dye at the RBC. lIt is a Fe-containing Protein. lHaemoglobin is exclusively produced by Erythroblasts in the bone marrow. lHemoglobin is an important part of the Erythrocytes. lThere is a direct relation between Haematokrit, Erythrocyte-concentration and Haemoglobin. lThe main function of Haemoglobin in the body is to transport Oxygen and CO2. Principles and Technology SYSMEX EUROPE GMBH Haemoglobin Histogram Newborn14 - 26 g/dl Children10 - 26 g/dl Women 12 - 16 g/dl Men 14 - 18 g/dl 0 2 4 6 8 10 12 14 16 18 20 NewbornBabiesChildrenWomenMen Patient Limits Principles and Technology SYSMEX EUROPE GMBH Haemoglobin Histogram lA special reagent is needed, containing Cyanide for lysing RBC lF-Series = Quicklyser Sysmex KCN-Method Principles and Technology SYSMEX EUROPE GMBH Haemoglobin Histogram lSLS means Sodiumlaurylsulfate lSLS ins not toxic (ingredient in a lot of soaps tooth- paste) lSysmex Sulfolyser is now used with the majority of instruments (K-, E-, NE-,SF-,SE-, XE-systems) lOnly Sysmex hematology systems have a stable KCN -free Haemoglobin-measurement as standard. Sysmex SLS-Method Principles and Technology SYSMEX EUROPE GMBH Haemoglobin- Measurement a a b b Heamoglobin molecule Hydrophobic area Hydrophilic area SO 3 - Na + Haemoglobin Principles and Technology SYSMEX EUROPE GMBH a a bb Haemoglobin molecule a Hydrophobic area Hydrophilic area SO 3 - Na+ Haemoglobin Cyanide-free SLS-method Principles and Technology SYSMEX EUROPE GMBH lHydrophobic area of SLS bind to Globin lConformation change lOxidation: Fe2+ Fe3+ lHydrophilic area of SLS bind to Fe3+ SLS-Hb. Fe 2+ Fe 3+ Fe 3+ SLS Haemoglobin Cyanide-free SLS-method Principles and Technology SYSMEX EUROPE GMBH lphotometric measurement at 540 nm (F-Series, K-800, K-1000, K-4500) Lamp Lens Filter Sample Cellpack Flowcell Photosensor Haemoglobin Cyanide-free SLS-method Principles and Technology SYSMEX EUROPE GMBH Lamp Haemoglobin LampLens Filter Diluent Flow CellPhoto Sensor First step: BLANK MEASUREMANT Principles and Technology SYSMEX EUROPE GMBH Lamp Haemoglobin LampLens Filter Diluent Flow CellPhoto Sensor Sample LampLens Filter Sample Flow CellPhoto Sensor Second step: MEASUREMENT WITH LYSED SAMPLE Principles and Technology SYSMEX EUROPE GMBH Lamp Haemoglobin Start Flow Cell is rinsed with Diluent. Measurement and saving of Blank. Measurement of sample dilution Sample value -Blank = HB-value Printing of result End Principles and Technology SYSMEX EUROPE GMBH Haematocrit lHaematocrit is determined by comparing the total or cumulative volume of red blood cells to the volume of whole blood. lDetermined normally by centrifugation or by electric pulse measurement lHaematocrit is besides Haemoglobin an important parameter in anemia-diagnosis. Haematocrit Principles and Technology SYSMEX EUROPE GMBH Haematocrit 0 0,1 0,2 0,3 0,4 0,5 0,6 NewbornChildrenMenBabiesWomen Newborn0,51 - 0,65 Children0,35 - 0,43 Women 0,38 - 0,48 Men 0,42 - 0,52 Patient Limits Principles and Technology SYSMEX EUROPE GMBH Haematocrit lDetermining the volume of corpuscles in the blood lNormally equivalent to mass of erythrocytes. lBlood is drawn into special capillaries, which contain dried heparin. lAfter closing, capillaries are placed into a centrifuge. lIn the centrifuge blood is divided into its corpuscular (cells) and fluid (plasma) components Centrifugation Principles and Technology SYSMEX EUROPE GMBH Haematocrit- Measurement Haematocrit Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Principles and Technology SYSMEX EUROPE GMBH Total volume VT Centrifugation Haematocrit Principles and Technology SYSMEX EUROPE GMBH Centrifuge Centrifugation Haematocrit Total volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifuge VT V Hct (%) = V / VT x100 Centrifugation Haematocrit Total volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Centrifugation lReading is done with a scale out of triangles. 0 0,1 0,2 0,3 0,4 0,5 0,6 0,7 0,8 0,9 1,0 lThe auxiliary line through the border between erythrocytes- and leukocytes gives the heamatocrit. lCV app. 1% Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Transducer Start-Sensor Stop-Sensor defined volume VT Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Pulse Height Ph Pulse Diagram Ph = k x VEry VEry = 1 / k x Ph Ph Pulse hight k Constant VEry Volume of 1 Erythrocyte Principles and Technology SYSMEX EUROPE GMBH Centrifugation Haematocrit Cumulative Pulse Hct (%) = V / VT x 100 V= VEry Ph Pulse hight k Constant VEry Volume of 1 Erythrocyte VT V Principles and Technology SYSMEX EUROPE GMBH Transducer Ph = k x VEry VEry = Ph x 1/k Ph Pulse Height k Konstant VEry Volume Ery Ph Hct (%) = V / VT x 100 V= VEry = 1/k x Ph Hct (%) = VT 1/k x Ph x 100 V VT Cumulative Pulse Height Detection K-Series Haematocrit Principles and Technology SYSMEX EUROPE GMBH Erythrocyte Indices Calculated (Parameters) Principles and Technology SYSMEX EUROPE GMBH Erythrocyte-Indices Mean Corpuscular Volume (MCV) Hct (%) RBC (x 106/l) MCV (fl) = Mean Corpuscular Haemoglobin (MCH) Hb (g/dl) RBC (x 106/l) MCH (pg) = Mean Cellular Haemoglobin Concentration (MCHC) Hb (g/dl) Hct (%) MCHC (g/dl) = RBC-Indices Principles and Technology SYSMEX EUROPE GMBH Biological Variation -40 -20 0 20 40 60 80 100 of quantitative parameters of CBC Deviation from Median in % WBC 103/l 5,0 190 PLT 103/l 14 Hgb g/dl 4,5 RBC x 106/l 42 HCT % 90 MCV fl 34 MCHC g/dl 31 MCH pg Biological Variation Principles and Technology SYSMEX EUROPE GMBH Specifications Example: SYSMEX K-4500 Specifications Principles and Technology SYSMEX EUROPE GMBH l18 Parameter: WBC, RBC, HBG, HCT, PLT MCV, MCH, MCH RDW-CV, RDW-SD, MPV, PDW, P-LCR SCR% (LYMPH%), MCR% (MONO/EO/BASO%), LCR% (NEUT%) SCC# (LYMPH#), MCC# (MONO/EO/BASO#), LCC# (NEUT#) l3 Histograms: WBC, RBC, PLT Specifications Principles and Technology SYSMEX EUROPE GMBH Reliable Measurement Principles for highest accuracy WBC/RBC/PLT DC Detection Method HBGSLS-Haemoglobin Method HCTCumulative Pulse Height Detection WBC-DiffDC Detection Method Specifications Principles and Technology SYSMEX EUROPE GMBH Throughput l80 Samples / Hour Cycle Time with single Samples l45 Seconds / Sample Specifications Principles and Technology SYSMEX EUROPE GMBH Sample Volume lClosed Mode (Sampler)200 l lManual Open Mode100 l lMCP-Mode200 l lCapillary Mode 40 Specifications Principles and Technology SYSMEX EUROPE GMBH Specifications ParametersWBC, NEUT%, LYMPH%, EO/MONO/BASO%, NEUT#, LYMPH#, EO/MONO/BASO#, RBC, HKT, HGB, MCV, MCH, MCHC, RDW-SD, RDW-CV, PLT, PDW, MPV, P-LCR ReagentsDiluentCELLPACK Lyse-ReagentSTROMATOLYSER-3WP haemoglobin-Reagent SULFOLYSER Cleaning Reagent (Detergent)CELLCLEAN Diameter of AperturesWBC100 m RBC/HCT/PLT75 m Throughput80 Samples / Hour (appr. 45 Seconds Cycle-Time / Sample) Principles-DC Detection Method (RBC, WBC, PLT) -SLS-Hemoglobin (HGB) -Cumulative Pulse Height Detection -Calculation from RBC and HCT (MCV) -Calculation from RBC and HGB (MCH) -Calculation from HCT and HGB (MCHC) Sample VolumeSampler Mode200 l Closed Mode200 l Manual Open Mode100 l MCP-Mode200 l Capillary Blood Mode 40 l Data MemoryExternal data memory available (K-DPS) Principles and Technology SYSMEX EUROPE GMBH -PrecisionOne fresh normal whole blood sample was measured 10 times consecutively in Sampler Mode. From those results the Coefficient of Variation has been calculated. WBCCV T 1 flag T1 was detected but not T2 T2 flag Attention: Confirm the result with the microscope if T1 or T 2 flag was indicated. The WBC result will be correct if no flag is indicated. All Leukocytes are counted. Histogram Leukocytes-Histogram Flags Principles and Technology SYSMEX EUROPE GMBH 3. Flag “F1” , “F2” and “F3” The Histogram of the Leukocytes is limited from the outer discriminators LD and UD. F1 F2 F3 There is a potential of mixing populations. F 1 and F 2 move together. Also F2 and F3. Histogram Leukocytes-Histogram Flags All Leukocytes are counted; WBC total is correct. (Assumption: no other flags) T 1 and T 2 were detected. Conspicuous is: The troughs are away from the basis line. Principles and Technology SYSMEX EUROPE GMBH Maintenance Maintenance Interpretation Principles and Technology SYSMEX EUROPE GMBH Maintenance 1.External Check lIs there enough reagent for routine daily analyses? CELLPACKca. 750 Cycles / 20 l STROMATOLYSER-