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2 VP-ITC VP-ITC. VP-ITC Y-Axis ii

3 10-12 V( ) VP-ITC 150cm 75cm NMR LAN VP-ITC VP-ITC 100VOA 3 VP-ITC 1

4 VP-ITC *.itc Origin *.opj VP-ITC / 500ml 5% dcn90 P23 VPViewer 2000 Setup / Maintenance Extended Data Mode / 3 / Tel: Fax:

5 . VP-ITC. VP-ITC rpm µcal/sec ( ) Time (min) Output (Reference Power) T T 0 Differential Power=DP VP-ITC C276 HastelloyC ml 2 2 ( T) T T DP DP T =0 1.4ml l 0.6ml Hastelloy C-276 Cr-Mo-W-Fe ph=2 3 CORROSION RESISTANCE OF HASTELLOY ALLOYS Booklet 3

6 DP H Kb VP-ITC OriginITC (N)K b ( H)( S). 16 Raw Data H Data µcal/sec Time (min) kcal/mole of injectant H Molar Ratio ( K B ) ( N ) G = -RT lnk B = H T S G < 0 K B G H S G 2 H G : H : S : K B : R : T : *K b =1/K d 4

7 . VP-ITC 1) PC Windows UPS ON 2) 3) VPViewer VP-ITC 5

8 VP-ITC 2 1) VPViewer2000 Main Window Window 2) Origin7.0 Real Time Plot Window Origin7.0-Real Time Plot VPViewer2000 Origin VPITCPlot.opj Real Time Plot VPViewer2000 VPViewer2000 Origin7.0-Real Time Plot VPViewer2000 System-Open Origin Origin Origin VPViewer2000 6

9 . 1) VPViewer2000 Main Window System QuitProgram VPViewer2000 VPViewer2000 Main Window Origin7.0 Real Time Plot Window Origin7.0 Real Time Plot VPiewer2000 2) VP-ITCOFF 3) PC 4) UPS OFF OFF ON ON 2 OFF VP-ITC ON VPViewer2000 PC ON 7

- - 1.5 ul 150 ncal I. VP-ITC VPViewer2000 Main Window Thermostat/Calib.

10 ul 150 ncal I. VP-ITC VPViewer2000 Main Window Thermostat/Calib. Default ThermoVac 1) Thermo Vac ) Thermo Vac VP-ITC

11 3) 4) Vacuum O- 5) 15 6) ml. 1) 800RPM ) 2 3mm 9

12 1 2mm 3) 4) 2 3mm

13 5) 2 3mm 6) ) Hastelloy C

14 . 8) 9). 10) VPViewer2000 Main Window ITC Controls 2 Experimental Parameters Total injections Cell Temperature Reference Power DP - 10uCal/sec Initial Delay 60sec - 60sec Syringe Concentration Syringe Concentration Cell Concentration 12

15 ( : M Cell Concentration ( : M) Stirring Speed Data File Name Feedback Mode/Gain DP(Diffarential Power) High LowNone High ITC equilibration Options No Check Temp Fast Equil. Auto - Injection Parameters Volume 1 ml 3 12ul 15 l - 10 ul Duration Volume2-20 sec Spacing sec Filter Period Edit mode All Same Injection Parameters Unique Injection Parameters Spacing Spacing Injection Parameters Injectin Parameter Table Edit mode Injectin Parameters Update Run Parameter Display Run Parameter 13

16 Apply To Rest Injection Parameters 14

17 . 11) 5. HEX DRIVER 0.05inch 12) ITC Controls Open Fill Port 15

18 () 13) 14) Up Dn - 16

19 15 16) Close Fill Port 2Perge >ReFill 17

20 . 17) Start Pre-Stirring Equilibration 18) DP DP Final Baseline Equilibration 19) DP DP ITC Injection #1 X Pre-Stairing Equilibration Final Baseline Equilibration 2 X 1200 sec. Y 0.15 cal/sec. DP X - 20) 18

21 . Thermostatting Thermostat/Calib. Heating/Cooling Jacket/Cells to Set Temperature PreStirring Equilibration ( T) DP ReferencePower Final Baseline Equilibration PreStirring Equilibration DP DP 19

Ranges Saved View 1-Y Min, Sav ed View 1-Y Max Saved View 2-Y Min,

22 Rescale To Show All Auto-View, Auto-View 2 DP YEdit Ranges Full Scale Auto View-1, Full Scale Auto View-2 Saved View 1, Saved View 2 Edit Ranges Saved View 1-Y Min, Sav ed View 1-Y Max Saved View 2-Y Min, Saved View 2-Y MaxY Edit Ranges Auto-View Saved View X Axis OptionsDP X 20

23 X Axis Options Disabled XDP Rescale 25 Scroll 25 21

24 . 1) 3 2) Thermostat/Calib. 30 3) O dcn90 dcn90 VP-ITC 10 4) 5 (v/v) dcn ml 5) ThermoVac 22

25 Thermo Vac ThermoVac 5 dcn90 6) 7) 8) Thermo Vac ml 9). 1) 3010 dcn90 2) VPViewer2000 Thermostat/Calib. 65 3) 65 4) 4 1 dcn

26 . 1) 2) ThermoVac 3) ThermoVac 4) (20 ml ) (20ml )(5ml ) 5) 24

27 . 1.5 ~3.0mm 1.5mm 1) 2)

28 3) 4) 5) mm (0.05 ) 6) 26

29 . 30 1) 2) 1. Remove Old Tip 3) VPViewer2000 Setup/Maintenance Pipete Maintenance 1.Remove Old Tip 27

30 4) 5) 6) 7) 8) 28

31 9) 2. Install New Tip 10) Pipette Maintenance 2.Install New Tip 11) 12) 13) 29

32 3. Poilsh New Tip 14) Pipette Maintenance 3.Polish New Tip 15) 16) 17) ) 30

33 4.Ne w Tip Polished 19) Pipette Maintenance 4.New Tip Polished 20) 21) 31

. Y-Axis Y-axis DP Deferential Power 3 1 1) 15 2) ThermoVac 15 3) VP-Viewer2000 ITC>Start ITC Calibration Run>Yaxis Y-Axis ITC Controls ITC Equilibration Options 4)

Power: 2 Pluse Duration: 300 Pluse Spacing: 600 Pulse #3 Caliblation Power: 5 Pluse Duration: 300 Pluse Spacing: 600 Pulse #4 Caliblation Power: -1 Pluse Duration:

34 . Y-Axis Y-axis DP Deferential Power 3 1 1) 15 2) ThermoVac 15 3) VP-Viewer2000 ITC>Start ITC Calibration Run>Yaxis Y-Axis ITC Controls ITC Equilibration Options 4) ITC Y Axis Calibration 4) ITC Y Axis Calibration Data File Name Default 6 Pulse #1 Caliblation Power: 1 Pluse Duration: 300 Pluse Spacing: 600 Pulse #2 Caliblation Power: 2 Pluse Duration: 300 Pluse Spacing: 600 Pulse #3 Caliblation Power: 5 Pluse Duration: 300 Pluse Spacing: 600 Pulse #4 Caliblation Power: -1 Pluse Duration: 300 Pluse Spacing: 600 Pulse #5 Caliblation Power: -2 Pluse Duration: 300 Pluse Spacing: 600 Pulse #6 Caliblation Power: -5 Pluse Duration: 300 Pluse Spacing:

35 5) Start Run Pre-Stirring Equilibration 6) DPDP Final Baseline Equilibration 7) DPDP 8) 1.txt X 2 X 1200sec. Y 0.15 cal/sec. DP X OK Script Window 9)

36 . c K b tot n c K b tot n c c Kb tot Kb n H c c Kb tot n tot tot M 10 M 34

37 Kb M tot M M 10 8 M ml1.4ml 0.3ml 10 9 Displacement Sigurskjold B,W.(2000) Exact analysis of competition ligand binding by displacement isothermal titration calorimetry. Anal. Biochem. 277, Xtot v/v= tot n 1.5 Xtot v V ( 1.4ml) 250ul 2 n=1 Mtot ul 3 H Kb n mM ph DMSO DMSO DMSO 35

38 . DTT ( ) ITCDTT DTT DTT 1mM DTT TCEP ( (2-) ) 1 2mM TCEP 10mMDTT TCEP. 36

39 Biocalo rimetry John E. Ladbury and Babuer Z. Chowdhry WILEY Biocalo rimetry John E. Ladbury and Michel L. Doyle WILEY 37

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目　次 MicroCal, LLC MicroCalorimetry System Tutrial Guide itc200 System 輸入代理店 :DKSH ジャパン株式会社 MAU130030 Rev. A 4/3/09 目次はじめに...1 ご使用の前に設置環境日本シイベルヘグナー株式会社からのサポート 1. itc200 基本原理...4 Ⅰ. 装置ダイアグラム Ⅱ. 解析データ 2. システムの起動...6

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