CONFOCOR 3 User Manual
Contents
1. Mode Switch track eve A Traki amp Light Path Acqutsiton LSM Online Acquisibon Acquisition Mode Obyce Dt Scan Mode Frame n Speed Averaging Number simultaneous gt annels a Pinhote 1 00 Ay Units 1 0 ym section ain Amp Offset Track v 453 477 4838 514 S61 633 Status Transmessaon 488 nm Foe urent Z Postion pm tep Size um Zero Position BP 375 445 IR a A li BG39 NFT490 LP470 BP 465 510 IR e Mirror lt gt Stage D Regions Apochromat 4thd t 1 W Kor U 9 1 93 msec Bit Depth Direction a IR Ww Multidimensional Acqursiton O Z Stack D Bleaching 9 Time Series Qs Tie Scan i information On Experiment ContoCor Court rate Methods g Carrier Position System Configuration Light Path lt 2 Ab42 HLF ag Pinhole 0 97 Airy Units Latest Adjustment 0472 Adjust Pinhole Pinhole Diameter pm Auto Adjust o Coarse Position o Fig 3 Software overview 465 510IR and a low laser power of around 0 2 See Fig 4 Confocor3 Panel BP 373 445 IR OF BG 39 NFT4 90 P 470 APD O A KEIR Ch s B A LO Clad L Mirror LP S05 Ch3 ale mad 8 None Stage Focus Fig 4 Sample Light Path 2 Choose scan mode Line Pinhole 1 AU Gain of 100 300 don t want spikes See fig 5 and 6 Acquisition Mode2. CONFOCOR 3 User Manual Location MI 278 inner room 5 Door Code 514 Fig 1 Power Switch Startup Turn on the 2 switches behind the monitor to start the microscope and the lasers See Fig 1 Turn on the power switch for the PC and then log in with username administrator and password imaging 3 Start the Zen 2007 software Then click the Start System button 4 Lower the microscope objective all the way down till the Lower z limit message shows on the microscope panel Then change the objective to the 40x Water Add a drop or 2 of DI water and then place the sample in the holder Raise the objective so that it just touches the bottom of the sample chamber Place the cover on top of the sample and then lower the microscope head 5 In Zen turn on the 488 and or 561 laser line s The 488 line needs to be first set to standby before the On option is enabled See Fig 2 aw Laser Argon 458 477 486 514 Chameleon 690 1064 HeNeia3 633 Fig 2 Laser Controls Software View LSM Panel Finding the Coverslip 1 Inthe LSM panel choose the appropriate light path and filters E g 488 excitation NT 80 20 BP equisition Maintain Macro imaging Configuration AFS F555 AF647 Olver Fast Fmd Continuous Setup Manager Laser vetoed 458 477 514 e osso se Putt nsope Sean Chameteon 690 1064 HeNe633 633 Laser information S amp Imaging Sonfaguration
3. e save the adjustment See fig 10 7 Pinhole TO Pinhole 0 97 Airy Units Latest Adjustment O4 27 010 02 46 27 PM a Adjust Pinhole Pinhole Diameter pm Auto Adjust Position Fig 10 Pinhole settings Under Measure choose Acquisition and set bleach time to Os Measure Time to 10s and Repeat to 5s Click Count Rate to see CPM and count rates to make sure you are studying the sample in the right z plane See Fig 11 Count rate Mye 2tan Fig 11 Confocor Main Controls Click New and then Start to start taking FCS correlation data When done save the data in the E drive See Fig 11
4. po Objective LOC Apochromat 4001 7 W Konm ly virtkh Scan Mode Line Frame Size 512 T Fig 5 Scan Mode Settings w Channels Channel Ch232 Pinhole 1 00 Airy Unis 10 pm Section Gain Amp Offset Fig 6 Pinhole Settings 3 Click New to open a new window and then choose Line Scan See fig 7 Find Continuous Fig 7 Main Controls 4 Increase z of the objective till you get a signal Then zero the z You have now located the bottom of the coverslip See fig 8 a Focus Current lt Positon ym 451 12 Zeno Position Fig 8 z control 5 Go up about 125um in z to find the other end of the coverslip Then zero again 6 Fora 3D sample go up 300um into the solution If 2D sample will be a few microns above this position FCS Measurement 1 Go to the 3 panel Confocor3 and set up the beam path E g 1 color HFT 488 561 and LP505 2 Color NFT 545 BP 505 540 LP 580 See fig 9 beta olig 486 Ab42olig HL i v BP 505 540 IR W CC Chi gt ch Cc Ch gt Chi f beta olig 488 Q ema CC Chi Ch2 CG Che gt Chi LP 580 Fig 9 On the left 1 color Right 2 color sample paths Select the appropriate lasers 488 561 etc and use about 1 power 3 Start the pinhole with 1AU 4 Each pinhole needs to be calibrated Use the coarse setting and then auto adjust the x and y for each When it s don
Download Pdf Manuals
Related Search