1. FROM IMAGES TO ANSWERS July 2005 Silver Spring and San Diego Advanced Quantitative Image Analysis

2. Agenda Day One 9:00-10:00Advanced Microscopy Suite Setup 10:00-11:00Multi-Dimensional Acquisition 11:00-NoonMulti-Dimensional Analysis Noon-1:00Lunch (Catered) 1:00-2:00Simulated Live Cell Acquisition & Analysis 2:00-3:00Simulated Ratio Acquisition and Analysis 3:00-4:00Piezo Z-Axis Setup & Fast Acquisition 4:00-5:00Group Discussion 5:00PMAdjourn 6:00PMDinner at a Nearby Restaurant

3. Advanced Microscopy Setup FROM IMAGES TO ANSWERS

4. 9-Step Career Program Follow the Following Steps for a Rewarding Career… Install Application Program(s) Install Capture Driver and Save Settings (VPF File) Calibrate Objective(s) Configure and Save Microscope Settings Configure AFA from Saved Settings Acquire Multi-Dimensional Images Get Bigger Grant to Buy More Stuff Win Nobel Prize Retire

5. Camera Setup Install Capture Driver from WEB Site (CDs are Always Out of Date)

6. Camera Setup Create Video Preference File (VPF)

7. Microscope Setup Set up Scope-Pro Configurator Select Microscope and Devices

8. Microscope Setup Set up Scope-Pro Configure Devices Save Common Settings Calibrate Objectives

9. Microscope Setup Calibrate Objectives Carefully Bad Calibrations = Bad Results!

10. Microscope Setup Set up Stage-Pro When in Doubt, Accept the Defaults (They’re Usually Right)

11. Microscope Setup Import Calibrations (Previously Created) When at Least One Objective is Calibrated, the Other Five Tabs Become Available

12. Microscope Setup Here Are the Next Four Tabs

13. Microscope Setup Set Z-Limits Just Above & Below the Focal Plane The Fifth Tab (Acquire) is Most Often Used

14. Microscope Setup Configure AFA using Saved Microscope & Capture Settings

15. FROM IMAGES TO ANSWERS Live Cell Imaging - Practical Issues Silver Spring & San Diego, June 2005

16. QED Setup Install Application Program* Select Capture Driver from Pull-Down List Calibrate Objective(s) Select Microscope Settings from Pull-Down List or Auto-Configure Acquire Multi-Dimensional Images * Check for Program Updates (Support, Update this Software)

17. QED Camera Setup

18. QED Spatial Calibration Setup

19. QED Parfocality & Paracentricty Setup

20. QED Microscope Setup

21. QED Fluoroescence Acquisition Setup

22. Live Cell Practical Considerations

23. When selecting which system to use for imaging living cells, one should consider three things: –Sensitivity of Detection –Speed of Acquisition –Viability of the Specimen See “Light Microscopy Techniques for Live Cell Imaging”, David J. Stephens et al, Science, Vol. 300, pp. 82-86, 4 April 2003

24. What Do I Need to Control? –Camera (Gain, Binning & Averaging) –Speed of Acquisition (Exp. Time, Time Intervals, ROIs) –Dimensions of Acquisition (X,Y,Z, Time, Wavelength, Location) –Sample Environment (Temperature, Gases, etc.)

25. Controlling the Sensitivity of Detection Use a Combination of Gain, Binning and Averaging to control the Sensitivity of Detection.

26. Controlling the Speed of Acquisition Region of Interest Use a Combination of ROI, Binning, Exposure Time and Frame Rate to control the Speed of Acquisition.

27. Control the Dimensions of Acquisition Use a Combination of XY, Z, Color to control the Dimensions of the Live Cell Acquisition. 3D XY Color

28. Control the Viability of the Specimen Use a Combination of Trend Charting, Signals & Actions and Vital Systems Control to control the Viability of the Specimen. Trend Charting Vital Systems Control Signals & Actions

29. Setup of Live Cell Imaging in 5 Dimensions

30. Long Time Lapse (24h single channel) 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Select “Start” to acquire Long Time Lapse

31. High Speed Time Lapse (1000 I/30 sec) 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Calculate # of Images 4.Select “Burst” to acquire at High Speeds

32. High Speed Ratio Time Lapse 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Define Ratio Channels 4.Calculate # of Images 5.Select “Burst” to acquire at High Speeds

33. 3D Time Lapse 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Define # of Planes 4.Define Z Spacing 5.Select 3D Time Lapse 6.Select “Start” to acquire 3D Time Lapse

34. Multiple Log-points Time Lapse 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Define # of Log Points 4.Select Time Lapse 5.Select “Start Multi Acquire” to acquire Multiple Log-points Time Lapse

35. Multi Dimensional Live Cell Analysis

36. Live Ratio Analysis 1.Define Sensitivity of Detection 2.Define Speed of Acquisition 3.Select Time Lapse 4.Define Multiple ROIs 5.Select Trend Feature 6.Display Live or Compute on Movie

37. Ion Channel Imaging 1.Define Background Correction 2.Define Intensity Calibration 3.Define Psuedo Color Display 4.Define Multiple ROIs

38. FRET Imaging 1.Define Background Correction 2.Define Channels 3.Calculate FRET Image

39. Tracking of objects 1.Define Tracking Measurements 2.Define Tracking Options 3.Define Multiple ROIs 4.Filter Data

40. 3D Deconvolution 1.Verify Imported Deconvolution Parameters 2.Define Deconvolution Options 3.Deconvolve

41. 3D Reconstruction and Measurements 1.Verify Imported 3D Parameters 2.Define 3D Options 3.Measure 3D Colocalization

42. Image Pro 3D Suite Automatic Volumetric Measurements Over Time 4D Tracking of objects!! 1.Verify Imported 3D and Time Parameters 2.Define 3D Options 3.Define Tracking Options 4.Track Objects

43. Manage Images and Data with IQbase Manages Images and Data Drives Discovery Facilitates Collaboration Easy to Use Works the Way You Do Grows with Your Organization (Scalable)

44. Competitive Feature Comparison Image-Pro MC Most Comprehensive Package in the Industry; No Charge for Drivers MetaMorph Extra Fee for Camera drivers, Hardware Drivers, 2D & 3D Blind Deconvolution & 3D Simple PCI Extra Fee for Automated Image Capture, Image Processing and Analysis, Dynamic Intensity Analysis, Motion Tracking and Analysis IPLab Extra Fee for Ratio Plus, Fluorescence CV for Mac, MultiProbe for Win, Motion Control, Shutter & Filter Control OpenLab Extra Fee for 3D Module, Morphology Module, Ratio Module, FRET Module, Colocalization Module, Registration Module Intelligent Imaging Extra Fee for Ratio, TTL, 3D, 4D Tracking, Multiwell Plate Module, Statistics Package AxioVision Extra Fee for Z-Stack, Time Lapse, Tiling, Autofocus, Multichannel, Measurements, Imaging Plus

45. Competitive Feature Comparison Smart Camera Control YNNNNN Zoom & Pan Images YNNNNN Advanced Microscope Control YYYYYY Tiling, Mosaic, Slide and Well Scanning YYYYYY Multi-dimensional Acquisition YYYYYY Multi-Color Imaging YYYYYY Time Lapse Acquisition YYYYYY Color 3D with Piezo Focus Y, Extra $ YNYNN

46. Competitive Feature Comparison MCMMIPLab3ICompixAxioV Advanced Morphometric Measurment, Analysis & Classification YYYYYY Colocalization, Translocation FISH, and Fluorescence YYYYYY Ratio Imaging YYYYYY FRET Analysis YYYYYY Cell Tracking YYNYYN Vital Systems Control & Measurement YYNYNN Signals and Systems Control YYNYNN Data Analysis, Recording, and Reporting YYYYYY 3D rendering and measurements YNNYNN 3D Time Lapse Analysis YNNNNN Analog Digital signals support YYNYYN Trend chart analysis Intensity tracking YYYYYN

47. With Similar Features, How to Choose? Price (Highest to Lowest) MM > Zeiss > 3I > Compix > Scanalytics > MC Support (Best to Worst) MC > Compix > 3I > Zeiss > Scanalytics > MM Installed Base (Largest to Smallest) MC >> All Others Combined

48. With Similar Features, How to Choose? Zeiss Loyal Base due to Highty Integrated Systems and Direct Sales Few User Choices Molecular Devices (formerly UIC) Loyal (but Frustrated Base) Vulnerable due to Recent Changes in Personnel and Direction? Intelligent Imaging Innovations Lately Favored by Olympus; a Longtime Zeiss Collaborator (Strange Bedfellows) Reported to Have “More Mouse Clicks are Better” Syndrome Compix Released v. 4.0 this Spring with a Number of New Features and Improvements Considered the Low Price Leader for Basic Image Capture and Microscope Control Scanalytics Dealer Channel Remains Weak with Limited Representation Products Lag Behind Leaders; Typically Partners with Researchers for New Products

49. Updated Software Competitive Assessment Comparison Criteria: It is Impossible to Make an “Apples to Apples” Comparison (I wish There was) This Comparison Basis is Image Pro Plus MC versus Comparable Features in Other Packages The Data were Obtained from Respective Companies on their WEB Sites and Presumed to be Accurate as of June 2005..\My Documents\2005\Image Analysis Competitive Assessment 2005.xls

50. So What! What Do We Tell Customers? If You Have the Need for Speed: Choose QED for Fastest Acquisition when an External Device is Attached If You’re Prowling for Power: Choose Image Pro ®, the Most Powerful Analysis Package on the Planet If You Need Speed and Power: Choose a QED/IPA Bundle Combining the Speed of QED with the Horsepower of Image Pro

51. www.mediacy.com Silver Spring, MD (Headquarters) Worldwide Offices in: Boston, MA, Pittsburgh, PA & San Diego, CA Leiden, The Netherlands Goettingen, Germany Wokingham, United Kingdom Singapore, Republic of Singapore Tokyo, Japan Shanghai, China

52. “Elvis Has Left the Building…”