User Manual Software Version 5.6
Contents
1. Number of objects to be tracked simultaneously This value depends on the performance of your Automatic Tracking computer and should be set between 50 and 100 Number of objects 50 Search Pos Number of search positions in the Search Pos Pixel 500 image where the system searches for objects using TT random generated positions Search Field Cs Radius of the search area used to find the new Radius Pixel 20 object position during tracking Step width Pixel f2 Step width of search positions within the search area should be smaller than object sizes Exclude Button Show makes the search field visible in the oie Bb video image aten factor Match factor A factor calculated during tracking to Min length Frames 20 check the object parameters e g to identify Er collisions of objects A large match factor makes the Timing system less sensible Setting the match factor to O Runtime sec E deactivates checking Max Stops Frames lo Min length Motile objects which have been i tracked for a short time in frames will be excluded from evaluation Interval cycles Show protocol Runtime Time during which the system is running Load Parameter Save Parameter Max Stops The number of frames during which an object does not move before it is excluded from DK further evaluation Interval cycles Allows reduction of temporal resolution Fig 11 Dialog for setting the sys2. This function is not available in the demo version 2 2 4 Video Options Adjustment Gain Offset etc When you are working with a camera system before working with the system for the first time or when you have changed the optical system you have to set some video options Choose menu command Video Options to open the video options dialog and press the Device Options button To view the result of the settings the video grabber has to be in continuous grabbing mode Change the gain and offset settings until you get an image of high contrast so that the objects appear very clearly separated from the background Meteor Il Standard see also Kap 10 2 Meteor 1394 FireWire see also Kap 10 3 3 Setting of system parameters To ensure a correct image processing by the system you should perform the following steps Calibration 3 1 setting of grey color thresholds 3 2 and setting of object features 3 3 have to be done according to the different image conditions 3 1 Calibration 1 For calibration either an 100x100 um grid e g Makler chamber or an object micrometer has to be put under the microscope 2 Take a snapshot from the scale Menu Video Snapshot 3 Choose from the menu Options Calibration 4 Draw a line between two positions on the scale Click with the mouse onto the first position and then with the left mouse button being pressed draw a line to the second position After this operation the calibrat
3. V DC output of Power Supply Unit to the DC12 labelled socket of the camera Take care of the polarity of the cable Connect the Power Supply Unit to the mains 220 V AC Important information No matter if the camera is switched on or off never aim at the sun or other extremely bright objects Otherwise blooming or smear may be caused Clean the CCD faceplate carefully Do not clean the CCD with strong or abrasive detergents Use lens tissue or a cotton tipped applicator and ethanol 10 2 Video Options Matrox Meteorll Standard 21 gt meldea LAB IMAGING SYSTEM Channel Selection of the video channel Delay Delay Time waiting for signal RE no synchronisation while channel was changed J cht R Brightness A 155 Tuning Brightness Contrast Setting of brightness Delay pas and contrast of the video signal before digitalisation 4 E ae Lut Choose a predefined lockup table to transform aa the image for pseudo colour or inverse 0 transformation only BAW Mode CCIR or RS170 Eo sa Hue Red Blue Adjustment only colour mode Mace Show Info Show Info of the camera when initialising the video i videorecorder MZ Double Buffering activate for CASA Standard Select the video norm for the used Double Video Video video camera see also text Video Standards below Taa Mode Video recorder Time base correction Used MU CCIR E only for older Meteor boards Double Buffering Asynchro
4. choose menu File Open for other formats use File Import Using this command every loaded image is displayed in a new window To change the current image in an existing window choose menu File Load 9 Technical description medeaLAB works in real time i e each image is processed and evaluated with video frequency 40 ms per frame CCIR PAL 30 ms per frame NTSC The user can observe the tracked object whose contour line is marked white during tracking over a user defined or unlimited period of time 10 Appendix 10 1 Matrox Hardware installation Mount the Matrox Board to a PCI Slot in your computer After starting the windows first time the Found New Hardware Wizard appears Check not this time and cancel the wizard Put in the medeaLAB TRACKING CD and follow the steps 1 2 Mount the camera to the C mount connector of your microscope FireWire Camera Connect the IEEE 1394a connector to the connection socket of the Matrox Meteor 1394 situated in the computer and connect the other end to the socket of the camera No Power Supply is necessary because power is transported also via the IEEE 1394a cable 20 meldea LAB IMAGING SYSTEM ca Fig 19 IEEE 1394a connector and cable Analog Camera Connect the video cable to the BNC connection socket of the Matrox Meteor II situated in the computer and connect the other end of the video cable to the Video labelled BNC socket of the camera Connect the 12
5. input you need a special DB 44 connector cable Using a PAL or CCIR camera you are able to select 12 channels Using S Video Y C input 4 channels are available 22 IMAGING SYSTEM Tuning Brightness Control the black level of the picture 128 Brightness Gain Control the camera circuit gain 198 aii Auto Exposure Set to automatic exposure mode wherein the camera controls the exposure level 255 Auto Exposure automatically and continuously fe White Bal U White Bal U Set the U chrominance color component of the data 64 White Bal Y White Bal V Set the V chrominance color component of the data C tti i i SMAA See Camera setting Select the resolution and color M_640X480_Y 30FPS mode for the used camera M Double Bunerine Double Video Only used if multiple boards l Show Info mounted in the computer Resolution Show Info Show Info of the camera when Grabber 100 C 75 Kg initialising the video Display lx Resolution Resolution of the grabber and the l display Video file processing This settings are only available when the video is not initialized Write Sequence of 25 Images Video file processing Writing an image sequence as an AVI File to the disk Type in file name and Load Sequence First Frame Reset number of images and press Grab Sequence OK Set Default MIL configuration Only for advanced users and technical service To reserve enough RAM memory for the MIL system Non
6. paged memory start the tool MiLConfig exe You find this tool on the installation CD in the directory MIL6 1 23 IMAGING SYSTEM E MIL Configuration IIS as New settings Increase the Non paged Memory Size according to your video sampling needs For example With grabber resolution 50 half frames and video standard CCIR b w for a sequence of 30 frames which is a little more than one second you need 3 245 MB memory Warning Do not reserve too much memory from your system Leave min 64 MB for the operating system otherwise Windows will not boot next time 24
7. you can automatically set the grey colour level thresholds for object detection A grey colour histogram in which the ranges of the grey value distribution is shown Select first if the objects are more brighter or darker than the background Radio Button Object brightness Dark or Bright Than you can set the system threshold by pushing the button Set Threshold Finish the dialog by pushing the button Finish 3 5 Counting of objects for evaluation the form parameters In order to prove the correct setting of thresholds and form parameters Start a measurement after activation of the image window by choosing from the menu Image Count 10 E Count Result 29 Index 1 2 3 4 5 6 7 8 9 Fig 10 Count evaluation Your filter settings are correct if all sperms displayed in the video window are marked and listed in the table All changes of your optical system which affect the illumination of your field of view are critical for these filter settings You therefore should always check your filter settings after having changed your system 11 gt meldea LAB IMAGING SYSTEM 3 6 Setting and control of the system options for tracking Activate the tracking window and choose the option Options Tracking Options from the menu The duration in seconds of the tracking measurement can be set by Timing Runtime sec For the measurement of sperm motility the settings shown below have proven as a good default
8. B e lt gt pl Object Appearance Dak O Bright Middle Dark Bright Pseudo Live 4 Default Learn Object Leam Back i Cancel Load Save DK Cancel bead Save Preview on Push Preview again to leave Fig 5 Preview of the image after correct setting of thresholds for optimal segmentation 3 3 Selecting the object features for object recognition Several features are calculated continuously during image processing By setting the proper upper and lower levels for these features unwanted objects can be excluded from being counted For setting choose in the menu Options Object Features Object Features Motility Selected Object Features Area Filter E Fomfactor Filter Flagellum Sensitivity Filter Elites Ura Filter Cutting active Sensitivity SO m Default Load Save e Wax Area dum 200 OF Abbrechen bernehmen Hilfe Fig 6 Dialog for setting the form parameters for a motility and concentration measurement Description of the from measuring parameters Area Object area calculated on the base of the contour Due to the actual calibration the value is converted into metrical units Formfactor Value without unity which is calculated from the ratio of circumference and area Flagellum On the basis of available contour information the system decides whether the object has an appendix flagellum or not When the filter is acti
9. Bake CL IFAN IMAGING SYSTEM medeaLAB Tracking mage Analysis System User Manual 0 50 100 150 200 250 300 350 200 220 240 260 280 O m VG A uan O m O es 5 15 25 35 45 55 65 75 65 935 105 115 125 135 145 155 165 175 185 135 Speed Distribution meldea LAB IMAGING SYSTEM IMAGING SYSTEM is a product of _ meldea a Medea AV Multimedia und Software GmbH Medea AV GmbH Am Weichselgarten 23 D 91058 Erlangen Germany www medealab de Software Version 5 6 3 4 Date January 2010 1997 2010 Medea AV GmbH gt meldea LAB IMAGING SYSTEM Contents 1 PTT SU AUTO e EAR DAS Do 4 bi Progam AS LECCE he ee re ee ee 4 2 Starting and configuration the software cccccccceeccceeeeceeeceeeesseeesseeesseeeseaeessaeessaeeseaeessaeesaeeesaaees 4 Zola Be COMO O ee nr De nadando dE EEE 4 Ei VIGO SONS er E nasa tient etnias RE EAEE 5 2 2 1 Video input source SEISCTION ccccccscccscceeceecececeeccueceeseeecuecaeeceeseuesaeecseseueseeeseeseuesaeenaes 5 Zee VIOCOMMMANZAMON juris AAAA RRR ASAR ANIAR EASAN AAAA AANER 5 2 2 3 Video Source File Playback eee errar rre narra aereas ee narra nara anna 5 2 2 4 Video Options Adjustment Gain Offset eic errar rre aaa 6 3 Setting of system parameters ccccecccceececeececeeceeeecee cess taiii et iieiea 6 Sede CDO sencceossscdescennst vesecedsneenscconacereaccesseequsecuespe A A E A AA E EE
10. E EEE 6 3 2 Setting of the grey level colour intensity thresholds for image segmentation 7 3 3 Selecting the object features for object recognition ci erre erraram 8 3 4 Checking microscopy light conditions and automatic setting of the grey level colour intensity thresholds for image segmentation erre erre eee rr ee aerea cre area eee ce anca e area rena anna 9 3 5 Counting of objects for evaluation the form parameters erre 10 3 6 Setting and control of the system options for tracking erre 12 4 Selection of Image Regions Areas of Interest AOI erre reeanea 12 o MAC UNC sra erhedeuute ater snteuniaancuanenuesaeseatenedsannatecnt unre ts casieelinasineancesnaaaeusaltasntesnbaracesmaaaenhwentneeennpceasmscenas 13 5 1 Contour Tracking and Contour Scan Tracking errar erre 13 52 Shape TPACKINO sesuais sado ia ed ne ee ee 13 5 3 Starting a tracking MEASUEMENN ccccecececesceeceeecceneccenecceneccenecceneccenecceeesceaeecenteneaeensnes 13 6 ECKO AD ICAI ON ett secre eek tl ee ce eg le la aerate aden aiet op eteadiaebia 14 ol Cama ce 61 4196 MOOG rennene DD RS A A na nee ener tert eer 14 6 2 Worm Tracking Mode Caenorhabditis elegans cccccccsccccseeeeeeeeaeeeeaeeeaeeeeaeeeeaeeesaeeesaaeesaaees 16 Oo ONPE BIACKING iV OAC SOME assa aaa aaa anes nase sect encnsiea oes 17 6 4 Shape Tracking demo versi
11. Max Trigger processing When the Trigger is active and a trigger impulse is send the radio button on the right side is also checked Create Level Create a new level by clicking with the mouse to first point and draw a line to the second point while holding down the left mouse button Calc Max Deflection If this box is active during a measurement the system automatically finds the maximal deflection points based an the actual level Trigger Test You can check the trigger signal on the Matrox meteor 2 4 Standard board Pressing trigger test both trigger signals are active until one video cycle 40ms Null trigger signal Matrox Pin24 5V Max trigger signal Matrox Pin8 5V To create a new measurement document choose menu Measure New or push the button j Write in a new filename or accept the automatic generated filename in the format yymmddHHMMSS A new window for showing the track data is created Start the tracking by choosing menu Tracking Automatic or push the button e again The tracking process is running until the runtime is reached Starting the measurement again you will be asked to overwrite or not the previous measurement If you choose no the new track data is added to the document 14 IMAGING SYSTEM If the document contains more than one track you can merge tracks with the function menu Evaluate Merge Tracks The data of the tracking document can be exported as an csv Format f
12. de Continuous grabbing live video and the snapshot mode Snapshot freeze video Select the continuous grabbing mode to focus the image 2 Step Create a new tracking document Menu Measure new In the Select method dialog choose tracking and write in a filename After this operation you see two windows on the screen The video window and the tracking document window 3 Step Activate the tracking document and select Menu Tracking automatic The tracking is running as long as the runtime is set Tracking options 13 IMAGING SYSTEM 6 Tracking Applications 6 1 Fish Tracking Mode After initialisation of the video In the medeaLAB Fish Tracking Version a window showing Fish Tracking parameters appears Fish Tracking a a D Actual Deflection 0 51 Threshold Max Trigger lo M Null Trigger I Mas Trigger Lreate Level Calc Max Deflection Help Trigger Test Fig 12 Actual Deflection During a tracking measurement the actual deflection value is printed in this box The value is in the range O 1 0 Left maximum deflection point reached leads to a value of 0 0 Right deflection maximum reached leads to a value of 1 0 If the actual position is outside the range the output value is 1 Threshold max trigger User defined threshold for Max Trigger signal If the actual deflection falls under the threshold a max trigger signal is send Null Trigger MaxTrigger Activate this boxes for Null and
13. ding down the left mouse button By this all the object grey colour values are added to the threshold range The opposite operation can be done in the Learn Back Mode In this mode all of the background grey colour values are removed from the actual range Three learn modes are available lower fixed upper fixed and band In the band mode both lower and upper levels are adjusted Having dark objects on a bright background please select lower fixed When you have bright objects on a dark background please select upper fixed The band mode is useful when you have objects in an background with dark and although bright regions For increasing the object area you although have the button lt gt and for decreasing the object area you can choose the button gt lt Thresholds Channel 1 Color Thresholds lower level high level Red 4 S Shea gt 255 Green aj ia E 160 gt 255 Blue 4 _ 2 160 ET 255 Mark Color 0 0 0 A e lt gt Object Appeatance J Dark Bright Middle Dark Bright Default Learn Object Learn Back OK Cancel Load Save Ready Fig 4 Left Dialog for setting the grey and colour thresholds The video window on the right shows the object in the current grey color image IMAGING SYSTEM Thresholds Channel 1 Color Thresholds 2 lower level high level Fed da C fien qi gt 255 i Green E _ 160 A 255 Blue 4 E plano 255 Mark Color po
14. he table can be exported to MS Excel menu File Excel Export For export the program MS Excel has to be installed To filter out short tracks or tracks from objects which do not move use the function menu command Evaluate Filter Tracks Track Filter Exclude Tracks Distance lt i Unitas Speed WCL i 0 Unitas Cancel Fig 17 Track Filter Dialog Write in the thresholds for min distance and min speed VCL Curve line velocity After pressing OK the filter result is shown in the track view To Undo this function select menu command Evaluate Undo You can continue also with the next filter operation because the tracks filtered out are not deleted before you press File Save To view statistical information about the tracks speed histograms and speed charts are available by menu command View Show Speed Histogram and View Show Speed Chart 19 meldea LAB IMAGING SYSTEM MM Histogramm Humber Total 52 Mean20 29 Std Dev 5 29 11 25 16 25 Elas 26 25 atas 36 25 4125 46 25 Speed Distribution Fig 18 Speed Histogram 8 Saving and loading images medeaLAB TRACKING allows you to save and load images in BMP TIF JPEG GIF or PNG format for documentation Take a snapshot from the scale Menu Video Snapshot If the video window is active the system offers you the functions Menu File Save and Save as to save the image to the hard disk For loading an image in Windows BMP format
15. hen the intensity of the microscopy light was changed the segmentation threshold settings have to be readjusted So in medeaLAB an function for checking the microscope illumination brightness and comparing with the brightness of a reference image allows you to find the old settings Menu Options Check Illumination Setup Microscope Illumination e Reference Brightness 0 255 ESEZ pen o 11 E Contrast 1 01 E 102 Reference Image Start Live Video Save Save Reference Image Save Reference Image Fig 8 First page of the dialog for checking the microscope illumination settings The video window on the right shows sperms in the current grey image IMAGING SYSTEM Push the Button Start Live Video and adjust the microscope brightness until the difference between the actual image and the reference image is smaller than 5 In this case the green light goes on To store a new reference image please push the button Save Reference Image Push the button Next when ready Setup Microscope Illumination EE mm video Channel t gm 20 40 60 80 100 120 140 160 180 200 220 240 260 280295 o inc a 0 50 100 150 200 250 120 Minimum Maximum Object brightness 160 Red 57 ira Cb Bagh a Green 57 i74 Ble 57 174 Set Threshold CE coca t Fig 9 Second page of the of the dialog for checking the microscope illumination settings dialog On this side of the dialog
16. ile 3 Check that the settings of the grey level colour intensity thresholds for image segmentation Cap3 2 and the object features for object recognition Cap 3 3 are correct 16 IMAGING SYSTEM 4 Choose menu Tracking Start or push the button again to start the tracking 5 You can change the view for the track tables as described in Cap 7 To display information about the stop classification choose menu View Show Table Stops 6 3 Shape Tracking Trypanosome For trypanosome tracking the shape tracking mode is the best solution 1 Open the program window choosing menu Measure New or push the button j Select an existing file or write in a new filename If you are writing a new filename you will be asked to create the new file A new window for displaying the track data and the video is created 2 Load the video to analyse player menu command File Open 3 If using the automatic mode you can also select an rectangle AOI to reduce the image area for object searching see Cap 4 4 Change to Shape Tracking Mode menu command Tracking Shape Tracking 5 Choose menu Tracking Start or push the button again to start the tracking measurement 6 Stop the tracking process by menu command Tracking Stop or pressing ESC The tracking process stops automatically when the video file reaches the end 6 4 Shape Tracking demo version After start of the tracking demo version the tracking for
17. ile choosing menu File Export CSV File 15 IMAGING SYSTEM 6 2 Worm Tracking Mode Caenorhabditis elegans 1 Open the program window choosing menu Measure New or push the button e Select an existing file or write in a new filename If you are writing a new filename you will be asked to create the new file A new window for displaying the track data and the video is created medeaLAB Demo fow ae te Measure Evaluate Mac video Options r a a re T Player Demoversi Di S ta db Cl T FE E 1 Heese em L Demo fow Caenorhabditis elegans Tracking Demo Version G 4 O 22 mm View Table Stop Reactions x ee ree eng Sree eeice 3 2 8 7 1 31 0 0 9 E Hd Ls Fig 13 Form for Worm Tracking 2 Initialisation of the video system Choose either Video Init Video in the menu or click on the toolbar In the Demo Version the video is initialized automatically After initialisation the live video will be displayed in the video window There might be a slight delay You can switch in the menu Video between the continuous grabbing mode Continuous grabbing live video and the snapshot mode Snapshot freeze video Choose the button i to move to the beginning and to the end of the video file Choose the Botton Load Video to load a new video file Demo Version automatically switches to the next available video f
18. ion dialog opens 5 Use the calibration dialog to transform the distance in pixel to a metric unit Choose the relevant metric unit um mm cm For calculations of concentrations cells ml you have to set also the chamber depth meldea LAB IMAGING SYSTEM 6 For performance of a test measurement choose menu option Image Count You should see the measurement result in form of a table where the area value is assigned to a metric unit Check the values for plausibility 3 2 Setting of the grey level colour intensity thresholds for image segmentation The procedure is as follows First take a still image by choosing Video Snapshot By choosing Option Thresholds you can check the current thresholds and if needed set them by the slides When having grey level images the channels red and blue should be set to O lower level and 255 upper level The settings for grey level images are done by the green channel only Using the preview function the resulting image can be viewed on the screen Set the thresholds in such a way that the objects appear with highest contrast possible dark on a bright background Pushing the button Pseudo Live in preview mode shows the binary image in live mode To learn the object grey colour level you can also use the automatic learning functions by pushing the Button Learn Object and Learn Back In Learn Object Mode you can click in the object area and move the mouse over the object by hol
19. k circles in the track view If pointing to a circle by the mouse arrow the main parameters of this track will be displayed in a toolbar Pointing to a track in the track data table below the track view will show the corresponding track will start blinking Std Dev 0 27 Mean Linearity 0 33 7 Fig 14 Track pattern view The numerical data of the tracks are shown in a table view Index Frames Distance mm Speed VAP Linearity Short Stop Long Stop l 153 2 24 0 0696 13 1 E 3 153 2 01 0 0969 251 Z 2 4 153 4 505 0 18 4a 7o 1 T 5 153 Er 0 10 Po to E 2 Mean 0 1l Fig 15 Track table view All Tracks view To analyse the data of a single track you can switch between the modes Show all tracks and Show 2 single track by checking the menu item Evaluate All Tracks 18 IMAGING SYSTEM Index Frame Time ms Pose Post rea mm Form Distance Pixel O O 19 45 104 a9 DON l l 149 elf eal 111 B20 3 61 E 2 295 214 cod 125 lb 4 24 3 3 447 ela eat 123 Ol 1 00 7 4 4 v96 ela eat 115 ob 0 00 z a 4a ele 295 116 vob LA 6 6 Ud elo cod 115 260 1 00 E F 1045 214 293 114 64 rd Fig 16 Track table view Single track view In the mode Show single track the navigation through all of the tracks is possible menu Evaluate Next Track and Previous Track A single track can be deleted by menu command Evaluate Delete Track gt x All data of t
20. ll 1394 when having a fire wire system Select 16 MB DMA memory After installation choose yes want to restart my computer now Hardware Installation when using a Matrox imaging board see Kap 10 1 2 Starting and configuration the software 2 1 Base configuration Before working with the program you can select a path for storing the image and track files using the system options dialog This dialog is avaiable from the program base menu command System Options System Options Image Files Dt TrackinotImages E Track Files D Trackingsfowd Ea Analysis Profiles Window Mode Standard x Save Delete OF Fig 1 System Options dialog Type in a path to store the Track Files and the image files or select an existing directory with the button If the directory does not exist it will be created automatically Track Files have the extension fow Window Mode Switch to window mode to run the system in a free configurable window mode In this mode the video the tracks are represented in single windows Analysis Profiles meldea LAB IMAGING SYSTEM An analysis Profile allows to store and restore the complete system configuration all video options settings motility classification illumination threshold filters calibration illumination etc All the settings will be stored in the xml files btwin32 xml and bv xml To create a new profile with the button Save 2 2 Video setting
21. m is opened and the video file is automatically loaded The player control appears To start the measurement choose menu command Tracking Start or push the buiton j Stop the tracking process by menu command Tracking Stop or pressing ESC The tracking process stops automatically when the video file reaches the end To repeat the measurement navigate to the first frame of the video file using the navigation function of the player control see Cap 2 2 3 and start the tracking process again as described above If the tracking process is running you see the tracking points marked as red crosses over the video The tracks are shown in the track view After the tracking process is finished the track data are also shown in the table view below In the demo version the system is automatically configured to the setting Shape Tracking Automatic Mode In the automatic mode the object tracking positions are automatically selected by the system You have the capability to switch to manual mode In the manual mode you can select the tracking positions manually by multiple mouse clicks Change to Manual Mode by choosing menu command Tracking Manual Mode Choose menu Tracking Start or push the button again to start the tracking measurement and click over the object you want to process 17 IMAGING SYSTEM 7 Data evaluation The tracks area shown in a view on you tracking form The endpoints of the tracks are marked by small blac
22. nous grabbing An image is grabbed same time while the last grabbed image can be processed Useful to perform real time processing of 25 Frames sec Display 1x Double Video Only used if multiple boards mounted in the computer Resolution Grabber 100 75 sS0g video file processing EWideos Sterlett 4100 avi Resolution Resolution of the grabber and the display 100 768 x 576 european standard or Write Sequence of 25 iisliise 640 x 480 american standard This settings are only available when the video is Load Sequence First Frame Reset not d 4 Set Default Video file processing Writing an image sequence as an AVI File to the disk Type in file name and number of images and press Grab Sequence Video Standards e M CCIR European video norm b w BNC or S Video S VHS Connector e M PAL European video norm colour BNC Connector e M PAL YC European video with Y C colour S Video Connector e M_RS170 American video norm b w e MNTSC American video norm colour e M NTSC YC American video norm colour Using a S Video S VHS Connector e Use DCF Use a special Matrox format file to initialize the frame grabber The DCF Files are available on the installation CD in the directory MIL7 5 Matrox drivers meteor_II dctf 10 3 Video Options Matrox Meteorll 1393 1 When connecting to the BNC video connector video channel1 is used For connecting to other video input channels or Y C
23. ntast The difference between Contour Tracking and the Contour Tracking Scan is that in the Contour Tracking mode the system search for objects in restricted positions while in Scan mode the whole image is processed which is more time consuming The Contour Tracking mode is available for real time video stream processing while Scan mode is useful in playback processing The setting of the grey threshold and object features are essential for these tracking modes menu command Options Thresholds and Options Object features 5 2 Shape Tracking In this mode the system looks automatically for relevant tracking positions which normally are found on a object edge etc This mode is very powerful for tracking objects which appear not very homogenous and have a low contrast While this method is not very specific to the objects you normally want to track you get a lot of tracks which are not relevant for your measurement The system allows to filter out these data by data evaluation filters see Kap 7 In the manual mode you can add tracking positions by left mouse click and remove tracking positions by right mouse click 5 3 Starting a tracking measurement 1 Step Initialisation of the video system Choose either Video Init Video in the menu or click on the toolbar After initialization the live video will be displayed in the camera window There might be a slight delay You can switch in the menu Video between the continuous grabbing mo
24. on cccceccceecceeeeeeeeeeeeeeeeseeeseeeseueesueeseeeseeseeeeueeseeeseeeseeeseeesaeeenes 17 7 Data evaluation eee eee era ee area a esse eeseeeeeeeesaeeseeeeeseeeseesaeeseeeseeseeesaeeeeseeeseeegeees 18 8 Saving and loading IMAGES assirian eanan Nae Eaa aa EEEE EEE aaia aaa 20 9 MESSI ACLS SF ONO es E E E E EEE 20 1 PP ONO a EEEE ES 20 10 1 Matrox Hardware installation ccccccceccscceeceeceececceeeceeceeceecaeeeeesseseeseeeaeeseesaesueseueseeseeseeeass 20 10 2 Video Options Matrox Meteorll Standard erre erra ren re area a arena 21 10 3 Video Options Matrox Meteorll 1393 errar rear re erre nnaa 22 IMAGING SYSTEM 1 Installation Minimal system requirements are a PC with e Pentium Ill 850 MHz processor e 64MB RAM e Operating system of type Windows 2000 or Windows XP Optional e High resolution video camera or Fire Wire camera e Image processing board Matrox Meteor ll Standard or Meteor ll 1394 1 1 Program installation To install the medeaLAB Software put the installation CD into your drive The medeaLAB setup staris automatically otherwise start the program setup msi Choose Typical setup and follow the instructions To install the software administrator rights are essential Optional only when using a Matrox Grabber Card Move to the directory MIL8 0 on your medeaL AB CD and start the Setup batch routine Choose Meteor ll Standard when having an analog system or Meteor
25. s 2 2 1 Video input source selection Before first video initialization please choose a video input source The dialog is avaliable by menu command Video Options Video Options File Select Input Source Video File Input File Video File Camera YVDM FileMame Z debug ida MIL Digicam Prosilica GigE Fig 2 Video Options dialog You can switch between Video File Camera WDM USB and FireWire cameras MIL Matrox Imaging Adapter DigiCam Digital Camera and GigE Camera video input Input File Select the video file for Video File Inout Source or Playback Mode 2 2 2 Video initialization To initialize the video system choose menu command Video Init o Toolbar After initialization the video window shows up possibly with a short delay You can switch in the menu Video between the continuous grabbing mode Continuous grabbing live video and the snapshot mode Snapshot freeze video Select the continuous grabbing mode to focus the microscope image Toolbar d Continuous grabbing Snapshot 2 2 3 Video Source File Playback In the case of Video File as the actual video source the Player dialog appears meldea LAB IMAGING SYSTEM Player File Edit Options Fig 3 Player dialog AC ER the start and stop buttons o D to start and stop the playback or the navigation buttons for navigation through the video file To load another video file choose menu command File Open
26. tem parameters needed for motility analysis Show protocol After the tracking is finished a protocol is printed 4 Selection of Image Regions Areas of Interest AOI AOl s allow you to decrease the image area for tracking and other image operations There are 4 different AOI types available Rectangle Parallelogram Ellipse Circle und Freehandline Start the AOI creation process by menu command Image AOI Toolbar Rectangle AOI E Parallelogram Ellipse Circle le lole Freihand 12 IMAGING SYSTEM Delete Active AOI x To create a rectangle AOI please move the mouse to left top position and press left mouse button Move to bottom right position by holding left mouse button downs and release 5 Tracking In the system 3 tracking modes are available One can select a specific tracking mode by menu command Tracking Contour Tracking Shape Tracking Contour Scan Tracking All tracking modes can run in a automatic mode and a manual tracking mode Switch to manual mode with menu command Tracking Manual Mode In the manual mode you can select objects by moving over them by mouse and click left mouse button 5 1 Contour Tracking and Contour Scan Tracking This mode is for objects which appear relative homogenous and can be separated by a threshold operation The position of the object is calculated from the contour Use this mode when you have a good image quality and the objects have a good co
27. vated only objects with a flagellum are counted The sensitivity for the flagellum detection ranges from O 10 IMAGING SYSTEM Cutting active Activation of an algorithm to cut conglomerates When two or more objects are within a cluster and are counted as one object this algorithm can divide the cluster and this leads to more realistic counting results Fig 7 Cutting of 2 clustered objectss With the exception of the form parameters Area and Formfactor all form parameters can be activated or deactivated The parameters Area and Formfactor are evaluated constitutively By use of the button Filter the upper and lower level of the respective form parameter filter can be set Example In Fig 3 4 the filter for the form parameter area has been set for a minimum area of 20 um and for a maximum area of 200 um The unit um has been selected during system calibration With this setting only objects will be counted which are gt 20 um and lt 200 um Note The parameter settings are separately store for motility and morphology measurements Please change to the desired mode an make the settings 3 4 Checking microscopy light conditions and automatic setting of the grey level colour intensity thresholds for image segmentation Proper setting of the grey colour level thresholds is of utmost importance for automatic object recognition Therefore these settings should not be changed after having been set for a certain preparation W
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