Import into excel as a CSV (comma-separated values) file.įor experienced users, a quicker version of the original macro has been included.When colony counting is complete, save the Log file.Run the Count Colonies command as many times as needed.Do not close this window until you are done!!! The counted colonies will be printed into your Log window.Use the left mouse click to select more colonies, andĬtrl +Click to remove colonies. You can now manually correct the detected colonies.Automatically detected colonies will be marked by a red dot.To increase detection of small/faint colonies, decrease prominence. To reduce background noise, increase prominence. These values can be approximated to mean “how much brighter should a region of pixels be compared to its surroundings, before it is recognized as a bright spot” Typical values range from 10 – 35 for densely packed colonies, up to 100 - 120 for colonies spaced further apart. This the parameter that ImageJ’s “Find Maxima” function uses to define bright spots. Select and name the area of interest when prompted.Locate and select the Count Colonies command, then run it.Your image will now be processed to facilitate colony counting.To crop, select relevant area then press Ctrl + Shift + X You will be given the chance to crop your image if needed.Locate and select the Process Image command.Īlternatively, the macro has been assigned the keyboard shortcut F1, simply press this key with the ImageJ toolbar in focus and the macro will run.Open your image containing colonies to be counted.Please ensure that your FIJI is up to date before proceeding. *This macro was written for FIJI is just ImageJ (FIJI). In the ImageJ toolbar, Plugins > Macros > Install > Select CoCo.ijm This is quicker, but needs to be repeated every time FIJI is started up. Copy the contents of CoCo.ijm to the end of StartupMacros.ijm.Open CoCo.ijm in Notepad (or your word editor of choice).Find your ImageJ directory, under the macros folder, open StartupMacros.ijm in Notepad (or your word editor of choice).This permanently installs the macros to your ImageJ toolbar so you do not have to re-install it every time FIJI starts up. Resolution of 300dpi was found to be sufficient, however 720dpi is recommended especially for small colonies. See the example scanned image available in the GitHub Repository. A BLACK background must be used behind the plate to enable sufficient contrast between the colonies and the background. Plates should be placed without their lids, and with the top of the agar facing the scanner. Once the lanes dry on to the agar, place the lid back onto the plate and allow bacteria to grow.Ī scanner is recommended for imaging.Tilt the plate so that the lanes do not merge. The bacterial suspension should flow downwards and create lanes. Tilt the plate towards you at a 45 degree angle, and gently pipette the bacteria on to the plate.Using a multi-channel pipette, take up 10 ul of bacteria from each well.Make dilutions of bacteria suspension and technical replicates using a multi-channel pipette and a 96-well plate.Ensure the surface of the agar is thoroughly dry, to facilitate the “flow” of bacterial suspension droplets down the agar later (see image below).Īn example bacterial growth experiment that CoCo was optimized for is as follows: This is to enable better contrast between colonies and background when the plates are scanned later. If you are using standard round Petri plates this will be around 15(?) ml of media. CoCo can also be used as a standard counter of colonies on agar plates.Īt least 40 ml of media should be poured into a standard square Petri plate, to make up a depth of 5 mm. This is an example of a standard bacterial growth experiment which CoCo was optimized for. The user can modify the detected colonies to enable more accurate colony counting. Then the user is prompted to select area of interest, and the Find Maxima command is used to automatically count colonies. Images are first processed in using standard ImageJ commands such as Subtract Background, Enhance Contrast etc. This macro enables easy counting of bacterial colonies on Petri plates. Bacterial colony counter written in ImageJ macro language
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |