down for the count w
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The Scientist raquo Magazine raquo Lab Tools
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BLOODY COUNTING A stack of Silvio Bruggerrsquos bloodagar plates for growing bacterial coloniesPHOTO BY PASCAL M FREYCOURTESY OF SILVIOBRUGGER
Down for the CountOne two three four Counting colonies and plaques can be tedious but tools exist tostreamline the process
By Amber Dance | May 1 2013
hile studying bacterial resistance topenicillin in medical school Silvio Brugger
was horrified to discover that he and a colleaguewould have to count colonies growing on 100 orso petri dishes every day He thought ldquoOh myGod I cannot do thisrdquo says Brugger now workingat the University of Bern in Switzerland ldquoIt wasquite painfulrdquo
Brugger is hardly alone if you want to know howmany bacteria or viruses you have the standardmethod is to plate serial dilutions and count theresulting colonies or plaques While somescientists may not mind the repetitive activitymdashldquoIdonrsquot find it too badrdquo says virologist VincentRacaniello of Columbia University in New Yorkmdashothers find the work incredibly tedious Yet tallyingeach speck by hand remains the technique ofchoice in most labs that study bacteria or yeastsays Martin Smith sales manager at Synbiosis inCambridge UK which makes automated colonycounters In virology plaques are the best way toconfirm that viruses completed the full infectiouscycle says Racaniello who studies polioshy andrhinovirus in animal cell culture
However scientists faced with towers of hundredsor thousands of plates can get overwhelmed ifcounting by hand To help the latest automatedcounters have highshyres color cameras and multiplelighting options to identify spots barely visible tothe naked eye Advanced systems offer an array offeatures such as the ability to read barcode labelson plates calculate the number of colonyshyformingunits spread by a spiral plater or measure zonesof inhibition to name a few
Brugger on the other hand built his own colony counter for his current work studying the capsule ofpneumococcus (PLOS ONE 7e33695 2012) The DIY option may provide an affordable alternative tocostly commercial systems for mediumshythroughput counting
Here The Scientist profiles tools and tips for colony counting whether you have a few plates or a wholeincubator full
One If By Hand
At its most basic level counting colonies orplaques is as easy as 1 2 3 Many scientists usea marker like a Sharpie ($299 for two) to markeach spot on the plate as they do the tally so asnot to doubleshycount Racaniello keeps count in hishead but many researchers prefer to use ahandheld clicker ($13 or so) as they markSetting the plate on a light box ($100ndash$250)may make it easier to see the spots
ColeshyParmer of Vernon Hills Illinois sells a digital counter pen that senses the pressure on the dish witheach mark you make beeps and records the score ($21) However users complain the ink dries upquickly Similarly the SC6PLUS from Bibby Scientific Ltd in Staffordshire UK is a plate holder thatsenses your markerrsquos pressure It has a builtshyin magnifying glass and can also calculate an average tally($640) IUL Instruments SA based in Barcelona Spain makes a similar colony counter system ($900ndash
1 Comments 0Like 47 1
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SPOTTING DOTS Barcelonashybased IUL Instrumentsrsquo Flash amp Go (pictured on the right) identifies colonies andplaques as small as 006 mm It weighs about 10 pounds and comes with FDAshycompliant software Fort ValleyUniversityrsquos Ajit Mahapatra and his assistant Donna Harris (left) use the machine to count E coli colonies fortheir research on food and agricultural engineeringCOURTESY OF AJIT MAHAPATRA IUL SA
THE MANUAL METHOD Many researchers use amagnifying glass or a simple stereo microscope aclickershycounter and a Sharpie pen for a lowshytech buttimeshyintensive solutionPHOTO BY PASCAL M FREYCOURTESY OF SILVIOBRUGGER
$2200 from the US distributor NeushyTec GroupInc)
Throughput Counting by hand works fine forbasic researchers who donrsquot have too manyplates Brugger estimates it takes up to half a dayto go through 100 85shymillimeter petri dishes with30ndash300 bacterial colonies each For viruscounting Racaniello reports that 100 6shywellplates with 10ndash100 plaques per well take an houror two
Accuracy ldquoItrsquos as good as whoever is doing itrdquosays Elizabeth Murphy a product manager atColeshyParmer Racaniello suspects that differentresearchers would probably come within a fewplaques of each other By counting each platethree times he feels his results are reliable
Prosbull Economicalbull Easy
Consbull Tediousbull Open to human error both in counting andtranscribing data into a computerbull Spending hours hunched over plates full ofbacteria could cause eyestrain And breathing the air around such samplesmight not be healthy Murphy suggests
Tipsbull For plates crowded with more than 200colonies you can divide the area into sectorsmdash such as quarters or eighthsmdashcount one sectorand use that figure to estimate the total ldquoIt saves you a lot of time but itrsquos not very accuraterdquo Brugger cautionsbull For consistency one person should count all the plates says Ajit Mahapatra of Fort Valley State University in Georgia A second counter on the plate set can increase reliability
Automatic Counters
Scientists in Mahapatrarsquos lab now use an automatic colony counter These range from simple models tofancy machines with builtshyin analysis options and the ability to count both microbial colonies and viralplaques
Basic One straightforward affordable starter option is the aCOLyte 3 from Synbiosis ($418889 includingthe machine and software) It identifies colonies or plaques as small as 03 mm across and adds up thetotal for you IULrsquos Flash amp Go ($9000) which Mahapatra uses in his research on eliminating bacterialcontamination of foods offers similar basic counting capabilities These systems donrsquot have a lot ofadditional functions they just count spots
Advanced Other counters do much more than enumerate colonies and plaques For example Synbiosisoffers an advanced machine the Protocol 3 ($1098889 $14500 with a touchshyscreen computer) thatcan also provide data on the size or shape of your spots or measure zones of inhibition with the includedsoftware With additional optional software modules the ProtoCOL 3 can perform even more tasks For
example it can count and calculate concentrations of spiralshyplated colonies or analyze plates that use theAmes testmdashidentifying carcinogenic compounds by measuring the rate at which histidineshydependentbacteria mutate and acquire the ability to grow on histidineshyfree media ($94286 for each of theseadditional software plugins)
The ProtoCOL 3 with a scientificshygrade camera spots colonies as small as 43 micronsmdashldquojust barelyrdquo visibleto the eye says Smith Many other advanced counters can also identify dots in the subndash100shymicron range
Those who need more than just counts can also get an upgraded system from IUL called Flash amp Growstarting at $17000 which includes features such as color and size differentiation and other softwaremodules For example you can add zoneshyofshyinhibition measurements to test antibiotic drugs for about$4000
Most advanced systems use color cameras allowing users to distinguish colonies of different hues TheColonyDocshyIt Imaging Station from UVP LLC of Upland California ($6900) can also identify coloniesexpressing green fluorescent protein using an addshyon emission filter ($800) Both the ColonyDocshyIt andthe ProtoCOL3 have doors to shield plates from ambient light ensuring even illumination of the images
Throughput Automatic counters enable highshythroughput counting of hundreds or thousands of platesPharmaceutical companies as well as watershy and foodshysafety labs would likely find them useful says SmithWhen Mahapatra timed manual counting it took an average of 73 seconds per plate he says compared toa second or two with the Flash amp Go For those with truly highshythroughput needs IUL offers a companionplate handler for the Flash amp Grow which robotically feeds a stack of up to 80 plates into the imager($60000 including the imager and software)
Accuracy Itrsquos pretty good Mahapatra compared the Flash amp Go to counting by hand when he first got themachine and says the imager was quite accurate It came up with the same tally several times on thesame plate and correlated well with manual counts (Agricultural Engineering International the CIGREjournal 11MS1368 2009) He feels more confident of his results using the automatic system
Prosbull Fastbull No transcribing errors and therersquos a data trail of who performed the counting whenbull Builtshyin analysis for a variety of assays
Consbull Expensivebull Funnyshyshaped or closeshytogether colonies mold or debris writing on the plate or bubbles in the agar may interfere with an accurate count and require user correctionbull Since the software is proprietary you canrsquot customize it if the commercial version doesnrsquot fit your needs
Tipsbull Each lab should validate the instrument and its counts as compared to their usual method says Smithbull Keep the bottom of the plate free of writing and smudges Instead label the plate lid or side
DIY Solutions
Brugger wasnrsquot aware of commercial systemswhen he decided that crafting an automaticcounter would be a fun project for his lab Theteam built an aluminum apparatus with a 33megapixel camera mounted above a drawer thatholds the petri dish and a circular light sourcesurrounding the dish The Brugger group spentnearly $8000 but you could choose a cheapercamera or lighting he says To identify circularspots they wrote MATLAB software now freelyavailable online (wwwcoccuseu) The smallestspots Brugger has tried are Moraxella catarrhaliscolonies of 1ndash3 millimeters
Like Brugger Kevin Claytor wasnrsquot happy with the prospect of counting bacterial colonies by hand when asa Rice University student he spent the summer of 2008 in a Research Experience for Undergraduatesprogram at the University of Illinois in UrbanashyChampaign Playing with a webcam at home Claytor
DIY AUTOMATION The hardware setup of KevinClaytorrsquos apparatus is shown here (left) The webcamis clipped to the holder on the top of the box (middlepanel left) The sample sits on the diffuser in thecenter of the box and the lamp is placed so the bulbis below the sample (bottom panel)KEVIN CLAYTOR UIUC
thought why not use it for his experiments Hebuilt his apparatus out of a cardboard box 15shywatt light bulb and highshyres webcam for about$50
(physicsillinoiseduundergradreu2008Claytor_Kevinpdf) ldquoMost of the difficulty is the softwarerdquo saysClaytor now a physics graduate student at Duke University in Durham North Carolina He used MATLABto program the software for the counter which he has made available for free(peopledukeedu~kec30colonyhtml) While Claytor designed his system for bacterial colonies it shouldbe adaptable for viral plaques too he thinks The program provides spot count as well as statistics such asaverage size and shape symmetry
The DIY option is not necessarily limited to MATLAB aficionados Claytor notes because imageshyprocessingsoftware is freely available online For example Anne Carpenter at the Broad Institute in CambridgeMassachusetts codeveloped an imageshyanalysis program called CellProfiler (BioTechniques 4271shy752007) ldquoItrsquos designed for biologistsrdquo she says ldquoYou donrsquot need to adapt the code yourself you just playaround with different settingsrdquo With CellProfiler you could count spots or measure zones of inhibition andtrain the program to ignore artifacts such as bubbles You could use a digital camera or a flatbed scanner tomake the images for less than $100 she suggests
Throughput Homemade counters fill the space between smallshyscale figuring by hand and largeshyscalecommercial tallies Claytor says Bruggerrsquos lab might have 100 or 200 plates to count from time to timeHe estimates his system takes up to 20 seconds to count a dish depending on the number of coloniespresent so it could easily complete 100 plates within an hour Claytorrsquos took fewer than 10 seconds tocount a plate
Accuracy Bruggerrsquos system offers better accuracy than manual counting he found particularly for plateswith hundreds of colonies Claytor in his summer project reached an accuracy of 93 percent compared tothe byshyhand tally Some spotshyidentifying algorithms work better than others he notes
Prosbull Inexpensivebull Openshysource code means you can modify the program to fit your experiments
Consbull Limited featuresbull You may have to update the software yourself if you upgrade your computer or camera
Tips
copy 1986shy2013 The ScientistHome News amp Opinion The Nutshell Multimedia Magazine Advertise
About amp Contact Privacy Policy Job Listings Subscribe Archive
Now Part of the LabX Media Group Lab Manager Magazine | LabX | LabWrench
You
awPosts 1
Week in Review April 29ndash May 2By Jef Akst
The brainrsquos role in aging trackingdisease understanding the newflu virus no autismshyLyme linkone drugrsquos journey from bench tobedside
Easy Jump for H5N1from Bird to MammalBy Ruth Williams
Hybrid viruses derived from anH5N1 bird flu strain can infectguinea pigs through the air
BRET Meets FRETBy Ruth Williams
Scientists create biocompatibleselfshyluminescing nanoparticles forin vivo imaging
bull ldquoThe most important thing is to get the illumination of the plate rightrdquo Carpenter says You want even lightwith minimal shadows Brugger applied indirect ldquodarkshyfieldrdquo blue light from the sides of his apparatus to help distinguish colonies on dark bloodshycontaining agarbull Keep a consistent distance between the sample and the camera so the resolution is the same in all picturesbull ldquoYou should shoot for each colony or each lsquoblob of interestrsquo being at least five pixels across in the final imagerdquo Carpenter says ldquoAny less than that it gets hard to distinguish two little colonies side by siderdquo Ten pixels across would be even better she says
Tagsvirology techniques microbiology and colony counters
Add a Comment
Sign In with your LabX Media Group Passport to leave acomment
Not a member Register Now
Comments
May 2 2013
there is also a petty of nice appsweb available for counting colonies plus uhav an extra benefit of a photo backup
Sign in to Report
Related Articles
1 Comments 0Like 47 1 Link this Stumble Tweet this
SPOTTING DOTS Barcelonashybased IUL Instrumentsrsquo Flash amp Go (pictured on the right) identifies colonies andplaques as small as 006 mm It weighs about 10 pounds and comes with FDAshycompliant software Fort ValleyUniversityrsquos Ajit Mahapatra and his assistant Donna Harris (left) use the machine to count E coli colonies fortheir research on food and agricultural engineeringCOURTESY OF AJIT MAHAPATRA IUL SA
THE MANUAL METHOD Many researchers use amagnifying glass or a simple stereo microscope aclickershycounter and a Sharpie pen for a lowshytech buttimeshyintensive solutionPHOTO BY PASCAL M FREYCOURTESY OF SILVIOBRUGGER
$2200 from the US distributor NeushyTec GroupInc)
Throughput Counting by hand works fine forbasic researchers who donrsquot have too manyplates Brugger estimates it takes up to half a dayto go through 100 85shymillimeter petri dishes with30ndash300 bacterial colonies each For viruscounting Racaniello reports that 100 6shywellplates with 10ndash100 plaques per well take an houror two
Accuracy ldquoItrsquos as good as whoever is doing itrdquosays Elizabeth Murphy a product manager atColeshyParmer Racaniello suspects that differentresearchers would probably come within a fewplaques of each other By counting each platethree times he feels his results are reliable
Prosbull Economicalbull Easy
Consbull Tediousbull Open to human error both in counting andtranscribing data into a computerbull Spending hours hunched over plates full ofbacteria could cause eyestrain And breathing the air around such samplesmight not be healthy Murphy suggests
Tipsbull For plates crowded with more than 200colonies you can divide the area into sectorsmdash such as quarters or eighthsmdashcount one sectorand use that figure to estimate the total ldquoIt saves you a lot of time but itrsquos not very accuraterdquo Brugger cautionsbull For consistency one person should count all the plates says Ajit Mahapatra of Fort Valley State University in Georgia A second counter on the plate set can increase reliability
Automatic Counters
Scientists in Mahapatrarsquos lab now use an automatic colony counter These range from simple models tofancy machines with builtshyin analysis options and the ability to count both microbial colonies and viralplaques
Basic One straightforward affordable starter option is the aCOLyte 3 from Synbiosis ($418889 includingthe machine and software) It identifies colonies or plaques as small as 03 mm across and adds up thetotal for you IULrsquos Flash amp Go ($9000) which Mahapatra uses in his research on eliminating bacterialcontamination of foods offers similar basic counting capabilities These systems donrsquot have a lot ofadditional functions they just count spots
Advanced Other counters do much more than enumerate colonies and plaques For example Synbiosisoffers an advanced machine the Protocol 3 ($1098889 $14500 with a touchshyscreen computer) thatcan also provide data on the size or shape of your spots or measure zones of inhibition with the includedsoftware With additional optional software modules the ProtoCOL 3 can perform even more tasks For
example it can count and calculate concentrations of spiralshyplated colonies or analyze plates that use theAmes testmdashidentifying carcinogenic compounds by measuring the rate at which histidineshydependentbacteria mutate and acquire the ability to grow on histidineshyfree media ($94286 for each of theseadditional software plugins)
The ProtoCOL 3 with a scientificshygrade camera spots colonies as small as 43 micronsmdashldquojust barelyrdquo visibleto the eye says Smith Many other advanced counters can also identify dots in the subndash100shymicron range
Those who need more than just counts can also get an upgraded system from IUL called Flash amp Growstarting at $17000 which includes features such as color and size differentiation and other softwaremodules For example you can add zoneshyofshyinhibition measurements to test antibiotic drugs for about$4000
Most advanced systems use color cameras allowing users to distinguish colonies of different hues TheColonyDocshyIt Imaging Station from UVP LLC of Upland California ($6900) can also identify coloniesexpressing green fluorescent protein using an addshyon emission filter ($800) Both the ColonyDocshyIt andthe ProtoCOL3 have doors to shield plates from ambient light ensuring even illumination of the images
Throughput Automatic counters enable highshythroughput counting of hundreds or thousands of platesPharmaceutical companies as well as watershy and foodshysafety labs would likely find them useful says SmithWhen Mahapatra timed manual counting it took an average of 73 seconds per plate he says compared toa second or two with the Flash amp Go For those with truly highshythroughput needs IUL offers a companionplate handler for the Flash amp Grow which robotically feeds a stack of up to 80 plates into the imager($60000 including the imager and software)
Accuracy Itrsquos pretty good Mahapatra compared the Flash amp Go to counting by hand when he first got themachine and says the imager was quite accurate It came up with the same tally several times on thesame plate and correlated well with manual counts (Agricultural Engineering International the CIGREjournal 11MS1368 2009) He feels more confident of his results using the automatic system
Prosbull Fastbull No transcribing errors and therersquos a data trail of who performed the counting whenbull Builtshyin analysis for a variety of assays
Consbull Expensivebull Funnyshyshaped or closeshytogether colonies mold or debris writing on the plate or bubbles in the agar may interfere with an accurate count and require user correctionbull Since the software is proprietary you canrsquot customize it if the commercial version doesnrsquot fit your needs
Tipsbull Each lab should validate the instrument and its counts as compared to their usual method says Smithbull Keep the bottom of the plate free of writing and smudges Instead label the plate lid or side
DIY Solutions
Brugger wasnrsquot aware of commercial systemswhen he decided that crafting an automaticcounter would be a fun project for his lab Theteam built an aluminum apparatus with a 33megapixel camera mounted above a drawer thatholds the petri dish and a circular light sourcesurrounding the dish The Brugger group spentnearly $8000 but you could choose a cheapercamera or lighting he says To identify circularspots they wrote MATLAB software now freelyavailable online (wwwcoccuseu) The smallestspots Brugger has tried are Moraxella catarrhaliscolonies of 1ndash3 millimeters
Like Brugger Kevin Claytor wasnrsquot happy with the prospect of counting bacterial colonies by hand when asa Rice University student he spent the summer of 2008 in a Research Experience for Undergraduatesprogram at the University of Illinois in UrbanashyChampaign Playing with a webcam at home Claytor
DIY AUTOMATION The hardware setup of KevinClaytorrsquos apparatus is shown here (left) The webcamis clipped to the holder on the top of the box (middlepanel left) The sample sits on the diffuser in thecenter of the box and the lamp is placed so the bulbis below the sample (bottom panel)KEVIN CLAYTOR UIUC
thought why not use it for his experiments Hebuilt his apparatus out of a cardboard box 15shywatt light bulb and highshyres webcam for about$50
(physicsillinoiseduundergradreu2008Claytor_Kevinpdf) ldquoMost of the difficulty is the softwarerdquo saysClaytor now a physics graduate student at Duke University in Durham North Carolina He used MATLABto program the software for the counter which he has made available for free(peopledukeedu~kec30colonyhtml) While Claytor designed his system for bacterial colonies it shouldbe adaptable for viral plaques too he thinks The program provides spot count as well as statistics such asaverage size and shape symmetry
The DIY option is not necessarily limited to MATLAB aficionados Claytor notes because imageshyprocessingsoftware is freely available online For example Anne Carpenter at the Broad Institute in CambridgeMassachusetts codeveloped an imageshyanalysis program called CellProfiler (BioTechniques 4271shy752007) ldquoItrsquos designed for biologistsrdquo she says ldquoYou donrsquot need to adapt the code yourself you just playaround with different settingsrdquo With CellProfiler you could count spots or measure zones of inhibition andtrain the program to ignore artifacts such as bubbles You could use a digital camera or a flatbed scanner tomake the images for less than $100 she suggests
Throughput Homemade counters fill the space between smallshyscale figuring by hand and largeshyscalecommercial tallies Claytor says Bruggerrsquos lab might have 100 or 200 plates to count from time to timeHe estimates his system takes up to 20 seconds to count a dish depending on the number of coloniespresent so it could easily complete 100 plates within an hour Claytorrsquos took fewer than 10 seconds tocount a plate
Accuracy Bruggerrsquos system offers better accuracy than manual counting he found particularly for plateswith hundreds of colonies Claytor in his summer project reached an accuracy of 93 percent compared tothe byshyhand tally Some spotshyidentifying algorithms work better than others he notes
Prosbull Inexpensivebull Openshysource code means you can modify the program to fit your experiments
Consbull Limited featuresbull You may have to update the software yourself if you upgrade your computer or camera
Tips
copy 1986shy2013 The ScientistHome News amp Opinion The Nutshell Multimedia Magazine Advertise
About amp Contact Privacy Policy Job Listings Subscribe Archive
Now Part of the LabX Media Group Lab Manager Magazine | LabX | LabWrench
You
awPosts 1
Week in Review April 29ndash May 2By Jef Akst
The brainrsquos role in aging trackingdisease understanding the newflu virus no autismshyLyme linkone drugrsquos journey from bench tobedside
Easy Jump for H5N1from Bird to MammalBy Ruth Williams
Hybrid viruses derived from anH5N1 bird flu strain can infectguinea pigs through the air
BRET Meets FRETBy Ruth Williams
Scientists create biocompatibleselfshyluminescing nanoparticles forin vivo imaging
bull ldquoThe most important thing is to get the illumination of the plate rightrdquo Carpenter says You want even lightwith minimal shadows Brugger applied indirect ldquodarkshyfieldrdquo blue light from the sides of his apparatus to help distinguish colonies on dark bloodshycontaining agarbull Keep a consistent distance between the sample and the camera so the resolution is the same in all picturesbull ldquoYou should shoot for each colony or each lsquoblob of interestrsquo being at least five pixels across in the final imagerdquo Carpenter says ldquoAny less than that it gets hard to distinguish two little colonies side by siderdquo Ten pixels across would be even better she says
Tagsvirology techniques microbiology and colony counters
Add a Comment
Sign In with your LabX Media Group Passport to leave acomment
Not a member Register Now
Comments
May 2 2013
there is also a petty of nice appsweb available for counting colonies plus uhav an extra benefit of a photo backup
Sign in to Report
Related Articles
1 Comments 0Like 47 1 Link this Stumble Tweet this
example it can count and calculate concentrations of spiralshyplated colonies or analyze plates that use theAmes testmdashidentifying carcinogenic compounds by measuring the rate at which histidineshydependentbacteria mutate and acquire the ability to grow on histidineshyfree media ($94286 for each of theseadditional software plugins)
The ProtoCOL 3 with a scientificshygrade camera spots colonies as small as 43 micronsmdashldquojust barelyrdquo visibleto the eye says Smith Many other advanced counters can also identify dots in the subndash100shymicron range
Those who need more than just counts can also get an upgraded system from IUL called Flash amp Growstarting at $17000 which includes features such as color and size differentiation and other softwaremodules For example you can add zoneshyofshyinhibition measurements to test antibiotic drugs for about$4000
Most advanced systems use color cameras allowing users to distinguish colonies of different hues TheColonyDocshyIt Imaging Station from UVP LLC of Upland California ($6900) can also identify coloniesexpressing green fluorescent protein using an addshyon emission filter ($800) Both the ColonyDocshyIt andthe ProtoCOL3 have doors to shield plates from ambient light ensuring even illumination of the images
Throughput Automatic counters enable highshythroughput counting of hundreds or thousands of platesPharmaceutical companies as well as watershy and foodshysafety labs would likely find them useful says SmithWhen Mahapatra timed manual counting it took an average of 73 seconds per plate he says compared toa second or two with the Flash amp Go For those with truly highshythroughput needs IUL offers a companionplate handler for the Flash amp Grow which robotically feeds a stack of up to 80 plates into the imager($60000 including the imager and software)
Accuracy Itrsquos pretty good Mahapatra compared the Flash amp Go to counting by hand when he first got themachine and says the imager was quite accurate It came up with the same tally several times on thesame plate and correlated well with manual counts (Agricultural Engineering International the CIGREjournal 11MS1368 2009) He feels more confident of his results using the automatic system
Prosbull Fastbull No transcribing errors and therersquos a data trail of who performed the counting whenbull Builtshyin analysis for a variety of assays
Consbull Expensivebull Funnyshyshaped or closeshytogether colonies mold or debris writing on the plate or bubbles in the agar may interfere with an accurate count and require user correctionbull Since the software is proprietary you canrsquot customize it if the commercial version doesnrsquot fit your needs
Tipsbull Each lab should validate the instrument and its counts as compared to their usual method says Smithbull Keep the bottom of the plate free of writing and smudges Instead label the plate lid or side
DIY Solutions
Brugger wasnrsquot aware of commercial systemswhen he decided that crafting an automaticcounter would be a fun project for his lab Theteam built an aluminum apparatus with a 33megapixel camera mounted above a drawer thatholds the petri dish and a circular light sourcesurrounding the dish The Brugger group spentnearly $8000 but you could choose a cheapercamera or lighting he says To identify circularspots they wrote MATLAB software now freelyavailable online (wwwcoccuseu) The smallestspots Brugger has tried are Moraxella catarrhaliscolonies of 1ndash3 millimeters
Like Brugger Kevin Claytor wasnrsquot happy with the prospect of counting bacterial colonies by hand when asa Rice University student he spent the summer of 2008 in a Research Experience for Undergraduatesprogram at the University of Illinois in UrbanashyChampaign Playing with a webcam at home Claytor
DIY AUTOMATION The hardware setup of KevinClaytorrsquos apparatus is shown here (left) The webcamis clipped to the holder on the top of the box (middlepanel left) The sample sits on the diffuser in thecenter of the box and the lamp is placed so the bulbis below the sample (bottom panel)KEVIN CLAYTOR UIUC
thought why not use it for his experiments Hebuilt his apparatus out of a cardboard box 15shywatt light bulb and highshyres webcam for about$50
(physicsillinoiseduundergradreu2008Claytor_Kevinpdf) ldquoMost of the difficulty is the softwarerdquo saysClaytor now a physics graduate student at Duke University in Durham North Carolina He used MATLABto program the software for the counter which he has made available for free(peopledukeedu~kec30colonyhtml) While Claytor designed his system for bacterial colonies it shouldbe adaptable for viral plaques too he thinks The program provides spot count as well as statistics such asaverage size and shape symmetry
The DIY option is not necessarily limited to MATLAB aficionados Claytor notes because imageshyprocessingsoftware is freely available online For example Anne Carpenter at the Broad Institute in CambridgeMassachusetts codeveloped an imageshyanalysis program called CellProfiler (BioTechniques 4271shy752007) ldquoItrsquos designed for biologistsrdquo she says ldquoYou donrsquot need to adapt the code yourself you just playaround with different settingsrdquo With CellProfiler you could count spots or measure zones of inhibition andtrain the program to ignore artifacts such as bubbles You could use a digital camera or a flatbed scanner tomake the images for less than $100 she suggests
Throughput Homemade counters fill the space between smallshyscale figuring by hand and largeshyscalecommercial tallies Claytor says Bruggerrsquos lab might have 100 or 200 plates to count from time to timeHe estimates his system takes up to 20 seconds to count a dish depending on the number of coloniespresent so it could easily complete 100 plates within an hour Claytorrsquos took fewer than 10 seconds tocount a plate
Accuracy Bruggerrsquos system offers better accuracy than manual counting he found particularly for plateswith hundreds of colonies Claytor in his summer project reached an accuracy of 93 percent compared tothe byshyhand tally Some spotshyidentifying algorithms work better than others he notes
Prosbull Inexpensivebull Openshysource code means you can modify the program to fit your experiments
Consbull Limited featuresbull You may have to update the software yourself if you upgrade your computer or camera
Tips
copy 1986shy2013 The ScientistHome News amp Opinion The Nutshell Multimedia Magazine Advertise
About amp Contact Privacy Policy Job Listings Subscribe Archive
Now Part of the LabX Media Group Lab Manager Magazine | LabX | LabWrench
You
awPosts 1
Week in Review April 29ndash May 2By Jef Akst
The brainrsquos role in aging trackingdisease understanding the newflu virus no autismshyLyme linkone drugrsquos journey from bench tobedside
Easy Jump for H5N1from Bird to MammalBy Ruth Williams
Hybrid viruses derived from anH5N1 bird flu strain can infectguinea pigs through the air
BRET Meets FRETBy Ruth Williams
Scientists create biocompatibleselfshyluminescing nanoparticles forin vivo imaging
bull ldquoThe most important thing is to get the illumination of the plate rightrdquo Carpenter says You want even lightwith minimal shadows Brugger applied indirect ldquodarkshyfieldrdquo blue light from the sides of his apparatus to help distinguish colonies on dark bloodshycontaining agarbull Keep a consistent distance between the sample and the camera so the resolution is the same in all picturesbull ldquoYou should shoot for each colony or each lsquoblob of interestrsquo being at least five pixels across in the final imagerdquo Carpenter says ldquoAny less than that it gets hard to distinguish two little colonies side by siderdquo Ten pixels across would be even better she says
Tagsvirology techniques microbiology and colony counters
Add a Comment
Sign In with your LabX Media Group Passport to leave acomment
Not a member Register Now
Comments
May 2 2013
there is also a petty of nice appsweb available for counting colonies plus uhav an extra benefit of a photo backup
Sign in to Report
Related Articles
1 Comments 0Like 47 1 Link this Stumble Tweet this
DIY AUTOMATION The hardware setup of KevinClaytorrsquos apparatus is shown here (left) The webcamis clipped to the holder on the top of the box (middlepanel left) The sample sits on the diffuser in thecenter of the box and the lamp is placed so the bulbis below the sample (bottom panel)KEVIN CLAYTOR UIUC
thought why not use it for his experiments Hebuilt his apparatus out of a cardboard box 15shywatt light bulb and highshyres webcam for about$50
(physicsillinoiseduundergradreu2008Claytor_Kevinpdf) ldquoMost of the difficulty is the softwarerdquo saysClaytor now a physics graduate student at Duke University in Durham North Carolina He used MATLABto program the software for the counter which he has made available for free(peopledukeedu~kec30colonyhtml) While Claytor designed his system for bacterial colonies it shouldbe adaptable for viral plaques too he thinks The program provides spot count as well as statistics such asaverage size and shape symmetry
The DIY option is not necessarily limited to MATLAB aficionados Claytor notes because imageshyprocessingsoftware is freely available online For example Anne Carpenter at the Broad Institute in CambridgeMassachusetts codeveloped an imageshyanalysis program called CellProfiler (BioTechniques 4271shy752007) ldquoItrsquos designed for biologistsrdquo she says ldquoYou donrsquot need to adapt the code yourself you just playaround with different settingsrdquo With CellProfiler you could count spots or measure zones of inhibition andtrain the program to ignore artifacts such as bubbles You could use a digital camera or a flatbed scanner tomake the images for less than $100 she suggests
Throughput Homemade counters fill the space between smallshyscale figuring by hand and largeshyscalecommercial tallies Claytor says Bruggerrsquos lab might have 100 or 200 plates to count from time to timeHe estimates his system takes up to 20 seconds to count a dish depending on the number of coloniespresent so it could easily complete 100 plates within an hour Claytorrsquos took fewer than 10 seconds tocount a plate
Accuracy Bruggerrsquos system offers better accuracy than manual counting he found particularly for plateswith hundreds of colonies Claytor in his summer project reached an accuracy of 93 percent compared tothe byshyhand tally Some spotshyidentifying algorithms work better than others he notes
Prosbull Inexpensivebull Openshysource code means you can modify the program to fit your experiments
Consbull Limited featuresbull You may have to update the software yourself if you upgrade your computer or camera
Tips
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Now Part of the LabX Media Group Lab Manager Magazine | LabX | LabWrench
You
awPosts 1
Week in Review April 29ndash May 2By Jef Akst
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Easy Jump for H5N1from Bird to MammalBy Ruth Williams
Hybrid viruses derived from anH5N1 bird flu strain can infectguinea pigs through the air
BRET Meets FRETBy Ruth Williams
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bull ldquoThe most important thing is to get the illumination of the plate rightrdquo Carpenter says You want even lightwith minimal shadows Brugger applied indirect ldquodarkshyfieldrdquo blue light from the sides of his apparatus to help distinguish colonies on dark bloodshycontaining agarbull Keep a consistent distance between the sample and the camera so the resolution is the same in all picturesbull ldquoYou should shoot for each colony or each lsquoblob of interestrsquo being at least five pixels across in the final imagerdquo Carpenter says ldquoAny less than that it gets hard to distinguish two little colonies side by siderdquo Ten pixels across would be even better she says
Tagsvirology techniques microbiology and colony counters
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Comments
May 2 2013
there is also a petty of nice appsweb available for counting colonies plus uhav an extra benefit of a photo backup
Sign in to Report
Related Articles
1 Comments 0Like 47 1 Link this Stumble Tweet this
copy 1986shy2013 The ScientistHome News amp Opinion The Nutshell Multimedia Magazine Advertise
About amp Contact Privacy Policy Job Listings Subscribe Archive
Now Part of the LabX Media Group Lab Manager Magazine | LabX | LabWrench
You
awPosts 1
Week in Review April 29ndash May 2By Jef Akst
The brainrsquos role in aging trackingdisease understanding the newflu virus no autismshyLyme linkone drugrsquos journey from bench tobedside
Easy Jump for H5N1from Bird to MammalBy Ruth Williams
Hybrid viruses derived from anH5N1 bird flu strain can infectguinea pigs through the air
BRET Meets FRETBy Ruth Williams
Scientists create biocompatibleselfshyluminescing nanoparticles forin vivo imaging
bull ldquoThe most important thing is to get the illumination of the plate rightrdquo Carpenter says You want even lightwith minimal shadows Brugger applied indirect ldquodarkshyfieldrdquo blue light from the sides of his apparatus to help distinguish colonies on dark bloodshycontaining agarbull Keep a consistent distance between the sample and the camera so the resolution is the same in all picturesbull ldquoYou should shoot for each colony or each lsquoblob of interestrsquo being at least five pixels across in the final imagerdquo Carpenter says ldquoAny less than that it gets hard to distinguish two little colonies side by siderdquo Ten pixels across would be even better she says
Tagsvirology techniques microbiology and colony counters
Add a Comment
Sign In with your LabX Media Group Passport to leave acomment
Not a member Register Now
Comments
May 2 2013
there is also a petty of nice appsweb available for counting colonies plus uhav an extra benefit of a photo backup
Sign in to Report
Related Articles
1 Comments 0Like 47 1 Link this Stumble Tweet this