HCS & in vivo imaging system
Locked
 
|
Nishigandha Naik |
HCS & in vivo imaging system
|
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Dear All, I am planning to procure High Content Screening system (HCS). I would like to know experience of users as I could not find many published reports on different systems. I am also debating on utility of confocal (Nipkow disk based) based HCS versus without confocal HCS. I will be thankful if anybody could share their experience or opine on the various available makes in the market. I am also looking for in vivo imaging system to procure images from small animals such as mouse, rats to study tumor formation, metastasis and angeogenesis of GFP or RFP expressing. I request to share your experience to enable me to choose the proper system for these applications. With thanks and best wishes for the year, Nishigandha Naik ____________________________________________________________________________________ Looking for last minute shopping deals? Find them fast with Yahoo! Search. http://tools.search.yahoo.com/newsearch/category.php?category=shopping |
|
|
George McNamara |
Re: HCS & in vivo imaging system
|
|
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Hi Nishigandha, As always - get a demo! Really more a question of budget and throughput. For in vivo, the Caliper/Xenogen IVIS Spectrum looks very good. http://www.caliperls.com/products/optical-imaging/ivis-spectrum.htm My colleagues at UM are upgrading our IVIS 200 to the Spectrum in a month. Advantage of the IVIS systems is that using cryogenic back-illuminated CCD's enables luciferase bioluminescence imaging (BLI) in vivo (and ex vivo - extremely valuable to take out the organs at the end and re-image without the rest of the animal in the way. also, can image multi-well plates, so can test the performance of cell lines before putting them in the mouse). A huge advantage of the Xenogen systems is that they have made the effort to provide quantitative data for bioluminescence. They use photons/second/cm^2/steradian. This enables every paper published with a Xenogen system to be compared to each other (of course, the depth of the cells, expression level, and amount of luciferin at the cells are additional variables). If you get a non-Xenogen BLI instrument, make sure the vendor validates the same units. Xenogen now does something similar for fluorescence tomography (a trickier issue). CRi has been in the in vivo fluorescence field for several years. Check out their http://www.cri-inc.com/products/maestro2.asp If your budget is on the low end, arrange a demo from John Fox of Lightools (http://www.lightools.com/tutorial.htm). I was especially impressed by a demo of his a couple of years ago, that included the Pan-a-See-Ya (http://www.lightools.com/LRTPDFS/panaseeypdf.pdf), a simple RGB color camera, and mouse in hand without anesthesia (you need an isofluorene rig for the other systems). Reagents, cells and animals for in vivo imaging: Marker Gene Technology and Promega both sell gal-luciferin, a LacZ substrate whose product is a firefly luciferase substrate. Sam Gambhir has lots of tribrid vectors of FP-luciferase-TK (TK = enzyme for use with PET reporter and therapy). Xenogen also has several transgenic mice and various cell lines expressing luciferase. GFP (and maybe by now RFP) and firefly luciferase transgenic mice are available from Jackson Labs (www.jax.org). Lots of plasmids are available from www.addgene.com Which FP works best is tissue dependent - see http://www.jhc.org/cgi/content/abstract/55/9/931 for one study. You can also demo by injecting known volumes of fluorescent beads (i.e. in matrigel) under the skin, though you should expect most of the research to be of deeper sites. Ralph Weissleder's company, Visen Medical, sells a variety of fluorescence reagents for mouse in vivo imaging (AngioSense 680, etc). Fluorescence angiography with indocyanine green (ICG) or fluorescein (http://msp.rmit.edu.au/Article_02/03d.html) has been clinically available for years, and is a good way to start with mice. HCS - Cellomics (Thermo Fisher) founded the field, worth demoing. They had a nice article on their web site on how adding an Apotome (Zeiss optical sectioning device) to their instrument improved Z'-scores, so confocal imaging should help. Drug Discovery Today, ASSAY and Drug Development Technologies, Nature Reviews Drug Discovery, and a bunch of other journals cover this field. Start with the following books: Methods Enzymol. 2006;414 Methods Mol Biol. 2007;356 Then, search PubMed for high content screen* George At 12:56 AM 1/12/2008, you wrote: >Search the CONFOCAL archive at >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > >Dear All, > >I am planning to procure High Content Screening system >(HCS). I would like to know experience of users as I >could not find many published reports on different >systems. I am also debating on utility of confocal >(Nipkow disk based) based HCS versus without confocal >HCS. I will be thankful if anybody could share their >experience or opine on the various available makes in >the market. > >I am also looking for in vivo imaging system to >procure images from small animals such as mouse, rats >to study tumor formation, metastasis and angeogenesis >of GFP or RFP expressing. I request to share your >experience to enable me to choose the proper system >for these applications. > >With thanks and best wishes for the year, > >Nishigandha Naik > > > > > >____________________________________________________________________________________ >Looking for last minute shopping deals? >Find them fast with Yahoo! >Search. http://tools.search.yahoo.com/newsearch/category.php?category=shopping George McNamara, Ph.D. University of Miami, Miller School of Medicine Image Core Miami, FL 33010 [hidden email] [hidden email] 305-243-8436 office http://home.earthlink.net/~pubspectra/ http://home.earthlink.net/~geomcnamara/ http://www.sylvester.org/health_pro/shared_resources/index.asp (see Analytical Imaging Core Facility) |
|
|
Mancini, Michael A |
Re: HCS & in vivo imaging system
|
|
In reply to this post by Nishigandha Naik
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
NIshigandha, I can't help with the animal imaging, but we've evaluated a lot of HTM scopes and software over the last several years for HCS. I list below a few questions regarding the project needs of your group that should help narrow the scopes/software to go after. 1. Not surprisingly, what is the budget? There are scopes from not much over 100K to about a million or more..... 2. What are the assays your group wants to run? Will they run effectively on low mag/low NA lenses, or do they require higher mag/NA lenses? Many manufacturers cater specifically to the larger market of lower mag/lower NA hardware since that's all some assays need. Fewer scopes will be able to autofocus on 40x/0.95NA or 60-100x water/oil lenses where, in my opinion, the most data can be obtained for any number of assays (albeit at a slower rate) . 3. how fast do you need to go? how big will your experiments be? If you're after a genome wide RNAi screen or a large compound library vs a focused compound/RNAi libraries, the speed requirements are obviously very different. 4. do you have a bunch of programers that will be in charge of custom development of assays, or do you need "canned" assays that anyone can push a button to run? it seems the earlier 'locked-down' software is shifting to being increasingly fully customizable, perhaps due to the increased academic market. 5. what is the processing speed of the software that comes with the scope and/or the 3rd party software for image analyses? I can't argue enough for being able to fully customize protocols, and the need for fast image processing (unless you have a large cluster to crunch data). 6. make sure you plan for a means to catch all the image data, and how you're going to store it short/long term. tape backups fail (i'm told) 10% of the time. hard drives WILL fail. 7. i haven't seen a commercial database for image analysis, yet, that comes close to handling all the info from LIMS data to image data to compound/RNAi management to image analysis, but there again it is a question of SCALE. if, for example, you're getting an HTM to run in a small lab or small departmental core, many of these questions would elicit different answers... Note to vendors: I invite vendors to write me on or off line to share specifications for HTMs that are validated to focus at 40x/0.95 and above, as we are considering new scope purchases for 2008. Also, vendors with full blown LIMS+HTM image data management + querying+stats+reporting are encouraged to provide information. Cheers, Mike Michael A. Mancini, Ph.D. Associate Professor Director, Integrated Microscopy Core Co-Director, Gulf Coast Consortium for Chemical Genomics Department of Molecular and Cellular Biology Baylor College of Medicine Houston, TX 77030 713 798 8952 voice 713 798 3017 fax 713 408 0179 cell On Jan 11, 2008, at 11:56 PM, Nishigandha Naik wrote:
|
|
|
Nishigandha Naik |
Re: HCS & in vivo imaging system
|
|
In reply to this post by George McNamara
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Dear George, Thanks indeed for elaborate information. Among high end in vivo imaging system, on papers Xenogen did appear very good, especially for deep seated bioluminescence, but I could not find reports on imaging of GFP or RFP expressing metastatic cells. I am interested on imaging metastasis of GFP or RFP expressing cells in liver and lymphnodes. What is your opinion and experience reagrding this application with Xenogen system? Among lower end systems, FluorVivo of INDECO BioSystems and Pan-a-See-Ya of Lightools looked good. But, I could not get demo of Pan-a See-Ya from John Fox. Do you know any users who have sucessfuly used any of these for imaging metastasis in lymphnodes and liver? Do you have any infromation on performance HCS system of BD that has in built Nipkow disc based confocal system? Once again thanks a lot for the information. With best regards, Nishigandha Naik --- George McNamara <[hidden email]> wrote: > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > Hi Nishigandha, > > As always - get a demo! Really more a question of > budget and throughput. > > For in vivo, the Caliper/Xenogen IVIS Spectrum looks > very good. > http://www.caliperls.com/products/optical-imaging/ivis-spectrum.htm > > My colleagues at UM are upgrading our IVIS 200 to > the Spectrum in a > month. Advantage of the IVIS systems is that using > cryogenic > back-illuminated CCD's enables luciferase > bioluminescence imaging > (BLI) in vivo (and ex vivo - extremely valuable to > take out the > organs at the end and re-image without the rest of > the animal in the > way. also, can image multi-well plates, so can test > the performance > of cell lines before putting them in the mouse). > A huge advantage of the Xenogen systems is that they > have made the > effort to provide quantitative data for > bioluminescence. They use > photons/second/cm^2/steradian. This enables every > paper published > with a Xenogen system to be compared to each other > (of course, the > depth of the cells, expression level, and amount of > luciferin at the > cells are additional variables). If you get a > non-Xenogen BLI > instrument, make sure the vendor validates the same > units. Xenogen > now does something similar for fluorescence > tomography (a trickier issue). > > CRi has been in the in vivo fluorescence field for > several years. > Check out their > http://www.cri-inc.com/products/maestro2.asp > > If your budget is on the low end, arrange a demo > from John Fox of > Lightools (http://www.lightools.com/tutorial.htm). I > was especially > impressed by a demo of his a couple of years ago, > that included the > Pan-a-See-Ya > (http://www.lightools.com/LRTPDFS/panaseeypdf.pdf), > a > simple RGB color camera, and mouse in hand without > anesthesia (you > need an isofluorene rig for the other systems). > > Reagents, cells and animals for in vivo imaging: > > Marker Gene Technology and Promega both sell > gal-luciferin, a LacZ > substrate whose product is a firefly luciferase > substrate. Sam > Gambhir has lots of tribrid vectors of > FP-luciferase-TK (TK = enzyme > for use with PET reporter and therapy). > Xenogen also has several transgenic mice and various > cell lines > expressing luciferase. GFP (and maybe by now RFP) > and firefly > luciferase transgenic mice are available from > Jackson Labs > (www.jax.org). Lots of plasmids are available from > www.addgene.com > Which FP works best is tissue dependent - see > http://www.jhc.org/cgi/content/abstract/55/9/931 for > one study. > You can also demo by injecting known volumes of > fluorescent beads > (i.e. in matrigel) under the skin, though you should > expect most of > the research to be of deeper sites. Ralph > Weissleder's company, Visen > Medical, sells a variety of fluorescence reagents > for mouse in vivo > imaging (AngioSense 680, etc). Fluorescence > angiography with > indocyanine green (ICG) or fluorescein > (http://msp.rmit.edu.au/Article_02/03d.html) has > been clinically > available for years, and is a good way to start with > mice. > > > > HCS - Cellomics (Thermo Fisher) founded the field, > worth demoing. > They had a nice article on their web site on how > adding an Apotome > (Zeiss optical sectioning device) to their > instrument improved > Z'-scores, so confocal imaging should help. Drug > Discovery Today, > ASSAY and Drug Development Technologies, Nature > Reviews Drug > Discovery, and a bunch of other journals cover this > field. Start with > the following books: > Methods Enzymol. 2006;414 > Methods Mol Biol. 2007;356 > Then, search PubMed for high content screen* > > > George > > > At 12:56 AM 1/12/2008, you wrote: > >Search the CONFOCAL archive at > >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > > >Dear All, > > > >I am planning to procure High Content Screening > system > >(HCS). I would like to know experience of users as > I > >could not find many published reports on different > >systems. I am also debating on utility of confocal > >(Nipkow disk based) based HCS versus without > confocal > >HCS. I will be thankful if anybody could share > their > >experience or opine on the various available makes > in > >the market. > > > >I am also looking for in vivo imaging system to > >procure images from small animals such as mouse, > rats > >to study tumor formation, metastasis and > angeogenesis > >of GFP or RFP expressing. I request to share your > >experience to enable me to choose the proper system > >for these applications. > > > >With thanks and best wishes for the year, > > > >Nishigandha Naik > > > > > > > > > > > >____________________________________________________________________________________ > >Looking for last minute shopping deals? > >Find them fast with Yahoo! > >Search. > > > > > > > > George McNamara, Ph.D. > University of Miami, Miller School of Medicine > Image Core > Miami, FL 33010 > [hidden email] > [hidden email] > 305-243-8436 office > http://home.earthlink.net/~pubspectra/ > http://home.earthlink.net/~geomcnamara/ > > (see > Analytical Imaging Core Facility) > ____________________________________________________________________________________ Never miss a thing. Make Yahoo your home page. http://www.yahoo.com/r/hs |
|
|
George McNamara |
Re: HCS & in vivo imaging system
|
|
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
Hi Nishigandha, Most papers imaging GFP or RFP in liver and other internal organs have been published by Robert M. Hoffman of AntiCancer Inc. http://www.anticancer.com/ (ex PubMed search: hoffman rm liver orthotopic). He typically uses skin flaps in his in vivo imaging models. See Bouvet M, Tsuji K, Yang M, Jiang P, Moossa AR, Hoffman RM. Cancer Res 66: 11293, for an example. He has published with the Olympus products and claims to have been involved in developing the FluorVivo ( http://www.anticancer.com/FluorVivoBrochure.pdf). I have not yet seen the Xenogen system in action for in vivo fluorescence. If you want sensitivity by in vivo whole (intact) mouse imaging, go with Xenogen and firefly luciferase. If you want to be cutting edge (maybe bleeding edge), be the first to publish on a FLuc-mKate BRET model. If you want to image single cells in lymph nodes in live mice at microscopy resolution, look at the Olympus IV100 with stick objective lens(es) (hopefully in multiphoton mode), or get yourself a multiphoton microscope. Not tumor papers, but see the reviews by Hauser, Shlomchik and Haberman 2007 Nat Rev Immunol 7: 499-504, and by Cahalan MD, Parker I. Annu Rev Immunol. 2008 Jan 2; [Epub] for starters. Multiphoton excitation will enable a lot deeper imaging into lymph node (or liver, if you can get the lens to it) than confocal - though mKate expressing or DiD, Cy5 or Cy5.5 labeled cells confocal imaging might not be too bad (long wavelength light scatters less). If the Pan-a-See-Ya fits your requirements (i.e. imaging mice at macroscopic scale), and John Fox continues to decline to do a demo, ask if you can buy it on contingency - that is, if it works, you pay for it, if it does not, you ship it back (negotiate in advance who should be for shipping and whether a restocking fee is in order - after all, nce you've used it, it cannot be sold as new). BD CARV HCS system (Pathway Bioimager) - I've seen the one in our building used for Fura-2 ratio calcium imaging. Looked ok. I just saw it in action for live pancreatic islet cell imaging, did not touch any knobs so can't say how sensitive it is compared to other systems (and did not see any focus changes, so cannot address how confocal it is). It was being run for sequential well timelapse assays, not HCS, so I cannot address how well it works for HCS. best wishes, George At 04:29 AM 1/20/2008, you wrote: Search the CONFOCAL archive at George McNamara, Ph.D. University of Miami, Miller School of Medicine Image Core Miami, FL 33010 [hidden email] [hidden email] 305-243-8436 office http://home.earthlink.net/~pubspectra/ http://home.earthlink.net/~geomcnamara/ http://www.sylvester.org/health_pro/shared_resources/index.asp (see Analytical Imaging Core Facility) |
|
|
Nishigandha Naik |
Re: HCS & in vivo imaging system
|
|
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Thanks a lot George. This is to acknowledge the receipt of mail. I will go through the suggested links and write back. Need to keep up with some dead-lines. Thanks. Best regards, Nishigandha Naik --- George McNamara <[hidden email]> wrote: > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > > > Hi Nishigandha, > > Most papers imaging GFP or RFP in liver and other > internal organs > have been published by Robert M. Hoffman of > AntiCancer Inc. > http://www.anticancer.com/ (ex PubMed search: > hoffman rm liver > orthotopic). He typically uses skin flaps in his in > vivo imaging > models. See > > > M, Tsuji K, Yang M, Jiang P, Moossa AR, Hoffman RM. > Cancer Res 66: > 11293, for an example. He has published with the > Olympus products and > claims to have been involved in developing the > FluorVivo > (http://www.anticancer.com/FluorVivoBrochure.pdf). > > I have not yet seen the Xenogen system in action for > in vivo > fluorescence. If you want sensitivity by in vivo > whole (intact) mouse > imaging, go with Xenogen and firefly luciferase. If > you want to be > cutting edge (maybe bleeding edge), be the first to > publish on a > FLuc-mKate BRET model. > > If you want to image single cells in lymph nodes in > live mice at > microscopy resolution, look at the Olympus IV100 > with stick objective > lens(es) (hopefully in multiphoton mode), or get > yourself a > multiphoton microscope. Not tumor papers, but see > the reviews by > Hauser, Shlomchik and Haberman 2007 Nat Rev Immunol > 7: 499-504, and > by > > > MD, Parker I. Annu Rev Immunol. 2008 Jan 2; [Epub] > for starters. > Multiphoton excitation will enable a lot deeper > imaging into lymph > node (or liver, if you can get the lens to it) than > confocal - though > mKate expressing or DiD, Cy5 or Cy5.5 labeled cells > confocal imaging > might not be too bad (long wavelength light scatters > less). > > If the Pan-a-See-Ya fits your requirements (i.e. > imaging mice at > macroscopic scale), and John Fox continues to > decline to do a demo, > ask if you can buy it on contingency - that is, if > it works, you pay > for it, if it does not, you ship it back (negotiate > in advance who > should be for shipping and whether a restocking fee > is in order - > after all, nce you've used it, it cannot be sold as > new). > > BD CARV HCS system (Pathway Bioimager) - I've seen > the one in our > building used for Fura-2 ratio calcium imaging. > Looked ok. I just saw > it in action for live pancreatic islet cell imaging, > did not touch > any knobs so can't say how sensitive it is compared > to other systems > (and did not see any focus changes, so cannot > address how confocal it > is). It was being run for sequential well timelapse > assays, not HCS, > so I cannot address how well it works for HCS. > > best wishes, > > George > > > > At 04:29 AM 1/20/2008, you wrote: > >Search the CONFOCAL archive at > >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > > >Dear George, > > > >Thanks indeed for elaborate information. > > > >Among high end in vivo imaging system, on papers > >Xenogen did appear very good, especially for deep > >seated bioluminescence, but I could not find > reports > >on imaging of GFP or RFP expressing metastatic > cells. > >I am interested on imaging metastasis of GFP or RFP > >expressing cells in liver and lymphnodes. What is > your > >opinion and experience reagrding this application > with > >Xenogen system? > > > >Among lower end systems, FluorVivo of INDECO > >BioSystems and Pan-a-See-Ya of Lightools looked > good. > >But, I could not get demo of Pan-a See-Ya from John > >Fox. Do you know any users who have sucessfuly used > >any of these for imaging metastasis in lymphnodes > and > >liver? > > > >Do you have any infromation on performance HCS > system > >of BD that has in built Nipkow disc based confocal > >system? > > > >Once again thanks a lot for the information. > >With best regards, > >Nishigandha Naik > > > >--- George McNamara <[hidden email]> > wrote: > > > > > Search the CONFOCAL archive at > > > > >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > > > > > Hi Nishigandha, > > > > > > As always - get a demo! Really more a question > of > > > budget and throughput. > > > > > > For in vivo, the Caliper/Xenogen IVIS Spectrum > looks > > > very good. > > > > >http://www.caliperls.com/products/optical-imaging/ivis-spectrum.htm > > > > > > My colleagues at UM are upgrading our IVIS 200 > to > > > the Spectrum in a > > > month. Advantage of the IVIS systems is that > using > > > cryogenic > > > back-illuminated CCD's enables luciferase > > > bioluminescence imaging > > > (BLI) in vivo (and ex vivo - extremely valuable > to > > > take out the > > > organs at the end and re-image without the rest > of > > > the animal in the > > > way. also, can image multi-well plates, so can > test > > > the performance > > > of cell lines before putting them in the mouse). > > > A huge advantage of the Xenogen systems is that > they > > > have made the > > > effort to provide quantitative data for > > > bioluminescence. They use > > > photons/second/cm^2/steradian. This enables > every > > > paper published > > > with a Xenogen system to be compared to each > other > > > (of course, the > > > depth of the cells, expression level, and amount > of > > > luciferin at the > > > cells are additional variables). If you get a > > > non-Xenogen BLI > > > instrument, make sure the vendor validates the > same > > > units. Xenogen > > > now does something similar for fluorescence > > > tomography (a trickier issue). > > > > > > CRi has been in the in vivo fluorescence field > for > > > several years. > > > Check out their > > > http://www.cri-inc.com/products/maestro2.asp > > > > > > If your budget is on the low end, arrange a demo > > > from John Fox of > > > Lightools > (http://www.lightools.com/tutorial.htm). I > ____________________________________________________________________________________ Looking for last minute shopping deals? Find them fast with Yahoo! Search. http://tools.search.yahoo.com/newsearch/category.php?category=shopping |
«
Return to Confocal Microscopy List
|
1 view|%1 views
| Free forum by Nabble | Edit this page |