York Microscopy Job
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Please find below information about a new role at York. The deadline is very short. https://jobs.york.ac.uk/wd/plsql/wd_portal.show_job?p_web_site_id=3885&p_web_page_id=161773 The post provides an exciting opportunity under the Knowledge Transfer Programme to be involved in the development, application and commercialisation of novel biological imaging technology involving the Imaging & Cytometry Laboratory at the University of York and Phase Focus Ltd whilst also gaining further skills training. The post is an ideal stepping stone that can help further skills associated and needed within academia and industry. The training elements that are associated with the programme are highly desirable. The chance to also join a technology development still in its early stages is also very exciting. Best regards Peter -- Dr Peter O'Toole Head of Imaging and Cytometry Technology Facility Department of Biology (Area 15) University of York YORK YO10 5DD Tel : +44 (0)1904 328722 Fax : +44 (0)1904 328804 email : [hidden email] www.york.ac.uk/biology/tf Times Higher Education University of the Year 2010 EMAIL DISCLAIMER http://www.york.ac.uk/docs/disclaimer/email.htm |
 
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Jerry (Gerald) Sedgewick |
Online Photoshop Course for Microscopy (commercial?)
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Fellow Microscopists, You can now go to quickphotoshop.com to either take an online course in the use of Photoshop specifically for images from microscopes, or recommend the course to those who need it. I suspect that many of you will run across a person or two in the course of the following week who have used post-processing in ways that resulted in saturated pixels, and other completely avoidable mistakes, if only they could be trained. My hope is that this course will remedy imaging "ills." You might also be surprised at some functionality in Photoshop. Image stacks can be z-projected using the normal methods (mean, max, min), but also by using median, standard deviation, kurtosis, skewness, range, variance and summation (the caveat is that image stacks need to be saved as uncompressed AVI, and a post-CS3 version of Photoshop must be used). Other niceties include photostitching with blended edges, extended focus, extended dynamic range, histogram matching, linear histogram matching, pseudocoloring with the ability to edit colors, colorizing/decolorizing and some functions to reduce/eliminate darkness at image edges (vignetting). Also, tools exist to measure tones, look at "live" histograms, and create graphic overlays (to avoid oversaturation) so that objective means can be used to make the right decisions. If it weren't for the availability of tools for making objective decisions versus how an image "looks" on the screen, I would look elsewhere for post-processing. The full gamut of what students learn is listed at quickphotoshop.com (which goes to imagingandanalysis.com). There is a student price, as well, for the course, and a certificate at completion. My hope is to get more people adequately trained so that images are not overly processed and well within acceptable community standards. Best, Jerry -- Jerry (Gerald) Sedgewick Scientific Imaging & Image Analysis Consultant Technical Writer / Regulatory Consultant 965 Cromwell Avenue Saint Paul, MN 55114 651-788-2261 http://www.imagingandanalysis.com http://www.quickphotoshop.com Author: “Scientific Imaging with Photoshop: Methods, Measurement and Output” |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Dear All Does anybody have experience in using td-tomato fluorescent protein for FRAP? I will be grateful to hear about your experiences. Thanks in advance Julia Julia Edgar University of Glasgow UK |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi Julia, Many here can no doubt tell you that tdTomato is quite amenable to FRAP. In our hands tdTom bleaches when you hit it with highish levels of almost any light from 405 to 561 nm, and it makes a nice basis for experiments where you bleach the red channel while using counterstaining in the green channel to, e.g., track endosomes as they move. You can see my own experience with it in Feinstein et al., Nature Chem. Biol., 7:278 (2011). All the best, TF Timothy Feinstein, PhD Visiting Research Associate Laboratory for GPCR Biology Dept. of Pharmacology & Chemical Biology University of Pittsburgh, School of Medicine BST W1301, 200 Lothrop St. Pittsburgh, PA 15261 On Feb 21, 2013, at 4:19 PM, Julia Edgar wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Dear All > Does anybody have experience in using td-tomato fluorescent protein for FRAP? > I will be grateful to hear about your experiences. > Thanks in advance > Julia > > Julia Edgar > University of Glasgow > UK |
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In reply to this post by Julia Edgar
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi Julia, it might be important to keep in mind that tdtomato forms dimers. If you are planning to study molecular kinetics using FRAP (e.g. extract estimates for the diffusion coefficient or rate constants of binding reactions) it would be better to chose a fluorescent proteins that does not easily dimerizes. For other studies, this might not be a concern. Best regards, Philipp ******************************************** Philipp A. Steffen Postdoctoral fellow - Ringrose laboratory Institute of Molecular Biotechnology Dr. Bohr-Gasse 3 1030 Vienna Austria [hidden email] On Feb 21, 2013, at 10:19 PM, Julia Edgar wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Dear All > Does anybody have experience in using td-tomato fluorescent protein for FRAP? > I will be grateful to hear about your experiences. > Thanks in advance > Julia > > Julia Edgar > University of Glasgow > UK |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** H Philipp, dsRed had dimerization problems, but Roger Tsien addressed that by expressing Tomato as a tandem dimer. Two FP's in a single tag makes it twice as bright (allowing for loss to quenching and homo-FRET) and lets them to form an intramolecular dimer rather than grab outside an outside binding partner like dsRed did. Do you mean that Tomato forms tetramers? That would be news to me. cheers, TF Timothy Feinstein, PhD Visiting Research Associate Laboratory for GPCR Biology Dept. of Pharmacology & Chemical Biology University of Pittsburgh, School of Medicine BST W1301, 200 Lothrop St. Pittsburgh, PA 15261 On Feb 21, 2013, at 4:38 PM, Philipp Steffen wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi Julia, > > it might be important to keep in mind that tdtomato forms dimers. If you are planning to study molecular kinetics using FRAP (e.g. extract estimates for the diffusion coefficient or rate constants of binding reactions) it would be better to chose a fluorescent proteins that does not easily dimerizes. For other studies, this might not be a concern. > > Best regards, > > Philipp > > ******************************************** > Philipp A. Steffen > Postdoctoral fellow - Ringrose laboratory > Institute of Molecular Biotechnology > Dr. Bohr-Gasse 3 > 1030 Vienna > Austria > [hidden email] > > On Feb 21, 2013, at 10:19 PM, Julia Edgar wrote: > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> Dear All >> Does anybody have experience in using td-tomato fluorescent protein for FRAP? >> I will be grateful to hear about your experiences. >> Thanks in advance >> Julia >> >> Julia Edgar >> University of Glasgow >> UK |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Dear List members Thank you for your helpful replies. Re. Tim's comment below, td-tomato (which behaves effectively as a monomer, as I understand it) is indeed a very bright fluorescent protein and does not form the puncta that I have observed with dsRed. With my FRAP studies, I aim to use tandem dimer (td)-tomato as a surrogate marker of SOLUBLE cytosolic proteins and hope to extract half-time to recovery estimates and diffusion co-efficients. Do any of the characteristics of td-tomato (e.g. such as its relatively large size in comparison to GFP) make this particularly difficult? Any further comments/suggestions will be gratefully received. Best wishes Julia ....dsRed had dimerization problems, but Roger Tsien addressed that by expressing Tomato as a tandem dimer. Two FP's in a single tag makes it twice as bright (allowing for loss to quenching and homo-FRET) and lets them to form an intramolecular dimer rather than grab outside an outside binding partner like dsRed did...... cheers, TF Timothy Feinstein, PhD Visiting Research Associate Laboratory for GPCR Biology Dept. of Pharmacology & Chemical Biology University of Pittsburgh, School of Medicine BST W1301, 200 Lothrop St. Pittsburgh, PA 15261 >> Dear All >> Does anybody have experience in using td-tomato fluorescent protein for FRAP? >> I will be grateful to hear about your experiences. >> Thanks in advance >> Julia >> >> Julia Edgar >> University of Glasgow >> UK |
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