Re: High speed spinning disc confocal with EMCCD camera - commercial response

Posted by Guy Cox-2 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/High-speed-spinning-disc-confocal-with-EMCCD-camera-tp7583142p7583220.html

Hang on, you think I don’t know this??  Huh??  That wasn’t what I was talking about.

                                                     Guy

Guy Cox, Honorary Associate Professor
School of Medical Sciences

Australian Centre for Microscopy and Microanalysis,
Madsen, F09, University of Sydney, NSW 2006

From: George McNamara [mailto:[hidden email]]
Sent: Tuesday, 6 January 2015 4:28 PM
To: Confocal Microscopy List
Cc: Guy Cox
Subject: Re: High speed spinning disc confocal with EMCCD camera - commercial response

Hi Guy,

Nyquist is dead.

For mitochondria way beyond, see Shim et al 2012,
http://www.ncbi.nlm.nih.gov/pubmed/22891300
http://www.pnas.org/content/109/35/13978.long   (open access)

also several papers from Hell and colleagues, such as;

http://www.ncbi.nlm.nih.gov/pubmed/20205711
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2838807/pdf/1757-5036-3-4.pdf

and
30 nm isotropic resolution (isoSTED)
http://www.ncbi.nlm.nih.gov/pubmed/19459703

George
p.s. http://en.wikipedia.org/wiki/Harry_Nyquist
Harry Nyquist (né Harry Theodor Nyqvist; /<http://en.wikipedia.org/wiki/Help:IPA_for_English>ˈ<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>n<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>aɪ<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>k<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>w<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>ɪ<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>s<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>t<http://en.wikipedia.org/wiki/Help:IPA_for_English#Key>/<http://en.wikipedia.org/wiki/Help:IPA_for_English>, Swedish: [nʏːkvɪst]<http://en.wikipedia.org/wiki/Help:IPA_for_Swedish_and_Norwegian>; February 7, 1889 – April 4, 1976)

On 1/5/2015 10:10 AM, Guy Cox wrote:

Michael,



         Both you and I have been telling users this for years.  Likewise, if they want to image a mitochondrion, they gain nothing by going beyond Nyquist, yet they still zoom it up to 1024 x 768.  These people are supposed to be scientists - but are they?  A microscope is a scientific instrument, not a black box.



                              Guy



Guy Cox, Honorary Associate Professor

School of Medical Sciences



Australian Centre for Microscopy and Microanalysis,

Madsen, F09, University of Sydney, NSW 2006





-----Original Message-----

From: Cammer, Michael [mailto:[hidden email]]

Sent: Tuesday, 6 January 2015 2:57 AM

To: Guy Cox

Subject: RE: High speed spinning disc confocal with EMCCD camera - commercial response



I constantly tell people doing live imaging with the multiphoton who want to track cells to turn down the excitation and do running averages or other noise filtering later.  I show them examples but when I leave the room they invariably crank up the light because they want to see a perfect  low noise high contrast image right now and then they are disappointed when the experiment doesn't work either because the sample bleaches or the cells stop moving.

=========================================================================

 Michael Cammer, Microscopy Core & Skirball Institute, NYU Langone Medical Center

                          Cell:  914-309-3270     Temporary location:  SK2-7

          http://ocs.med.nyu.edu/microscopy & http://microscopynotes.com/





-----Original Message-----

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox

Sent: Monday, January 05, 2015 10:45 AM

To: [hidden email]<mailto:[hidden email]>

Subject: Re: High speed spinning disc confocal with EMCCD camera - commercial response





Not being able to saturate excitation is a good thing, not a bad thing!  I am perpetually telling people to reduce their excitation to 50% and check that their excitation decreases by the same amount - if not they are saturating (and destroying) their fluorochrome.  Nobody ever follows my advice - they just want a bright (and usually saturated and therefore uninformative) image.



                                   Guy





Guy Cox, Honorary Associate Professor

School of Medical Sciences



Australian Centre for Microscopy and Microanalysis, Madsen, F09, University of Sydney, NSW 2006










--







George McNamara, Ph.D.

Single Cells Analyst

L.J.N. Cooper Lab

University of Texas M.D. Anderson Cancer Center

Houston, TX 77054

Tattletales http://works.bepress.com/gmcnamara/42