http://confocal-microscopy-list.275.s1.nabble.com/microbead-calibration-to-set-psf-tp2560403p2564208.html
a few beads of a larger size. Then you can easily find the surface of
> Europium chelates are phosphorescent rather than luminescent - that
> is to say, they have a very long - millisecond - fluorescence
> lifetime. That should still work in widefield but would be hopeless
> for confocal. Pretty much unfadeable, which is a plus.
>
> Tetraspeck bead are for calibrating colocalization rather than PSF -
> having multiple fluorochromes on one bead must reduce the intensity
> at any one wavelength. As I said before, I've used bead from Bangs
> Labs and I've had good images from beads down to 60nm. (Though at
> that scale it requires care and patience).
>
> Guy
>
>
> Optical Imaging Techniques in Cell Biology
> by Guy Cox CRC Press / Taylor & Francis
>
http://www.guycox.com/optical.htm> ______________________________________________
> Associate Professor Guy Cox, MA, DPhil(Oxon)
> Electron Microscope Unit, Madsen Building F09,
> University of Sydney, NSW 2006
> ______________________________________________
> Phone +61 2 9351 3176 Fax +61 2 9351 7682
> Mobile 0413 281 861
> ______________________________________________
>
http://www.guycox.net>
>
>
> From: Confocal Microscopy List [mailto:
[hidden email]
> ] On Behalf Of David Knecht
> Sent: Tuesday, 31 March 2009 11:57 PM
> To:
[hidden email]
> Subject: Re: microbead calibration to set psf?
>
> I am looking for some very bright 100nm beads. I tried to look for
> the beads you mention and it looks like Duke Scientific no longer
> exists and is now part of hte Thermo conglomerate. The closest
> thing I found to what you describe is the Europium chelate 100nm
> beads. Is that what you were referrring to? THey provide very
> little data on fluorescence properties and they are unusual in
> apparently being broad excitation spectrum but there is no emission
> spectrum in their literature I can find. Since they fluoresce
> maximally at 613nm it would seem they would be a poor fit for most
> FITC, TRITC, CY3 or CY5 type filter sets. Can you clarify your
> experience with them? Dave
>
> On Mar 31, 2009, at 3:36 AM, Mariette P.C. Kemner - van de Corput
> wrote:
>
>> I have good experience with fluorescent beads from Duke scientific.
>> The
>> 100nm beads are very bright and are perfect for determining the
>> PSF. I use
>> the red, green and blue 100nm beads to determine the PSF in three
>> channels
>> and to deconvolved my 3 colour FISH images.
>> I have trouble finding the 100nm and 200nm TetraSpeck beads on my
>> slide
>> beacuse they are rather weak in intensity. So that's why I switched
>> to the
>> single coloured 100nm beads from DS. The 500nm TetraSpeck beads are
>> very
>> good and I use them to determine the chromatic shift in 4 channels.
>> Once
>> made, bead slides can be used for a long period of time.
>>
>> Mariette
>>
>> ____________________________________________
>>
>> Dr. M.P.C. Kemner-van de Corput,
>> ____________________________________________
>>
>> MGC - Dept. of Cell Biology & Genetics
>> Erasmus Medical Center
>> Dr. Molewaterplein 50, 3015 GE Rotterdam
>> POB 2040, 3000 CA Rotterdam, The Netherlands
>>
>> Office: H-Ee751; tel: +31 10 704.3949
>> Lab: H-Ee710; tel lab: +31 10 704.3315
>> tel secr: +31 10 704.3169
>> ____________________________________________
>>
>>
http://www2.eur.nl/fgg/ch1/cellbiology/>>
http://www.thesis.kemner.biz/>> ____________________________________________
>>
>> Op Di, 31 maart, 2009 1:59 am, schreef Matiar Jafari:
>>> hey again,
>>>
>>> I'm trying to find out how and what the best way to setup my own psf
>>> is in AutoQuant. I know i need microbeads and im wondering what
>>> brand
>>> and what product i need and then how i need to shoot it and load it.
>>> Also what diameter does it need to be. A cat# would be great too
>>> also
>>> i don't know if it matters on what im shooting but just to put it
>>> out
>>> there im shooting gfp slices. Sorry if this is vague. Any help
>>> would
>>> be greatly appreciated. Thanks in advance
>>>
>>> Thank You
>>> --
>>> Matiar Jafari
>>>
>
> Dr. David Knecht
> Department of Molecular and Cell Biology
> Co-head Flow Cytometry and Confocal Microscopy Facility
> U-3125
> 91 N. Eagleville Rd.
> University of Connecticut
> Storrs, CT 06269
> 860-486-2200
> 860-486-4331 (fax)
>
>
>
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