http://confocal-microscopy-list.275.s1.nabble.com/Oil-vs-water-objectives-tp7579114p7579127.html
the water lens. )Find a point object: focus up
focus. DON'T just try to "make it sharper!" The
correct for. One can easily see the difference
>Make sure the coverslip is orthogonal to the
>image axis with your water objective.
>
>A common aberration with water-immersion objective lenses.
>J Microsc. 2004 Oct;216(Pt 1):49-51.
>
http://www.ncbi.nlm.nih.gov/pubmed/15369482>
>Doug
>
>^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^
>Douglas W. Cromey, M.S. - Associate Scientific Investigator
>Dept. of Cellular & Molecular Medicine, University of Arizona
>1501 N. Campbell Ave, Tucson, AZ 85724-5044 USA
>
>office: AHSC 4212 email:
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>-----Original Message-----
>From: Confocal Microscopy List
>[mailto:
[hidden email]] On
>Behalf Of Steffen Dietzel
>Sent: Thursday, October 04, 2012 7:30 AM
>To:
[hidden email]
>Subject: Re: Oil vs water objectives
>
>*****
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>
>I second Michael's comment.
>
>Apart from that, when you have an aqueous(!)
>preparation, spherical aberration decreases
>image quality faster for the oil objective. So,
>directly behind the coverslip the oil objective
>(1.4) will give you the better image due to the
>higher NA, at some depth both objectives will
>give you similar quality and beyond that, the
>water immersion will be better.
>
>I seem to remember that for oil 1.4 and water 1.2 the crossing point is
>10-20 µm into the sample.
>
>If you have the Handbook laying around, check
>the chapter on Lens aberrations. Chapter 20 by
>Hell and Stelzer in the second edition, chapter
>20 by Egner and Hell in the 3rd edition.
>
>Needles to say that all above considerations
>assume that both objectives have the same
>transparency for excitation and emission
>wavelengths - which they probably don't. So we
>are back at Michael's comment: You've got to
>test it. Beads attached to the coverslip and the
>slide (various preparations with various
>distances between the two) give good test
>objects.
>
>Steffen
>
>On 04.10.2012 15:15, Gabriel Lapointe wrote:
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>>
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>> *****
>>
>> Hi,
>>
>> I have a user who insist that using a 1,27NA water immersion objective
>> is brighter and would give better images than using a 1,4NA oil
>> immersion. I understand that deeper into the media that would be true.
>> But, in that particular case, we are talking about imaging GFP at less
>> than 100 micron away with a spinning disk.
>>
>> So, I was wondering at which distance from the coverslips do we start
>> seeing benefits of using a water immersion objective over an oil
>> objective in aqueous media.
>>
>> Thanks for your help.
>>
>> Sincerely
>> *Gabriel Lapointe, M.Sc.*
> > Lab Manager / Microscopy Specialist
>> Concordia University, Biology Department
>> 7141 Sherbrooke St. West SP 534
>> Montréal QC H4B 1R6 Canada
>>
[hidden email]
>> cmac.concordia.ca
>>
http://gabriellapointe.ca>>
>
>
>--
>------------------------------------------------------------
>Steffen Dietzel, PD Dr. rer. nat
>Ludwig-Maximilians-Universität München
>Walter-Brendel-Zentrum für experimentelle
>Medizin (WBex) Head of light microscopy
>
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