Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal-Valeria,There is a Zeiss publication where they give the formula that calculates the thickness of the optical sections as a function of pinhole diameter, wavelength, refractive index, and numerical aperture. As Michael said, it is a rather complex formula. The more you open the pinhole, the more out of focus contribution you will get from farther away from the focal plane, so it's a somewhat arbitrary definition... it all depends where you set the threshold for what you "see" and what you don't "see" (typically, expressed in units of FWMH or full-width at max height). In any event, if you send me an email, I can reply with an attachment of the Zeiss paper (I did not get your original posting, so I don't have your personal address).As a side note, the Zeiss LSM 510 acquisition software will tell you what your optical thickness is for any combination of objective, wavelength and pinhole aperture. This is used to for instance make sure you are collecting images of similar "thickness" in different channels by adjusting the pinhole aperture, for instance for colocalization studies.The publication is titled: Microscopy from Carl Zeiss. Principles. Confocal laser scanning microscopy. You may be able to Google it too...--Julio VazquezFred Hutchinson Cancer Research Center1100 Fairview Ave. N., mailstop DE-512Seattle, WA 98109-1024=On Jul 8, 2008, at 7:02 AM, MODEL, MICHAEL wrote:Search the CONFOCAL archive atHi ValeriaI don't think there's a simple formula, but in the 2nd edition ofConfocal Handbook there is a chapter by Wilson that talks about theeffect of pinhole on resolution.MikeMichael Model, Ph.D.Confocal Microscopy CoreDpt. Biological SciencesKent State UniversityKent, OH 44242tel. 330-672-2874-----Original Message-----From: Confocal Microscopy List [[hidden email]] OnBehalf Of Valeria BernoSent: Tuesday, July 08, 2008 7:47 AMTo: [hidden email]Subject: pinhole-thicknessSearch the CONFOCAL archive atDear all,I know this is a very basic question but I am having hard time to findtheanswer.How can I calculate the thickness of the optical section I am aquiringwith different pinhole size?So waht is its relation with objective resolution?Thanks in advanceValeria
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