Index of refraction of aqueous media and/or cell interior

[John Oreopoulos]

============================================================
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi all,

I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.

I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?

And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:

glass (used in coverslips): ~1.523
water ~1.3326
inside cells ~ 1.38

Thanks!

John Oreopoulos

[Glen MacDonald-2]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

John,
Look at Dirkxx etal, J Biomedical Optics, 10(4), 2005 for a start.

the coverglass values come from the glass manufacturers and is stated for a single line at 20 deg. C.  
Another issue is the effect of cross-linking fixatives on RI.  

Regards,
Glen
Glen MacDonald
Core for Communication Research
Virginia Merrill Bloedel Hearing Research Center
Box 357923
University of Washington
Seattle, WA 98195-7923  USA
[phone]
[hidden email]








On Sep 14, 2010, at 8:34 AM, John Oreopoulos wrote:

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos

[Jennifer Reiber Kyle]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

John,

A great source for RI information is "Optical Clearing of Tissues and
Blood" by V. Tuchin. This book lists many different sources for RI of
tissue and cell components and also how processing affects these
values.

Jennifer Reiber Kyle
Ph.D. Candidate
Department of Electrical Engineering
University of California Riverside

On Tue, Sep 14, 2010 at 8:34 AM, John Oreopoulos
<[hidden email]> wrote:

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos

[Craig Brideau]

============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ I wrote my Master's thesis on this very topic:

http://www.dal.worldcat.org/title/low-coherence-light-for-measuring-optical-properties-of-biological-systems/oclc/437599652&referer=brief_results

Not my best work, but enjoy!

Craig

On Tue, Sep 14, 2010 at 10:25 AM, Jennifer Reiber Kyle <[hidden email]> wrote:

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

John,

A great source for RI information is "Optical Clearing of Tissues and
Blood" by V. Tuchin. This book lists many different sources for RI of
tissue and cell components and also how processing affects these
values.

Jennifer Reiber Kyle
Ph.D. Candidate
Department of Electrical Engineering
University of California Riverside

On Tue, Sep 14, 2010 at 8:34 AM, John Oreopoulos
<[hidden email]> wrote:
> ============================================================
> To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos

[Rosemary.White]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Another good source for organics and inorganics is the Handbook of Chemistry
and Physics.  Not so good for cell components.....
cheers,
Rosemary

Rosemary White
CSIRO Plant Industry
GPO Box 1600
Canberra, ACT 2601
Australia

T 61 2 6246 5475
F 61 2 6246 5334
E [hidden email]



On 15/09/10 2:25 AM, "Jennifer Reiber Kyle" <[hidden email]>
wrote:

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> John,
>
> A great source for RI information is "Optical Clearing of Tissues and
> Blood" by V. Tuchin. This book lists many different sources for RI of
> tissue and cell components and also how processing affects these
> values.
>
> Jennifer Reiber Kyle
> Ph.D. Candidate
> Department of Electrical Engineering
> University of California Riverside
>
> On Tue, Sep 14, 2010 at 8:34 AM, John Oreopoulos
> <[hidden email]> wrote:
>> ============================================================
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> ============================================================
>>
>> Hi all,
>>
>> I was just wondering if there is a simple method using basic imaging tools
>> available with most research microscopes (DIC, phase contrast, confocal,
>> etc.) to quantitatively determine the index of refraction of of the aqueous
>> media in your imaging chamber or even in the cell interior.
>>
>> I know that the index of refraction inside the cell is not uniform, and I
>> believe that several modes of imaging actually depend on this fact, but
>> again, is there a way to quantitate this (and possibly map it in 2D by
>> imaging)? I remember I had seen some papers a few years ago about
>> quantitative phase contrast microscopy, but this is likely something very
>> complicated with non-traditional optics (ie: not something you'd find in a
>> basic microscope). I just need something quick and easy to get a handle on
>> these numbers in real samples and to see how much they vary from sample to
>> sample. How much variation in the index of refraction would one expect to
>> find in cells, and is this cell type and/or environment dependent? Are there
>> any table of values out there in the literature?
>>
>> And for reference, can someone point me to a good source of the numbers that
>> usually get quoted for glass (coverslips), water, and cell interiors? I think
>> these are typical values that are commonly used, but I don't know where they
>> come from and how they were measured:
>>
>> glass (used in coverslips): ~1.523
>> water ~1.3326
>> inside cells ~ 1.38
>>
>> Thanks!
>>
>> John Oreopoulos

[Jeremy Adler-3]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Perfusing media containing cells with roller pumps has the potential  
to damage suspended cells - does anyone have any quantitation or refs.

[Johannes Helm]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Good afternoon,

a comparatively simple method might be to actually measure the refractive
index, at least of the mounting medium, using an Abbé B refractometer.
Most institutes with a chemistry department will have one. It's a device,
which you can easily carry to your own lab, if you get it on loan.

The main problem with older Abbé refractometers is that they may be
limited to one wavelength, only. Other refractometers, which overcome this
problem, are of the Pulfrich or the Michelson type, though they are much
more cumbersome to use.

Best wishes,

Johannes Helm

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools
> available with most research microscopes (DIC, phase contrast, confocal,
> etc.) to quantitatively determine the index of refraction of of the
> aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I
> believe that several modes of imaging actually depend on this fact, but
> again, is there a way to quantitate this (and possibly map it in 2D by
> imaging)? I remember I had seen some papers a few years ago about
> quantitative phase contrast microscopy, but this is likely something very
> complicated with non-traditional optics (ie: not something you'd find in a
> basic microscope). I just need something quick and easy to get a handle on
> these numbers in real samples and to see how much they vary from sample to
> sample. How much variation in the index of refraction would one expect to
> find in cells, and is this cell type and/or environment dependent? Are
> there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers
> that usually get quoted for glass (coverslips), water, and cell interiors?
> I think these are typical values that are commonly used, but I don't know
> where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos

--
P. Johannes Helm, M.Sc. PhD
Seniorengineer
CMBN
University of Oslo
Institute of Basic Medical Science
Department of Anatomy
Postboks 1105 - Blindern
NO-0317 Oslo

Voice: [phone]
Fax: [phone]

WWW: folk.uio.no/jhelm

[mmodel]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi John

For liquid media you can use reflection from a spherical lens immersed in the liquid whose n you want to measure. (Model, M.A., Khitrin A.K. and Blank J.L. Measurement of the absorption of concentrated dyes and their use for quantitative imaging of surface topography. J. Microsc. 231:156-167)

Most measurements of n on cells and organelles that I know of produced values from 1.35 to 1.45

Mike

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of John Oreopoulos
Sent: Tuesday, September 14, 2010 11:34 AM
To: [hidden email]
Subject: Index of refraction of aqueous media and/or cell interior

============================================================
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi all,

I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.

I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?

And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:

glass (used in coverslips): ~1.523
water ~1.3326
inside cells ~ 1.38

Thanks!

John Oreopoulos

[Johannes-P. Koch]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

  John,

I do not know whether this might help you, but...

Do you really need to measure the RI using your microscope???

Typically, the RI (for any kind of liquid or solid stuff) is measured
using a refractometer (Snell's law). Since the RI is
temperature-dependent, you have to correct for that as well, but there
are interpolation formulae available (for small scale interpolation,
i.e. a few degrees). Presumably, you can measure the RI of a cell
suspension as well - however, your result will be the average of the RIs
and you won't have spatial resolution.

Again, I do not know whether this might help you, but I am sure that
this method is widely used and has also been used for determining the
values you indicated for water, glass,...

Best regards,
Johannes



Am 15.09.2010 15:22, schrieb MODEL, MICHAEL:

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi John
>
> For liquid media you can use reflection from a spherical lens immersed in the liquid whose n you want to measure. (Model, M.A., Khitrin A.K. and Blank J.L. Measurement of the absorption of concentrated dyes and their use for quantitative imaging of surface topography. J. Microsc. 231:156-167)
>
> Most measurements of n on cells and organelles that I know of produced values from 1.35 to 1.45
>
> Mike
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of John Oreopoulos
> Sent: Tuesday, September 14, 2010 11:34 AM
> To: [hidden email]
> Subject: Index of refraction of aqueous media and/or cell interior
>
> ============================================================
> To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos
>

--
Mag. Johannes-P. KOCH
Department of Biochemistry and Cell Biology
MFPL, University of Vienna
Dr. Bohrgasse 9/5
A-1030 Vienna
Austria

phone: [phone]
fax: [phone]

mail to: [hidden email]

[Rietdorf, Jens]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi John,

just as an idea, TIR measurements should work as the critical angle
depends on the ratio of refractive indices at the glass-medium
interface. The critical angle is easy to spot by the reflection of the
laser in an objective type TIR setup.
Essentially it should also work at a lipid bilayer-intracellular fluid
interface, but I guess that is technically more demanding.

Cheers, jens

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]]
On Behalf Of John Oreopoulos
Sent: Tuesday, September 14, 2010 17:34 PM
To: [hidden email]
Subject: Index of refraction of aqueous media and/or cell interior

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi all,

I was just wondering if there is a simple method using basic imaging
tools available with most research microscopes (DIC, phase contrast,
confocal, etc.) to quantitatively determine the index of refraction of
of the aqueous media in your imaging chamber or even in the cell
interior.

I know that the index of refraction inside the cell is not uniform, and
I believe that several modes of imaging actually depend on this fact,
but again, is there a way to quantitate this (and possibly map it in 2D
by imaging)? I remember I had seen some papers a few years ago about
quantitative phase contrast microscopy, but this is likely something
very complicated with non-traditional optics (ie: not something you'd
find in a basic microscope). I just need something quick and easy to get
a handle on these numbers in real samples and to see how much they vary
from sample to sample. How much variation in the index of refraction
would one expect to find in cells, and is this cell type and/or
environment dependent? Are there any table of values out there in the
literature?

And for reference, can someone point me to a good source of the numbers
that usually get quoted for glass (coverslips), water, and cell
interiors? I think these are typical values that are commonly used, but
I don't know where they come from and how they were measured:

glass (used in coverslips): ~1.523
water ~1.3326
inside cells ~ 1.38

Thanks!

John Oreopoulos

[Guy Cox-2]

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

If you can accurately control all other parameters you could just measure the RI using the correction collar on a water immersion lens.  High precision coverslips, of course, and standardised thickness of liquid (thin plastic shim?) then calibrate with standard solutions.  Bit of work but no extra equipment.  There are tables for refractive indices of sugar solutions since it's used in the food and wine industry - there may also be tables for salt solutions.  Very lightly smoke the slide and scratch to form a few small black bits (other methods could work) and correct SA with given solution.

                                          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)
Australian Centre for Microscopy & Microanalysis,
Madsen Building F09, University of Sydney, NSW 2006

Phone [phone]     Fax [phone]
             Mobile [phone]
______________________________________________
      http://www.guycox.net
 


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of P. Johannes Helm
Sent: Wednesday, 15 September 2010 8:17 PM
To: [hidden email]
Subject: Re: Index of refraction of aqueous media and/or cell interior

============================================================
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Good afternoon,

a comparatively simple method might be to actually measure the refractive
index, at least of the mounting medium, using an Abbé B refractometer.
Most institutes with a chemistry department will have one. It's a device,
which you can easily carry to your own lab, if you get it on loan.

The main problem with older Abbé refractometers is that they may be
limited to one wavelength, only. Other refractometers, which overcome this
problem, are of the Pulfrich or the Michelson type, though they are much
more cumbersome to use.

Best wishes,

Johannes Helm

> ============================================================
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> ============================================================
>
> Hi all,
>
> I was just wondering if there is a simple method using basic imaging tools
> available with most research microscopes (DIC, phase contrast, confocal,
> etc.) to quantitatively determine the index of refraction of of the
> aqueous media in your imaging chamber or even in the cell interior.
>
> I know that the index of refraction inside the cell is not uniform, and I
> believe that several modes of imaging actually depend on this fact, but
> again, is there a way to quantitate this (and possibly map it in 2D by
> imaging)? I remember I had seen some papers a few years ago about
> quantitative phase contrast microscopy, but this is likely something very
> complicated with non-traditional optics (ie: not something you'd find in a
> basic microscope). I just need something quick and easy to get a handle on
> these numbers in real samples and to see how much they vary from sample to
> sample. How much variation in the index of refraction would one expect to
> find in cells, and is this cell type and/or environment dependent? Are
> there any table of values out there in the literature?
>
> And for reference, can someone point me to a good source of the numbers
> that usually get quoted for glass (coverslips), water, and cell interiors?
> I think these are typical values that are commonly used, but I don't know
> where they come from and how they were measured:
>
> glass (used in coverslips): ~1.523
> water ~1.3326
> inside cells ~ 1.38
>
> Thanks!
>
> John Oreopoulos

--
P. Johannes Helm, M.Sc. PhD
Seniorengineer
CMBN
University of Oslo
Institute of Basic Medical Science
Department of Anatomy
Postboks 1105 - Blindern
NO-0317 Oslo

Voice: [phone]
Fax: [phone]

WWW: folk.uio.no/jhelm

[George McNamara]

============================================================ To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ============================================================ Hi John,

Someplace in the middle of Multi-Probe Microscopy, http://home.earthlink.net/~mpmicro/mpmicro.zip is a long section on refractive index of cells and media. Search for the phrase      Johnsen and Widder (1999) (their table 1) list     (it is on page 621 of the version I pulled up on my PC. No edits in a while - I moved from Sacramento in 2007).

PubMed - try searching for:    suhling refractive index

See also www.iatia.com.au    for example,
Refractive index measurement in viable cells using quantitative phase-amplitude microscopy and confocal microscopy.
Curl CL, Bellair CJ, Harris T, Allman BE, Harris PJ, Stewart AG, Roberts A, Nugent KA, Delbridge LM.
Cytometry A. 2005 May;65(1):88-92.PMID: 15800856


Graham Dunn published DRIMAS,  for dry mass of cells (just subtract water):

J Cell Sci. 1989 Mar;92 ( Pt 3):379-89.

Microinterferometry of the movement of dry matter in fibroblasts.

Brown AF, Dunn GA.


Q. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent?

A. Yes, every cytoplasm -> membrane -> lumen, is two changes in RI. Cells with lots of vesicles, pancreatic beta cells for example, scatter lots of light (bonus: insulin zinc granules may be practically protein crystals, potentially RI > 1.5).

Enjoy,

George
 


At 11:34 AM 9/14/2010, you wrote:

============================================================
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
============================================================

Hi all,

I was just wondering if there is a simple method using basic imaging tools available with most research microscopes (DIC, phase contrast, confocal, etc.) to quantitatively determine the index of refraction of of the aqueous media in your imaging chamber or even in the cell interior.

I know that the index of refraction inside the cell is not uniform, and I believe that several modes of imaging actually depend on this fact, but again, is there a way to quantitate this (and possibly map it in 2D by imaging)? I remember I had seen some papers a few years ago about quantitative phase contrast microscopy, but this is likely something very complicated with non-traditional optics (ie: not something you'd find in a basic microscope). I just need something quick and easy to get a handle on these numbers in real samples and to see how much they vary from sample to sample. How much variation in the index of refraction would one expect to find in cells, and is this cell type and/or environment dependent? Are there any table of values out there in the literature?

And for reference, can someone point me to a good source of the numbers that usually get quoted for glass (coverslips), water, and cell interiors? I think these are typical values that are commonly used, but I don't know where they come from and how they were measured:

glass (used in coverslips): ~1.523
water ~1.3326
inside cells ~ 1.38

Thanks!

John Oreopoulos

George McNamara, Ph.D.
Image Core Manager
Analytical Imaging Core Facility
University of Miami, Miller School of Medicine
Miami, FL 33136
[hidden email]
[hidden email]
305-243-8436 office
http://www.sylvester.org/AICF (Analytical Imaging Core Facility)
http://www.sylvester.org/AICF/pubspectra.zip (the entire 2000+ spectra .xlsx file is in the zip file)
http://home.earthlink.net/~geomcnamara