Confocal Microscopy List

reflection mode in material science

Classic

List

Threaded

5 messages Options

jlribas

reflection mode in material science

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Dear all,
Several groups in our University are moving now to the confocal to
perform z profiles of materials. I would like to get more information
about the way of doing a correct adquisition in reflection mode to
ensure a realistic depth map. I've found that a z profile in the same
region with different air objectives give very different measurements,
so I'm not sure which is the real value. And is very difficult to obtain
some useful information about this technique.
I'll be very grateful if someone could recomend me some link, book or
paper about reflection mode and z profile caracterization in materials.
All the best

Juan Luis

--
Juan Luis Ribas, PhD
Servicio de Microscopía
Centro de Investigación, Tecnología e Innovación
Universidad de Sevilla
Av. Reina Mercedes 4b
41012 Sevilla

Tfno: 954559983

mmodel

Re: reflection mode in material science

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

It's typically done using interferometry. Reflection mode in confocal can be tricky because the signal depends on the tilt of the surface (see P.C.Cheng's articles in Confocal Handbook)

Mike

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Juan Luis Ribas
Sent: Tuesday, November 06, 2007 1:19 PM
To: [hidden email]
Subject: reflection mode in material science

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Dear all,
Several groups in our University are moving now to the confocal to
perform z profiles of materials. I would like to get more information
about the way of doing a correct adquisition in reflection mode to
ensure a realistic depth map. I've found that a z profile in the same
region with different air objectives give very different measurements,
so I'm not sure which is the real value. And is very difficult to obtain
some useful information about this technique.
I'll be very grateful if someone could recomend me some link, book or
paper about reflection mode and z profile caracterization in materials.
All the best

Juan Luis

--
Juan Luis Ribas, PhD
Servicio de Microscopía
Centro de Investigación, Tecnología e Innovación
Universidad de Sevilla
Av. Reina Mercedes 4b
41012 Sevilla

Tfno: 954559983

James Pawley

Re: reflection mode in material science

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi Juan,

I encourage you to pursue the use of this
instrument for looking at transparent inorganic
materials!

However, I also echo Mike's caution about the
effects of surface angle and add the point that
"materials" seldom have a refractive index of 1.

When using the no-coverslip dry objectives common
in materials, you have spherical aberration the
moment you penetrate into the material and any
measurements in z will require at least a
"back-of-the-envelope" correction for the actual
RI of the material between the near surface and
the focus plane. You will get different results
with different lenses because of differences in
NA (and therefore SA).

You can reduce this all somewhat if you can
immerse your "rock" in oil and use an oil
objective but be careful that the "rock" doesn't
get a chance to touch (i.e., "Scratch!") the
front element of the objective.

While the oil will help some, as the "rock" is
unlikely to have an RI of exactly 1.515, there
will still be some SA and the directional
dependence of surface reflections can still be a
problem.

On the other hand, you should be able to get good
contrast from small inclusions. If these are
small with respect to the wavelength of the
light, they scatter in all directions rather than
reflecting. (hence, "backscattered light" rather
than "reflected light")

It turns out that living cells are often full of
very small highly refractile objects are can be
imaged very well using this signal.

Cheers,

Jim P.

>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>It's typically done using interferometry.
>Reflection mode in confocal can be tricky
>because the signal depends on the tilt of the
>surface (see P.C.Cheng's articles in Confocal
>Handbook)
>
>Mike
>
>-----Original Message-----
>From: Confocal Microscopy List
>[mailto:[hidden email]] On Behalf
>Of Juan Luis Ribas
>Sent: Tuesday, November 06, 2007 1:19 PM
>To: [hidden email]
>Subject: reflection mode in material science
>
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>Dear all,
>Several groups in our University are moving now to the confocal to
>perform z profiles of materials. I would like to get more information
>about the way of doing a correct adquisition in reflection mode to
>ensure a realistic depth map. I've found that a z profile in the same
>region with different air objectives give very different measurements,
>so I'm not sure which is the real value. And is very difficult to obtain
>some useful information about this technique.
>I'll be very grateful if someone could recomend me some link, book or
>paper about reflection mode and z profile caracterization in materials.
>All the best
>
>Juan Luis
>
>--
>Juan Luis Ribas, PhD
>Servicio de Microscopía
>Centro de Investigación, Tecnología e Innovación
>Universidad de Sevilla
>Av. Reina Mercedes 4b
>41012 Sevilla
>
>Tfno: 954559983

--
**********************************************
Prof. James B. Pawley, Ph. 608-263-3147
Room 223, Zoology Research
Building, FAX
608-265-5315
1117 Johnson Ave., Madison, WI, 53706
[hidden email]
3D Microscopy of Living Cells Course, June 14-26, 2008, UBC, Vancouver Canada
Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2008
"If it ain't diffraction, it must be statistics." Anon.

mmodel

Re: transparent materials

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Speaking of transparent materials, you can image their surface topography by partially immersing them in a liquid with a high extinction coefficient. Transmission image can show surface roughness with nanometer vertical resolution (depends of the extinction coefficient of the liquid) and whatever horizontal resolution is achievable with your objective. Use of the laser is optional: a narrow band pass with a regular lamp will work fine.

Mike

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of James Pawley
Sent: Tuesday, November 06, 2007 5:50 PM
To: [hidden email]
Subject: Re: reflection mode in material science

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi Juan,

I encourage you to pursue the use of this
instrument for looking at transparent inorganic
materials!

However, I also echo Mike's caution about the
effects of surface angle and add the point that
"materials" seldom have a refractive index of 1.

When using the no-coverslip dry objectives common
in materials, you have spherical aberration the
moment you penetrate into the material and any
measurements in z will require at least a
"back-of-the-envelope" correction for the actual
RI of the material between the near surface and
the focus plane. You will get different results
with different lenses because of differences in
NA (and therefore SA).

You can reduce this all somewhat if you can
immerse your "rock" in oil and use an oil
objective but be careful that the "rock" doesn't
get a chance to touch (i.e., "Scratch!") the
front element of the objective.

While the oil will help some, as the "rock" is
unlikely to have an RI of exactly 1.515, there
will still be some SA and the directional
dependence of surface reflections can still be a
problem.

On the other hand, you should be able to get good
contrast from small inclusions. If these are
small with respect to the wavelength of the
light, they scatter in all directions rather than
reflecting. (hence, "backscattered light" rather
than "reflected light")

It turns out that living cells are often full of
very small highly refractile objects are can be
imaged very well using this signal.

Cheers,

Jim P.

jlribas

Re: reflection mode in material science

In reply to this post by James Pawley

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Thanks to everybody for all the interesting comments about this topic.

Juan Luis

James Pawley escribió:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Hi Juan,
>
> I encourage you to pursue the use of this instrument for looking at
> transparent inorganic materials!
>
> However, I also echo Mike's caution about the effects of surface angle
> and add the point that "materials" seldom have a refractive index of 1.
>
> When using the no-coverslip dry objectives common in materials, you
> have spherical aberration the moment you penetrate into the material
> and any measurements in z will require at least a
> "back-of-the-envelope" correction for the actual RI of the material
> between the near surface and the focus plane. You will get different
> results with different lenses because of differences in NA (and
> therefore SA).
>
> You can reduce this all somewhat if you can immerse your "rock" in oil
> and use an oil objective but be careful that the "rock" doesn't get a
> chance to touch (i.e., "Scratch!") the front element of the objective.
>
> While the oil will help some, as the "rock" is unlikely to have an RI
> of exactly 1.515, there will still be some SA and the directional
> dependence of surface reflections can still be a problem.
>
> On the other hand, you should be able to get good contrast from small
> inclusions. If these are small with respect to the wavelength of the
> light, they scatter in all directions rather than reflecting. (hence,
> "backscattered light" rather than "reflected light")
>
> It turns out that living cells are often full of very small highly
> refractile objects are can be imaged very well using this signal.
>
> Cheers,
>
> Jim P.
>
>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> It's typically done using interferometry. Reflection mode in confocal
>> can be tricky because the signal depends on the tilt of the surface
>> (see P.C.Cheng's articles in Confocal Handbook)
>>
>> Mike
>>
>> -----Original Message-----
>> From: Confocal Microscopy List [mailto:[hidden email]]
>> On Behalf Of Juan Luis Ribas
>> Sent: Tuesday, November 06, 2007 1:19 PM
>> To: [hidden email]
>> Subject: reflection mode in material science
>>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> Dear all,
>> Several groups in our University are moving now to the confocal to
>> perform z profiles of materials. I would like to get more information
>> about the way of doing a correct adquisition in reflection mode to
>> ensure a realistic depth map. I've found that a z profile in the same
>> region with different air objectives give very different measurements,
>> so I'm not sure which is the real value. And is very difficult to obtain
>> some useful information about this technique.
>> I'll be very grateful if someone could recomend me some link, book or
>> paper about reflection mode and z profile caracterization in materials.
>> All the best
>>
>> Juan Luis
>>
>> --
>> Juan Luis Ribas, PhD
>> Servicio de Microscopía
>> Centro de Investigación, Tecnología e Innovación
>> Universidad de Sevilla
>> Av. Reina Mercedes 4b
>> 41012 Sevilla
>>
>> Tfno: 954559983
>
>

--
Juan Luis Ribas
Servicio de Microscopía
Centro de Investigación, Tecnología e Innovación
Universidad de Sevilla
Av. Reina Mercedes 4b
41012 Sevilla

Tfno: 954559983