Phase relief image with confocal setup + a piece of plastic or paper

[Peng Xi-2]

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Dear List,
      We have just published our work of achieveing DIC phase relief
image of transparent specimens with existing confocal setup, by just
adding a piece of plastic or paper (basically, anything that
fluorescent) on top of your sample. We also explained why so
mathematically. Link:
      http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100

      Also, a piece of advice: do NOT place printing paper on your
lips when you open your door ---- although not presented, we noticed
that the paper contain heavily whitening agent,  a strong fluorescence
emitter under UV excitation, which is toxic and may induce cancer.

Sincerely,
Peng Xi
Ph. D.    Associate Professor
Dept. of Biomedical Engineering, College of Engineering
Peking University, Beijing, China
Tel: [phone]
Email: [hidden email]
http://bme.pku.edu.cn/~xipeng

[Guy Cox-2]

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This is very neat, but to my non-mathematical mind looks very like the "poor man's Schlieren" (SSB) one can get by blocking half the illuminating aperture in widefield.   (I think that goes back to Abbe).  Maybe some of our more numerate members will put me right.

                                                                                  Guy

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Peng Xi
Sent: Tuesday, 12 June 2012 9:25 AM
To: [hidden email]
Subject: Phase relief image with confocal setup + a piece of plastic or paper

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Dear List,
      We have just published our work of achieveing DIC phase relief image of transparent specimens with existing confocal setup, by just adding a piece of plastic or paper (basically, anything that
fluorescent) on top of your sample. We also explained why so mathematically. Link:
      http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100

      Also, a piece of advice: do NOT place printing paper on your lips when you open your door ---- although not presented, we noticed that the paper contain heavily whitening agent,  a strong fluorescence emitter under UV excitation, which is toxic and may induce cancer.

Sincerely,
Peng Xi
Ph. D.    Associate Professor
Dept. of Biomedical Engineering, College of Engineering Peking University, Beijing, China
Tel: [phone]
Email: [hidden email]
http://bme.pku.edu.cn/~xipeng

[Peng Xi-2]

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I totally agree with you Guy. LOSOM is oblique microscopy, but
achieved with scanning optical microscope like confocal. It's image
quality is comparable to DIC though. Poor confocal man's good
Schlieren! :)

Sincerely,
Peng Xi
Ph. D.    Associate Professor
Dept. of Biomedical Engineering, College of Engineering
Peking University, Beijing, China
Tel: [phone]
Email: [hidden email]
http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100

On Tue, Jun 12, 2012 at 8:51 PM, Guy Cox <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> This is very neat, but to my non-mathematical mind looks very like the "poor man's Schlieren" (SSB) one can get by blocking half the illuminating aperture in widefield.   (I think that goes back to Abbe).  Maybe some of our more numerate members will put me right.
>
>                                                                                  Guy
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Peng Xi
> Sent: Tuesday, 12 June 2012 9:25 AM
> To: [hidden email]
> Subject: Phase relief image with confocal setup + a piece of plastic or paper
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear List,
>      We have just published our work of achieveing DIC phase relief image of transparent specimens with existing confocal setup, by just adding a piece of plastic or paper (basically, anything that
> fluorescent) on top of your sample. We also explained why so mathematically. Link:
>      http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100
>
>      Also, a piece of advice: do NOT place printing paper on your lips when you open your door ---- although not presented, we noticed that the paper contain heavily whitening agent,  a strong fluorescence emitter under UV excitation, which is toxic and may induce cancer.
>
> Sincerely,
> Peng Xi
> Ph. D.    Associate Professor
> Dept. of Biomedical Engineering, College of Engineering Peking University, Beijing, China
> Tel: [phone]
> Email: [hidden email]
> http://bme.pku.edu.cn/~xipeng

[Steffen Dietzel]

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I don't quite get it from the article. Assuming an upright confocal, you
put, say, a chroma test slide under the preparation, ok.

* How do you get to tilted/oblique illumination? Do you have to put the
specimen to the edge of the scan range? Or is it done with moving a
lense out of axis? (which would mean that this is not easily feasable on
commercial setups since they don't intend to give access to the beampath)

* Then you scan eg with 488 nm and record the green fluorescent signal
from the test slide, is that it?

* The actual image is then generated by the fluorescent light traveling
back through the specimen and getting deflected (phase shifted ?) or not
during the process, being finally recorded with a PMT.

* To get signal through you have to open the pinhole, since it is not a
confocal signal, right?.

Maybe you could clarify these points for the non-physicist.

Steffen


On 12.06.2012 01:25, Peng Xi wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear List,
>        We have just published our work of achieveing DIC phase relief
> image of transparent specimens with existing confocal setup, by just
> adding a piece of plastic or paper (basically, anything that
> fluorescent) on top of your sample. We also explained why so
> mathematically. Link:
>        http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100
>
>        Also, a piece of advice: do NOT place printing paper on your
> lips when you open your door ---- although not presented, we noticed
> that the paper contain heavily whitening agent,  a strong fluorescence
> emitter under UV excitation, which is toxic and may induce cancer.
>
> Sincerely,
> Peng Xi
> Ph. D.    Associate Professor
> Dept. of Biomedical Engineering, College of Engineering
> Peking University, Beijing, China
> Tel: [phone]
> Email: [hidden email]
> http://bme.pku.edu.cn/~xipeng
>

--
------------------------------------------------------------
Steffen Dietzel, PD Dr. rer. nat
Ludwig-Maximilians-Universität München
Walter-Brendel-Zentrum für experimentelle Medizin (WBex)
Head of light microscopy

Mail room:
Marchioninistr. 15, D-81377 München

Building location:
Marchioninistr. 27,  München-Großhadern

[Peng Xi-2]

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Dear Steffen,
    I am glad to answer your questions. See below line-by-line responses.
Thank you!
Peng

On Tue, Jun 19, 2012 at 8:47 PM, Steffen Dietzel <[hidden email]>wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/**wa?A0=confocalmicroscopy<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>
> *****
>
> I don't quite get it from the article. Assuming an upright confocal, you
> put, say, a chroma test slide under the preparation, ok.
>
> * How do you get to tilted/oblique illumination? Do you have to put the
> specimen to the edge of the scan range? Or is it done with moving a lense
> out of axis? (which would mean that this is not easily feasable on
> commercial setups since they don't intend to give access to the beampath)
>

In order to get tilted illumination, you have to either tilt the
galvonometer, or tilt the incident laser. Right now it is done with a
custom confocal setup. Since I do not have access to a commercial confocal
system in this level, I cannot evaluate how difficult it is to convert a
commercial system to do this.

>
> * Then you scan eg with 488 nm and record the green fluorescent signal
> from the test slide, is that it?
>
Yes, that's right. Keep  in mind that you have to be able to excite the
fluorescent medium to generate fluorescence illumination.

>
> * The actual image is then generated by the fluorescent light traveling
> back through the specimen and getting deflected (phase shifted ?) or not
> during the process, being finally recorded with a PMT.
>
Because the illumination is oblique, the phase-relief image can be
generated with the shift of collecting optics(lens+pinhole+PMT).

>
> * To get signal through you have to open the pinhole, since it is not a
> confocal signal, right?.
>
No, we have just tested that for AU=1, LOSOM still works, just as
theoretical predicted.

>
> Maybe you could clarify these points for the non-physicist.
>
> Steffen
>
>
>
> On 12.06.2012 01:25, Peng Xi wrote:
>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/**wa?A0=confocalmicroscopy<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>
>> *****
>>
>> Dear List,
>>       We have just published our work of achieveing DIC phase relief
>> image of transparent specimens with existing confocal setup, by just
>> adding a piece of plastic or paper (basically, anything that
>> fluorescent) on top of your sample. We also explained why so
>> mathematically. Link:
>>       http://www.opticsinfobase.org/**oe/abstract.cfm?uri=oe-20-13-**
>> 14100 <http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100>
>>
>>       Also, a piece of advice: do NOT place printing paper on your
>> lips when you open your door ---- although not presented, we noticed
>> that the paper contain heavily whitening agent,  a strong fluorescence
>> emitter under UV excitation, which is toxic and may induce cancer.
>>
>> Sincerely,
>> Peng Xi
>> Ph. D.    Associate Professor
>> Dept. of Biomedical Engineering, College of Engineering
>> Peking University, Beijing, China
>> Tel: [phone]
>> Email: [hidden email]
>> http://bme.pku.edu.cn/~xipeng
>>
>>
>
> --
> ------------------------------**------------------------------
> Steffen Dietzel, PD Dr. rer. nat
> Ludwig-Maximilians-Universität München
> Walter-Brendel-Zentrum für experimentelle Medizin (WBex)
> Head of light microscopy
>
> Mail room:
> Marchioninistr. 15, D-81377 München
>
> Building location:
> Marchioninistr. 27,  München-Großhadern
>

--
席鹏
特聘研究员
北京大学工学院生物医学工程系
地址：中关村北大街北京大学医院A536室
邮编：100084
电话：[phone]
Email:  [hidden email]
http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100


Sincerely,
Peng Xi
Ph. D.    Associate Professor
Dept. of Biomedical Engineering, College of Engineering
Peking University, Beijing, China
Tel: [phone]
Email: [hidden email]
http://www.opticsinfobase.org/oe/abstract.cfm?uri=oe-20-13-14100