Confocal Microscopy List

gated PMTs

Classic

List

Threaded

7 messages Options

Simon Wiegert

gated PMTs

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****

Dear Listers,

I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.

Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.

Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?

Any feedback is welcome.

Thanks, Simon

Simon Wiegert, PhD

Center for Molecular Neurobiology (ZMNH)

Institute for Synaptic Physiology

Falkenried 94

20251 Hamburg

Germany

Tel.: +49 40 7410 55085

simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

Steffen Dietzel

Re: gated PMTs

Simon,

with the gating it is my understanding that all the unwanted photons still arrive at the photocathode and still produce photoelectrons. Since the cycles are short, the voltage cannot be powered down during the gating phase (at least not in our hybrid photomultipliers), so secondary electrons are also produced. So all the bad things to damage the detector are there, they are just not counted during the gating. I am not a physicist nor an engineer, so I may be wrong on this. But I share your estimate that the detector may be doomed.

Many (20?) years ago Hans Tanke developed something that works like a rotating shutter in a normal fluorescence microscope. By the speed of the rotation nanosecond windows for excitation and emission could be selected to suppress background fluorescence in tissue sections, if I remember correctly. Maybe there is a thought for you.

Steffen

Am 01.11.2016 um 21:28 schrieb Simon Wiegert:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****

Dear Listers,

I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.

Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.

Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?

Any feedback is welcome.

Thanks, Simon

--

Simon Wiegert, PhD

Center for Molecular Neurobiology (ZMNH)

Institute for Synaptic Physiology

Falkenried 94

20251 Hamburg

Germany

Tel.: +49 40 7410 55085

simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

-- 
------------------------------------------------------------
Steffen Dietzel, PD Dr. rer. nat
Ludwig-Maximilians-Universität München
Biomedical Center (BMC)
Head of the Core Facility Bioimaging

Großhaderner Straße 9
D-82152 Planegg-Martinsried
Germany

http://www.bioimaging.bmc.med.uni-muenchen.de

Mark Cannell-2

Re: gated PMTs

In reply to this post by Simon Wiegert

Not quite clear what " photostimulation happening from various light sources that more or less point directly into the condenser” means but it sounds like you have other illumination wavelengths that are in the bandpass of the detector? Can this not be avoided in some way with suitable detector filters? You are right, old research on PMTs suggest that if turned off, exposure to bright light is well tolerated -the main effect being an increase in dark current that takes many hours to decline. Exposure to bright light while powered on causes a large permanent loss in sensitivity if cathode or anode current limits are exceeded. Fast protection circuitry can help save the detector but the damage seems to accumulate over time so plan replacements… I see no reasons why these concerns would not apply to Gaasps.

Hope this helps

Mark

On 1/11/2016, at 8:28 pm, Simon Wiegert <[hidden email]> wrote:

Mark B. Cannell Ph.D. FRSNZ FISHR
Professor of Cardiac Cell Biology
School of Physiology & Pharmacology
Faculty of Biomedical Sciences
University of Bristol
Bristol
BS8 1TD UK

[hidden email]

Benjamin Smith

Re: gated PMTs

In reply to this post by Simon Wiegert

Unfortunately, as far as hybrid PMTs go, this is a case of manufacturers using the same term to describe two things.

Because of capacitance and inductance alone, it is highly prohibitive to gate a high-voltage potential in microsecond time frames, and to do so would require large currents, which you are trying to avoid.

In gated PMTs, the gating is done by switching the focusing potential between the photocathode and the dynodes. Since this potential is only about 10V, it can be rapidly switched. With the potential flipped, the photoelectrons are scattered away from the dynodes, protecting the dynodes from what would otherwise be a large current. By switching the focusing polarity, you can protect the dynodes while keeping them at a stable high voltage.

In a hybrid detector, gating refers to synchronizing the photon counting to a pulsed laser. In this case, the detector is always on detecting photoelectrons, and what you are gating is the detection clock, basically a start and stop time when you want to actually count the photoelectrons vs ignore them. Since autofluorescense has a relatively short fluorescence lifetime, while fluorophores have longer lifetimes, by waiting a short time after each laser pulse, you can enrich for detecting just the fluorophores.

In short, in a gated PMT, the photocathode always sees the photons, but the gate protects the dynodes from the photoelectrons, protecting the rest o the PMT. in a hybrid detector, the entire detector circuit sees all of the photoelectrons, and the gating simply tells the counting circuit which counts to ignore.

According to Hamamatsu, it sounds like the GaAsP photocathode is pretty resilient, because the guide says to avoid direct intensity light such as direct laser light, which is pretty extreme when you thing about normal light levels a PMT encounters.

For more reading on the topic, this is a good guide: http://psec.uchicago.edu/links/pmt_handbook_complete.pdf

Hope this helps,

Ben Smith
Sent from my iPhone

On Nov 1, 2016, at 1:28 PM, Simon Wiegert <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Dear Listers,
I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.
Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.
Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?
Any feedback is welcome.
Thanks, Simon

--
Simon Wiegert, PhD
Center for Molecular Neurobiology (ZMNH)
Institute for Synaptic Physiology
Falkenried 94
20251 Hamburg
Germany
Tel.: +49 40 7410 55085
simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

Mark Cannell-2

Re: gated PMTs

I don’t think he’s asking about hybrid detectors but Gaasp PMTs. By the way, the cathode to d1 voltage is typically 100V and this voltage does not focus the photoelectrons, it accelerates them into the first dynode to cause secondary emission.

Regards Mark

On 2/11/2016, at 7:52 pm, Benjamin E Smith <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Unfortunately, as far as hybrid PMTs go, this is a case of manufacturers using the same term to describe two things.

Because of capacitance and inductance alone, it is highly prohibitive to gate a high-voltage potential in microsecond time frames, and to do so would require large currents, which you are trying to avoid.

In gated PMTs, the gating is done by switching the focusing potential between the photocathode and the dynodes. Since this potential is only about 10V, it can be rapidly switched. With the potential flipped, the photoelectrons are scattered away from the dynodes, protecting the dynodes from what would otherwise be a large current. By switching the focusing polarity, you can protect the dynodes while keeping them at a stable high voltage.

In a hybrid detector, gating refers to synchronizing the photon counting to a pulsed laser. In this case, the detector is always on detecting photoelectrons, and what you are gating is the detection clock, basically a start and stop time when you want to actually count the photoelectrons vs ignore them. Since autofluorescense has a relatively short fluorescence lifetime, while fluorophores have longer lifetimes, by waiting a short time after each laser pulse, you can enrich for detecting just the fluorophores.

In short, in a gated PMT, the photocathode always sees the photons, but the gate protects the dynodes from the photoelectrons, protecting the rest o the PMT. in a hybrid detector, the entire detector circuit sees all of the photoelectrons, and the gating simply tells the counting circuit which counts to ignore.

According to Hamamatsu, it sounds like the GaAsP photocathode is pretty resilient, because the guide says to avoid direct intensity light such as direct laser light, which is pretty extreme when you thing about normal light levels a PMT encounters.

For more reading on the topic, this is a good guide: http://psec.uchicago.edu/links/pmt_handbook_complete.pdf

Hope this helps,
Ben Smith
Sent from my iPhone

On Nov 1, 2016, at 1:28 PM, Simon Wiegert <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Dear Listers,
I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.
Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.
Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?
Any feedback is welcome.
Thanks, Simon

--
Simon Wiegert, PhD
Center for Molecular Neurobiology (ZMNH)
Institute for Synaptic Physiology
Falkenried 94
20251 Hamburg
Germany
Tel.: +49 40 7410 55085
<a href="x-msg://81/simon-wiegert.com" style="color: rgb(149, 79, 114); text-decoration: underline;">simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

Mark B. Cannell Ph.D. FRSNZ FISHR
Professor of Cardiac Cell Biology
School of Physiology & Pharmacology
Faculty of Biomedical Sciences
University of Bristol
Bristol
BS8 1TD UK

[hidden email]

Benjamin Smith

Re: gated PMTs

From the Hamamatsu handbook (p. 97): http://psec.uchicago.edu/links/pmt_handbook_complete.pdf

   If the photomultiplier tube output is OFF at a gate input of 0V, a reverse bias of about 10 volts with respect
   to the focusing electrode and first dynode is supplied to the cathode. This prevents photoelectrons, if emitted
   by the cathode, from reaching the dynode section. Here, if a pulse signal of +3 to +4 volts is applied to the gate
   input terminal, the driver circuit gives a forward bias to the cathode via capacitance coupling, and sets the
   photomultiplier tube output to ON during the period determined by the gate pulse width and the time constant
   of the capacitance-coupled circuit. This gating circuit provides a switching ratio (or extinction ratio) of 10^4 or
   more. The capacitors are connected from the first through the center dynode to absorb the switching noises
   often encountered with this type of gating circuit.

I made a quick mock-up of the circuit schematic: http://tinyurl.com/jkzg5uf

I know the capacitors and resistors aren't the exact values, but the point is more to show the potential switching behavior at the photocathode. As you can see, if the TTL signal is high (switch closed) then there is a negative potential from the cathode to the focusing anode/dynodes, which turns the PMT "ON" (electrons head toward the dynodes). If the TTL signal is low (switch open), then the polarity of the potential between the cathode and focusing anode/dynodes gets flipped, without effecting any of the potential of the dynodes themselves, which turns the PMT "OFF" (electrons stay at the photocathode). The MOSFETs are there simply as a crude surrogate for the PMT current, where negative dynode potentials drive the PMT current

Cheers,

Ben Smith

On Wed, Nov 2, 2016 at 3:37 PM, Mark Cannell <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Benjamin,

I don’t think he’s asking about hybrid detectors but Gaasp PMTs. By the way, the cathode to d1 voltage is typically 100V and this voltage does not focus the photoelectrons, it accelerates them into the first dynode to cause secondary emission.

Regards Mark

On 2/11/2016, at 7:52 pm, Benjamin E Smith <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Unfortunately, as far as hybrid PMTs go, this is a case of manufacturers using the same term to describe two things.

Because of capacitance and inductance alone, it is highly prohibitive to gate a high-voltage potential in microsecond time frames, and to do so would require large currents, which you are trying to avoid.

In gated PMTs, the gating is done by switching the focusing potential between the photocathode and the dynodes. Since this potential is only about 10V, it can be rapidly switched. With the potential flipped, the photoelectrons are scattered away from the dynodes, protecting the dynodes from what would otherwise be a large current. By switching the focusing polarity, you can protect the dynodes while keeping them at a stable high voltage.

In a hybrid detector, gating refers to synchronizing the photon counting to a pulsed laser. In this case, the detector is always on detecting photoelectrons, and what you are gating is the detection clock, basically a start and stop time when you want to actually count the photoelectrons vs ignore them. Since autofluorescense has a relatively short fluorescence lifetime, while fluorophores have longer lifetimes, by waiting a short time after each laser pulse, you can enrich for detecting just the fluorophores.

In short, in a gated PMT, the photocathode always sees the photons, but the gate protects the dynodes from the photoelectrons, protecting the rest o the PMT. in a hybrid detector, the entire detector circuit sees all of the photoelectrons, and the gating simply tells the counting circuit which counts to ignore.

According to Hamamatsu, it sounds like the GaAsP photocathode is pretty resilient, because the guide says to avoid direct intensity light such as direct laser light, which is pretty extreme when you thing about normal light levels a PMT encounters.

For more reading on the topic, this is a good guide: http://psec.uchicago.edu/links/pmt_handbook_complete.pdf

Hope this helps,
Ben Smith
Sent from my iPhone

On Nov 1, 2016, at 1:28 PM, Simon Wiegert <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Dear Listers,
I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.
Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.
Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?
Any feedback is welcome.
Thanks, Simon

--
Simon Wiegert, PhD
Center for Molecular Neurobiology (ZMNH)
Institute for Synaptic Physiology
Falkenried 94
20251 Hamburg
Germany
Tel.: <a href="tel:%2B49%2040%207410%2055085" value="+4940741055085" target="_blank">+49 40 7410 55085
simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

Mark B. Cannell Ph.D. FRSNZ FISHR
Professor of Cardiac Cell Biology
School of Physiology & Pharmacology
Faculty of Biomedical Sciences
University of Bristol
Bristol
BS8 1TD UK

[hidden email]

Benjamin E. Smith, Ph. D.
Microscopy Specialist, Vision Science
University of California, Berkeley
142 Life Sciences Addition
Berkeley, CA 94720-3200
Tel (510) 642-9715
Fax (510) 643-6791
e-mail: [hidden email]
http://vision.berkeley.edu/

Simon Wiegert

Re: gated PMTs

Dear Ben and others,

thank you very much for your detailed responses. Yes, I was referring to gated GaAsPs, which I want to switch “off” for some ms during light pulses directly applied to the sample for optogenetic photostimulation. As I somewhat expected, the electronics beyond the photocathode are protected ok. As you noted, the photocathode itself might be quite resilient – at least, according to Hamamatsu. Nonetheless, the statement they make regarding avoiding direct laser light is not very quantitative. This is why I was asking if anyone has some real-world experience with such experiments. Even if the photocathode does not get destroyed completely, it might lose sensitivity over time. As far as I know, GaAsP detectors cannot recover as efficiently as bi-/multialkaline detectors.

Best, Simon

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Benjamin E Smith
Sent: Donnerstag, 3. November 2016 04:09
To: [hidden email]
Subject: Re: gated PMTs

Cheers,

Ben Smith

On Wed, Nov 2, 2016 at 3:37 PM, Mark Cannell <[hidden email]> wrote:

Benjamin,

Regards Mark

On 2/11/2016, at 7:52 pm, Benjamin E Smith <[hidden email]> wrote:

For more reading on the topic, this is a good guide: http://psec.uchicago.edu/links/pmt_handbook_complete.pdf

Hope this helps,

Ben Smith
Sent from my iPhone

On Nov 1, 2016, at 1:28 PM, Simon Wiegert <[hidden email]> wrote:

***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy Post images on http://www.imgur.com and include the link in your posting. *****
Dear Listers,
I guess this question was discussed here previously, but I could not find any recent discussion on this topic. I am planning to set up a two-photon imaging microscope with forward detection through the condenser. Since there will be a lot of photostimulation happening from various light sources that more or less point directly into the condenser, the PMTs need to be protected. Until now we did it with Uniblitz shutters. However, there are some issues with these shutters (speed, noise, etc.) and I would rather like to use gated PMTs.
Does anyone have long-term experience with gated PMTs and (strong) light exposure? I would like to use GaAsPs and I am afraid that a lot of light might still kill the PMT since there is no physical light block in front of it. In my understanding, excessive light can still destroy the photocathode, i.e. the light-sensitive surface, even if the anode current is strongly suppressed by the gating circuit.
Does anyone have experience or even a systematic comparison with gated GaAsP PMTs that had been exposed to light vs. some non-exposed?
Any feedback is welcome.
Thanks, Simon

--
Simon Wiegert, PhD
Center for Molecular Neurobiology (ZMNH)
Institute for Synaptic Physiology
Falkenried 94
20251 Hamburg
Germany
Tel.: <a href="tel:%2B49%2040%207410%2055085" target="_blank">+49 40 7410 55085
simon-wiegert.com

--
Pflichtangaben gemäß Gesetz über elektronische Handelsregister und Genossenschaftsregister sowie das Unternehmensregister (EHUG):

Universitätsklinikum Hamburg-Eppendorf
Körperschaft des öffentlichen Rechts
Gerichtsstand: Hamburg

Vorstandsmitglieder:
Prof. Dr. Burkhard Göke (Vorsitzender)
Prof. Dr. Dr. Uwe Koch-Gromus
Joachim Prölß
Rainer Schoppik

Mark B. Cannell Ph.D. FRSNZ FISHR
Professor of Cardiac Cell Biology
School of Physiology & Pharmacology
Faculty of Biomedical Sciences
University of Bristol
Bristol
BS8 1TD UK

[hidden email]