Confocal Microscopy List - Re: Argon ion laser - age/noise and effect on images..?

Re: Argon ion laser - age/noise and effect on images..?

Posted by Arvydas Matiukas on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Argon-ion-laser-age-noise-and-effect-on-images-tp7586010p7586029.html

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Hi Jacqui,

I recently replaced a failed argon laser head after 6 years and about 5000 hrs of operation.

Before the failure I noticed higher tube current, lower output power and higher temporal

power variation (slow, on the minute time scale). Image quality was somewhat degraded

but not directly attributable to the laser noise (high frequency noise) and requiring significantly

slower scan and/or more averaging.

Based on my experience granularity of confocal images is related to the problems on the

detection path: noisy detector, degraded emission filter, beamsplitters, pinholes, and other components

depending on the design of your confocal. Especially when your laser output at focal

plane is in correct mW range. Additionally to other listers advice I suggest to run a simple scan

with (strong) backreflection of laser beam (from mirror slide or just coverslip edge)

and low detector voltage settings (manually configure

optics allowing the backreflected laser beam to reach the detector). That would diagnose

if low noise level images can be acquired.

Another potential problem outside the detection path may be AOTF failure as mentioned by Glen.

Our confocal experienced AOTF problems due to failing fan and AOTF overheating. I would

suggest to check that the ambient temperatures around/inside AOTF and laser head are

in the correct range and do not fluctuate.

Regards,

Arvydas

Arvydas Matiukas, Ph.D.
Director of Confocal&Two-Photon Core
SUNY Upstate Medical University
Neuroscience & Physiology Dept

Room 4607 IHP

505 Irving Ave

Syracuse, NY 13210

Email: [hidden email]

>>> Jacqui Ross <[hidden email]> 11/17/2016 10:55 PM >>>

Hi everyone,

One further comment which I forgot to include which may be very relevant. We replaced the power supply for the argon ion laser last year after experiencing some fluctuations including black lines in the image. Although I thought that lasers and power supplies came as a pair, I was told that we didn't need to replace the laser, only the power supply. If anyone else has knowledge about this, it would be very helpful.

Thanks to Patty who has advised me that she was also told that both needed to be replaced.

To Paul, Craig and Zdenek; thanks for your valuable suggestions for further testing. I will try these ideas next week.

Hi Rosemary, we do have our system serviced every year and the alignment is checked and should be OK as it was just serviced in September. However, I will go back to our engineer to check that. He also does the power level testing each time and I managed to get the historical data off him which is how I now know that it's dropped to 1/3 of its previous power level.

Cheers,

Jacqui

From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Jacqui Ross
Sent: Friday, 18 November 2016 1:20 p.m.
To: [hidden email]
Subject: Argon ion laser - age/noise and effect on images..?

Hi everyone,

I'm hoping that someone can help me to confirm whether an effect I am seeing with one of our confocal microscopes is likely to be due to the ageing of an argon ion multiline laser.

The laser is about 8 years old and the power level has dropped to about 1/3 of the original output but we are still able to excite the fluorophores very well at AOTF controlled percentages <5%.

What I'm noticing is that when I use the argon ion laser, e.g. 488nm line, the images look much more granular than when using other lasers when similar voltages are being used at the PMT. I've checked the other detectors and it doesn't seem to be related to the PMTs.

Much slower speeds and more averaging do improve the images but I'm puzzled by this as the issue is there even when the Master gain/voltage is at a level which I wouldn't normally need to go slower/average as much.

Can anyone tell me whether the age of the laser can cause issues like this? I'm wondering if we should just replace it. We only normally replace lasers when they expire.

Or if there are some tests that I can do to further define the issue?

The last measurements were as below for 8A with two different beam splitters;

488nm 0.51mW (NT 80/20) 2.31mW (MBS488)

Thanks in advance for any ideas.

Cheers,

Jacqui

Jacqueline Ross

Biomedical Imaging Microscopist
Biomedical Imaging Research Unit (BIRU)
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland 1142, NEW ZEALAND

Telephone: Ext 87438; DDI: +64 9 923 7438

Website: http://www.auckland.ac.nz/biru