Hello from Toronto

[wjuanxp]

Hi All,

So nice to find this forum and sorry if my English isn't perfect. I'm an analytical chemist in my late thirties. I always love to see small things, in fact, this is the reason for me to get into analytical chemistry, so I can "see" mulecules. At work, I work with mass spectrometer to see organic molecules in samples. Since it costs thousands of dollars and very noisy plus consumed tones of electricity, it is impossible to have one at home, so I thought a microscope might full fill my cravings on seeing small stuff in a different way. Last year, I was lucky to get a Leica DMIRB inverted microscope from an auction for 800 USD with flourecent ablity and three phase contrast objectives. It brought me and my three years old son a lot of joy during summer time trying to find something to see in our backyard. I'm currently trying to find the best options to get digital photography capability on our scope and please give us some advices if you can.

I'll post more pics and try to complete my profile in the near future.

[billbillt]

WELCOME!...

BillT

[zzffnn]

Welcome to the forum, wjuanxp!

I would be surprised to see a hobbyist with a mass spec at home. But you may rent user time from an electron microscope from some major universities. Member Robert Berdan from Calgary may have done that or is going to do so.

For a 3yo, I would suggest a dissecting macroscope (also called stereo microscope). My 4yo and 8yo prefer my macroscope over my high power compound microscopes and use it to look at seashells and insects.

Any macroscope would work, though I would recommend Bausch and Lomb StereoZoom (B&L SZ7). It is of very good optical quality, more than powerful enough for hobbyists and can be bought for less than $100 sometimes (due to its wide availability). I like mine.

You may want a camera with electronic (silent vibration -free) shutter to take great pictures on microscope. For fluorescence though, a dedicated cooled fluorescence camera may be better, but most consumer cameras with large sensors would work as well.

Note HBO lamps of fluorescence microscopes may blow up, if not well maintained.

[wjuanxp]

Welcome to the forum, wjuanxp!

Note HBO lamps of fluorescence microscopes may blow up, if not well maintained.

Thanks for you advices on digital solutions. What kind of maintenance do I need to do for HBO lamps? Is there a place to read more about it?

[zzffnn]

I would suggest using UV LED, instead of HBO (I don't know how to maintain HBO lamps).

"search all words" "LED fluorescence" at
http://www.photomacrography.net/forum/s ... de=results

Here is an example thread:
http://www.photomacrography.net/forum/v ... sc&start=0

[wjuanxp]

I would suggest using UV LED, instead of HBO (I don't know how to maintain HBO lamps).

"search all words" "LED fluorescence" at
http://www.photomacrography.net/forum/s ... de=results

Here is an example thread:
http://www.photomacrography.net/forum/v ... sc&start=0

That's cool! I might be able to make a LED light source like that too. Thanks for all those useful info.

[Hobbyst46]

I would suggest using UV LED, instead of HBO (I don't know how to maintain HBO lamps).

"search all words" "LED fluorescence" at
http://www.photomacrography.net/forum/s ... de=results

Here is an example thread:
http://www.photomacrography.net/forum/v ... sc&start=0

That's cool! I might be able to make a LED light source like that too. Thanks for all those useful info.

That example thread is a great display of a DIY fluorescence illumination setup. Note, though, that for chlorophyll fluorescence, UV is not the best wavelength, and neither is blue (450-460nm). Rather, the optimal wavelength is 400-430nm, i.e. violet light. Also, just for chlorophyll, an excitation filter is not a must.
About HBO: handling and alignment of high pressure Hg/Xe lamps requires appropriate safety measures. These bulbs have a limited lifetime, especially upon frequent on/off cycles.

[wjuanxp]

I would suggest using UV LED, instead of HBO (I don't know how to maintain HBO lamps).

"search all words" "LED fluorescence" at
http://www.photomacrography.net/forum/search.php?mode=r

Here is an example thread:
http://www.photomacrography.net/forum/v ... sc&start=0

That's cool! I might be able to make a LED light source like that too. Thanks for all those useful info.

That example thread is a great display of a DIY fluorescence illumination setup. Note, though, that for chlorophyll fluorescence, UV is not the best wavelength, and neither is blue (450-460nm). Rather, the optimal wavelength is 400-430nm, i.e. violet light. Also, just for chlorophyll, an excitation filter is not a must.
About HBO: handling and alignment of high pressure Hg/Xe lamps requires appropriate safety measures. These bulbs have a limited lifetime, especially upon frequent on/off cycles.

Thanks for your valuable input. Regarding to excitation wavelength, i know it depends on the molecule you want to excited, but which wavelength is the most commonly used in microscopy? i might want to do an open source project on LED light source for microscopy (both white light and narrow-band) if I have sometime this summer. I'll let you guys know if I decided to do it later.

[Hobbyst46]

...Regarding to excitation wavelength, i know it depends on the molecule you want to excited, but which wavelength is the most commonly used in microscopy?

1. Decades ago, I think it was ~490nm and ~560nm, for fluorescein and rhodamine, respectively, since these dyes were very popular stains. But the number of biological stains has grown exponentially, as well as fluorescent molecules that are artificially introduced into organism (fluorescent proteins etc). They are selected by researchers to serve as specific indicators (say, cell viability) - and other purposes (even to kill cancer cells). The number of stains far exceeds the number of available LED wavelengths, so a single LED will have to serve several dyes. This is possible since the spectral excitation peak is not narrow.

2. Auto-fluorescence in plants: for example chlorophyll but also vitamin E and similar molecules. I would like to clarify, UV (365-385nm) does excite autofluorescence, of those molecules for example, my point was that for chlorophyll it is not the best wavelength.

3. There are various other fluorescence sources, such as minerals.

i might want to do an open source project on LED light source for microscopy (both white light and narrow-band) if I have sometime this summer. I'll let you guys know if I decided to do it later.

There are commercial LED lamps for fluorescence microscopy. They are quite expensive (not the chips - the complete lamp). AFAIK they are monochromatic, as much as LEDs can be, peak wavelength +/- a few tens of nanometers. As for white light LED, I would suspect that the intensity at a specific excitation wavelength would be low, in comparison to the intensity derived from Hg/Xe lamps for example, but I could be wrong on this. Research-level confocal fluorescence microscopy is based on lasers, not LEDs, to provide a sufficiently high intensity at a (highly monochromatic) wavelength.

4. Nevertheless, with the following combination:
(a) powerful white light LED, especially cool white (which contains strong blue light around 430-450nm),
(b) efficient optical filtration setup, that rejects the excitation and lets only the emission reach the eyepiece and camera
(c) high concentration of fluorophore in the specimen,
(d) darkness around the microscope, to avoid masking by ambient light,
(e) sensitive camera for low-level light,
at least some fluorescence will be visible.