Notes on adapting a Nikon Labophot 1 stand for LED use.

by David Walker, UK

I recently had extended access to a Nikon Labophot 1 stand / optics; this stand with various outfits crops up quite regularly on eBay and is illustrated and described on Nikon's excellent MicroscopyU Microscopy Museum website. An aspect of the stand I found particularly useful was the ease of adapting this model's lamphouse to LEDs, and its design allowed a rapid interchange during microscopy studies to LEDs of different visible and near IR wavelengths. Other maker's models with similar lamphouses may also prove suitable (but note comments on electrical aspects below).

The ease of adapting the Labophot lamphouse is summarised in the annotated images below. A spare lamphouse is dedicated to LEDs and the original quartz halogen lamphouse can be reinstalled whenever needed. The LED can be changed out to study the effect of varying the wavelength on imaging a subject, e.g. diatoms for shorter wavelengths or dense insect exoskeletons for longer wavelengths.

Why use LEDs? Glass filters are of course quick and easy to place on the field iris but I find that the near monochromatic LEDs give superior results to a non specialised glass/acetate filter and the LEDs cost very little.

There's two diffusers on the Labophot 1, one after the lamp and another after the field condenser which reduce the available light with the supplied 6V / 20W lamp and even superbright LEDs for techniques like phase at higher mags. Immersion oiling a large coverslip to the accessible rear diffuser made it clear for more light throughput; and it seemed to have no effect on the evenness of light and is easily reversible if needed. (The lamphouse was uprated on the Labophot 2 but the lamphouse is bulkier).

Safety note: Please see safety footnote below if using high intensity visible LEDs and particularly if considering LEDs outside of the visible spectrum.

Diatom Frustulia rhomboides, Klaus Kemp 'Test plate 8 forms'. Hyrax. Blue 470 nm LED. Nikon oil NA 1.2 - 1.43 darkfield condenser, Zeiss 40x NA1.0 oil objective. Nikon D50 DSLR, 2 sec exposure. Zeiss 4:1 projection eyepiece.
LED details: Nichia NSPB500S, 5 mm envelope, 4.4 Candelas, 15 degree beam, (www.maplin.co.uk Maplin code NR85, £3.34). (LEDs can be up to 10x cheaper on eBay but the maker and full specifications may not always be supplied, which is important to know at short wavelengths to check if UV present.)

Diatom Pleurosigma angulatum, Klaus Kemp 'Test plate 8 forms'. Hyrax. Blue 470 nm LED (no UV component). Nikon oil NA 1.2 - 1.43 darkfield condenser, Zeiss 40x NA1.0 oil objective. Nikon D50 DSLR, 2 sec exposure. Zeiss 4:1 projection eyepiece.

Diatom Navicula lyroides, Klaus Kemp test plate. Zrax. Two image stack. Green LED. Nikon condenser, Zeiss 25x NA0.60 objective. Nikon D50 DSLR, 1/2 sec exposure. Zeiss 4:1 projection eyepiece. (LED 5 mm, 20 degree beam, 525 nm, 15 Candelas,  eBay sourced £2.99 for 10.)

 

The high NA 1.2 - 1.43 of the Nikon darkfield condenser also allowed a variation of the popular 'COL' technique to be tried, i.e. circular oblique lighting by using water not oil immersion with the condenser. This gave a narrow ring of light just inside an NA 1.3 objective's back focal plane, thus exploiting its near full NA. Coupled with the blue LED 470 nm, the combination suited some test diatom studies.

This is a technique Ted Clarke (USA) encouraged me to try with this condenser as he has used it to good effect with a LOMO darkfield oil immersed NA 1.2 condenser with a LOMO NA 1.3 apo objective. (Water is required for the higher NA of the Nikon condenser). Paul James has an excellent suite of Micscape articles on many aspects of COL.

The technique does have a low tonal range cf. conventional oblique, so tonal balance would normally be adjusted. See examples below using this technique.

 



High NA COL: Above: Amphipleura pellucida. Resolving the transverse striae (seen vertically in above images) to punctae is regarded to be near the limit of light microscope resolution. With high NA COL a Zeiss 100x NA 1.3 Neofluar objective is close to punctae resolution, LED 470 nm (blue, no UV). The full annular ring of light doesn't stretch the camera's sensitivity as much as conventional directional oblique may do.
Left: Victorian mount strew by 'S S Pilgram North Wales, Hyrax'. Specimens typically 140 µm long. Out of camera image crop.
Right: Klaus Kemp '8 form test plate', Hyrax. Specimen 90 µm long. Out of camera image crop with slight tonal balance adjust.
Opticstar PL-130M monochrome camera.

 

 

Near infra red: Cinnabar moth head, Nikon 4x NA0.1 objective, NBS slide. Left: visible light, right - near IR LED 880 nm.
(LED details: Siemens SFH487P, 3 mm envelope, typically £0.88, 130 degree beam - for low power work, I prefer a wide beam).

Near IR is useful for studying subjects where the loss of resolution from the longer wavelength isn't important e.g. the gross structure of the dense exoskeletons of some insects. There's no visible image of course with near IR (and eyepieces shouldn't be used) but the live view of the camera allows all microscopy work.

 

Comments to the author David Walker are welcomed.

Acknowledgements:
Many thanks to Klaus Kemp for his excellent diatom test plates and type slides (available from his website www.diatoms.co.uk), and for making them so affordable to the hobbyist.

Also thanks to Ted Clarke for personal emails where he has shared his skills and for encouraging me to try the high NA COL technique. He has shared his use of the LOMO stand and its NA1.2 darkfield oil immersion for COL with a 90x NA1.3 LOMO apo in a recent presentation at Inter / Micro 2007 entitled 'Using the E330 DSLR camera with compound and stereo microscopes'. The use of COL formed part of the presentation where he showed the ability of the 'live view' mode of the Olympus E330 DSLR to fine focus difficult subjects, e.g. the striae of Amphipleura pellucida on the camera's LCD screen. Update Aug. 15th. Ted Clarke has kindly allowed Micscape to host a copy of his Powerpoint presentation of his Inter / Micro lecture and can be dowloaded here, (file 3.6 Mbytes).

Safety notes: From the author's hobbyist perspective, trying to establish what is a safe eye exposure off and on 'scope from both an intensity / wavelength perspective for the many LEDs on the market can be complex, so it's best to err on the side of extreme caution with high intensity LEDS and especially any with wavelengths approaching or exceeding the boundaries of the visible light spectrum (near UV and near IR); we've only got one pair of eyes!

On a microscope a live view webcam and computer screen can become your eyes and with common sense and awareness of an LED maker's guidelines and those from other authorities, high power LEDs can be and are being used safely by many hobbyists but not suitable for unsupervised youngsters.

General guidelines, please see the articles linked to:

Modern visible light LEDS and those required to give a good camera sensitivity on a microscope at high mags and using 'light hungry techniques' can be very bright and potentially dangerous to view directly both on and off 'scope, but especially off 'scope as they are point sources.

Apparently the eye becomes increasingly insensitive to shorter or longer wavelength extremes of the visible spectrum so visible light LEDs such as blue (no UV) need to be used with care. (For eye sensitivity to colour, see 'The joy of physical perception' an online book by Peter Kaiser.)

This article High-Power LEDs Pose Safety Hazards on the Photonics website by G R Davies is invaluable reading. (With thanks to the Yahoo Microscope group members for highlighting it.)

My own view is err on the safe side and do all LED microscopy studies with potential safety queries with a webcam, which often have good sensitivity to shorter wavelengths (and longer wavelengths if no IR blocking filter) and can be used entirely for all studies after setting up in brightfield.

Check the maker's spec sheet for short wavelength LEDs e.g. deep blue / violet to be aware if any near UV component.  High power near IR LEDs also need caution from inadvertent viewing with eyes.

An LED with any UV component must not be used for visual work but only with a suitable camera with direct stray light from the microscope blocked. Near IR should also be used only with a webcam. (A maker's spec sheet (ETG-5UV400-30) for a 0.7 Candela 400 nm LED i.e. on the near UV / visible light border, has warnings of even brief eye exposure.)

Disclaimer: The author, site contributors, and site owner (of Microscopy-UK / Micscape, OnView.net) accepts no responsibility for any eye or other damage by using LEDs. It is up to the user to adopt appropriate safety precautions for an LED's intensity and/or wavelength as recommended by the maker or other suitable authorities.

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