Equipment for microscopy

First trial of centrifuge on pond water 14/12/2017

“First Light” for my centrifuge – the second hand Eppendorf 5415c. I followed a procedure detailed on the Micscape website for using the centrifuge to concentrate the organisms in a pond water sample from Stowe Pool in Lichfield collected today.

The centrifuge worked well and did indeed concentrate the organisms (see photos below for procedure I followed) although the variety was limited in the sample.

Andy

1-The-Eppendorf-5145c-centrifuge

2-Pond-water-sample-including-piece-of-plant-root

3-Stirring-sample-to-release-organisms-into-water

4-Filtering-out-large-particles using tea strainer

5-Filling-Ependorf-1-5ml-tubes-with-pond-water – each one needs to have same amount fluid in order for centrifuge to balance:

6-Placing-Eppendorf-tubes-in-centrifuge- note that they are evenly spread around centrifuge to obtain balance at high G-forces – you should always use pairs and never one on its own:

7-Screwing-on-airtight-lid-onto-rotor:

8-Centrifuged-samples-on-homemade-stand:

9-Pipetting-off-excess-fluid-after-pellet-the idea is to carefully remove the fluid above the pellet of centrifuged solids at bottom of tube without disturbing the pellet. Ideally, the pellet and single drop fluid left after pipetting.

10-View-of-pellet-at-bottom-of-Eppendorf-tube. after pipetting – it is the brown spot at bottom of tube in picture. I used pipette to transfer the pellets and small amounts of fluid from 6 tubes to 2 tubes and centrifuged again to further concentrate the contents into two pellets:

11-Slide-after-pellet-and-drop-water-pipetted-onto-it-showing-concentrated-protozoa & debris. I pipetted the final 2 pellets onto a slide and the picture below shows many dark areas in the water drop – these are from the pellets:

12 Coverslip & nail varnish to seal it before viewing under microscope:

Photos of organism from sample today seen under Zeiss IM microscope x32 magnification. The first photo shows well one particular feature evident in many of the photos – which is that the centrifuge has caused chloroplasts to be concentrated on one side of the cell rather than spread throughout. Other photos show large vacuoles which I think are from damage to the cell. Most of the cells were inactive – I presume killed by the process, although the banana-shaped ciliated organism in the third photo below was alive and moving.

 

Eppendorf 5415 C Micro-Centrifuge Variable Speed with Timer – photos & manual

An old but sturdy piece of kit – useful for microscopy – allows samples to be concentrated (e.g. pond water samples) making it easier to find targets of interest. Only problem is that the timer does not work so I need to source one of those.

I have found a replacement timer – I intend to ask Ed whether he can take a look with me to see if we can work out how to replace it – not asked him yet – help please, Ed!
I also picked up a spare rotor – the screw on top was missing from mine.
All these problems with second hand stuff – its cheap but often needs work to get it working – but then that is the fun of the thing.
Below are photos of the centifuge, spare rotor and new timer.

Click below to read the manual for this machine (PDF file):

Centrifuge_Eppendorf_5415C manual

Andy

Diehl Series 600 T85 springwound timer – finding a replacement timer for the Eppendorf 5315c micro-centrifuge

Diehl Series 600 T85 springwound timer – finding a replacement for the 5315c centrifuge

Is this the correct one?

https://www.ebay.co.uk/itm/Poltic-or-Diehl-Springwound-Timer-30-MIN-Tanning-Bed-Type-600-T85-with-BELL/371656358069?epid=1376092377&hash=item568871f8b5:m:mjcQ-kmGpI3cfy5GJ-pL_oQ

or is this one a better choice?

https://www.ebay.co.uk/itm/Poltic-or-Diehl-Springwound-Timer-30-MN-Tanning-Bed-Type-600-HOLD/391491109081?_trkparms=aid%3D222007%26algo%3DSIM.MBE%26ao%3D2%26asc%3D49479%26meid%3Dac3b581f3858496789ad0cb084fb3974%26pid%3D100623%26rk%3D2%26rkt%3D6%26mehot%3Dag%26sd%3D371656358069&_trksid=p2047675.c100623.m-1

or this one?

https://spasandmore.com/catalog/timers/poltik-t85-typ-600-30-min-twist-timer-with-bell-8302.html

or possible UK source:

http://www.plccenter.co.uk/Buy/DIEHL/T85600?redirect=true

 

Attaching Zeiss 100 W illuminator to epi-illumination port on Zeiss IM microscope

The following photos show the (now working as it has a new bulb) Zeiss 100 W illuminator attached to the epi-illumination port, and light coming through into the base of the objective turret through the epi-illumination port. Unfortunately, this is not sufficient to use epi-illumination yet. I have a flourescence cube holder which fits into the space under the objective turret and directs light up into the objective lens. This currently only has flourescence filters on it, which remove most of the light, seriously dimming the image. Hence I need to modify that cube holder to allow all light to be re-directed without filtering. I think this will involve purchasing a purpose-cut mirror…..but I still need to look into it.

Andy

Changing bulb on Zeiss 100 W halogen illuminator (Zeiss 40 80 19 illuminator)

Today, I changed the bulb on this illuminator, having picked up a couple of these halogen bulbs cheap from ebay. The process I went through to change the bulb is shown in the series of pictures below.

Andy

The 100 W Zeiss illuminator, before changing the bulb:

 

How to take the lamp holder out from the illuminator:

The bulb pulls out of its holder – but be careful – fragile and could easily break.

The bulbs purchased from ebay:

Replacing the bulb:

Final illuminator setup with working bulb:

 

Zeiss binocular head 47 30 11 9901

The binocular head I currently use on my Zeiss IM microscope is the Zeiss binocular head 47 30 11 9901. The dovetail is 43mm in external diameter.

I have included two photos of the IM microscope at the bottom of the page for colour comparison purposes – one taken using flash and one without. The binocular head is the same colour as the microscope. The Zeiss IM microscope is similar to the Zeiss IM35 microscope – in fact I have an IM35 as well and both are the same colour and use the same heads. Therefore, the colour of this microscope can be also seen on photos of the IM35 on the internet and many photos of the IM35 can be seen at this search link from Bing search engine:

https://www.bing.com/images/search?q=zeiss%20im35%20microscope&qs=n&form=QBIR&sp=-1&pq=undefined&sc=0-21&sk=&cvid=3A82742C156A4208ADB40A81ED1C4FD6

Andy

         

 

 

Microscopy S annectans Finis Shale Texas x4 objective Zeiss IM microscope 26/11/2017

Damian popped around and together we looked at a slide of fossilised coral from the USA.

Stereocorypha is a form of extinct coral. The slide here is from Jack County, Texas. Photographs were taken on my Zeiss IM microscope with x4 objective, using the Bresser Mikrocam 9.0 camera and Diagnostic Instruments adapter and ZU clamp, with Zeiss 910137 dual observation adapter (originally designed for a teaching microscope but bought into use here to allow both the binocular head and ZU clamp to be simultaneously used on the IM microscope).

The following slides show:

  • Three images of the same field of view to demonstrate the (limited) effect on this slide of using a single polarization filter. The filter used is one made by Zeiss.
  • A composite of 49 pictures combined using Microsoft’s Image Composite Editor.

Andy & Damian

Bresser Mikrocam 9.0 installed on Zeiss IM microscope:

The photos in the composite were taken using a blue filter. The set of three were the only ones using polarising filter – in this case one made by Zeiss. This is single linear polarisation NOT cross-polarisation:

 

A set of three images to demonstrate effect of polarisation on the same field of view:

Without polarising filter:

Polarising filter position I:

Polarising filter position II:

Composite of 49 images taken using blue filter (non-polarised):

This file is so large I have had to ZIP file into a compressed folder. The picture within it is a 70% quality JPEG version of the composite – the only way I could get such a large photo onto WordPress!

S annectans Finis Shale Texas x4 obj Zeiss IM 261117 (stitched x49 photos)JPEG-70percent-cropped

 

 

Trying to use Canon EOS 300D camera with Zeiss IM microscope

Today, I spent an hour or so trying to take photographs with my Canon EOS 300D on the Zeiss IM microscope…….and concluded that it is far too much hassle to be worth the effort – better off sticking with my dedicated Bresser MikrOkular and Mikrocam cameras with the microscopes.

Anyway, here is the account of my attempt to use the EOS with the microscope.

Firstly – an adapter issue – needed an EOS to microscope adapter – could not find one so resorted to buying an EOS-T2 adapter and a T2-C-mount adapter and pairing the two – some vignetting but not a major issue. This allowed me to mount the camera as in the picture below, using the Diagnostic Instruments ZU clamp (the ZU has correct dovetail for the IM and IM35 microscopes I have found).

I tried to directly download photos from camera to WIndows 7 computer but then ran into my next problem – no disc with my EOS 300D (bought few years ago without CD) and I can’t download software from Canon or find it on internet.

Also driver problem with Windows 7 – Canon driver does not support Windows 7 – found a workaround for Windows 7 – BUT it did not work on my laptop – don’t know why – involved changing communication settings on a Canon EOS camera menu.

Click download to view workaround for Windows 7 (PDF file):

Workaround Canon 300D with Windows 7 accessed 26/11/17

The alternative is to use a card reader – this does work although means card needs to be taken in and out – not good for helping with focusing where many quickly viewed images needed.

In spite of all of the above, I could work around these problems but the next one is somewhat  insurmountable.

The Diagnostic Instruments clamp includes a nifty focusing ring which allows the camera to be made parfocal with the binocular observing head – very handy for taking photos. It is circled in the image below.

Unfortunately, the clamp focusing mechanism does not have enough travel to bring the EOS par-focal with the binocular head – but it does work very well with the Mikrocam. So why would I chose the EOS over the Mikrocam, as this feature is so useful?

I did attempt some photos of a slide of fossilised material – you can see in the photos below that focusing is the major issue:

Andy