100 x oil for water life

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100 x oil for water life

#1 Post by janvangastel » Mon Jan 20, 2020 9:45 am

I have tried to use my 100 x oil objective to look at micro organisms in a water sample, but as soon as the eyepiece touches the oil, the water below the cover glass starts moving and the subject moves out of the FOV. Is it impossible to use an oil objective for this purpose, or do I do something wrong?

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Re: 100 x oil for water life

#2 Post by mrsonchus » Mon Jan 20, 2020 11:30 am

Hi Jan, a tricky manoeuvre for sure!
I'd try to secure or even seal the coverslip before going to oil. Perhaps a couple of dabs of quick-setting 'stuff' at a few points around the coverslip/slide edges. I wonder if some small dabs of wax may be able to secure the coverslip and remain unaffected by the water. Even a few small dabs of vaseline or similar may be enough to stop the coverslip wandering from the drag imposed by the oil/objective upon it?
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Re: 100 x oil for water life

#3 Post by MicroBob » Mon Jan 20, 2020 11:47 am

Hi Jan,
you can smear a bit of vaseline on a flat surface and scrape the cover slip endes along the surface. this leaves a tiny wall of vaseline that will seal the slide against the oil and stop the cover slip from moving. Then there are immersion oils of different viscosities. Obviously your water layer has to be thin due to the small working distance of the objective.


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Re: 100 x oil for water life

#4 Post by Hobbyst46 » Mon Jan 20, 2020 1:06 pm

MicroBob wrote:
Mon Jan 20, 2020 11:47 am
Then there are immersion oils of different viscosities.
There are two common types of immersion oil, named Type A and Type B respectively (Cargille nomenclature), of low and high viscosity respectively. The viscosity difference is large. I recommend Type A, but even with it, securing the coverslip to the slide with grease/vaseline/paraffin/other stuff 9as mentioned by others) is essential.
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Re: 100 x oil for water life

#5 Post by 75RR » Mon Jan 20, 2020 3:44 pm

I think one of the main points is to make sure that the water layer is as thin as possible.

Check the working distance of the objective - a cover slip lying on top of too much water may well exceed this distance.

If it there is too much water then the objective pushes against the cover slip as one attempts to focus deeper.

Dabbing the edges of the cover slip with absorbent/tissue paper is a good way to lower the water level quickly.

Make sure that there is little to no detritus on the slide, as it will impede the cover slip from lying flat.
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Re: 100 x oil for water life

#6 Post by apochronaut » Mon Jan 20, 2020 4:09 pm

You can use an oil immersion objective without securing it in theory, as long as your sample preparation and tools are accurate. Having to use paraffin or risers under the slip, certainly detracts from the ease and joy of using an oil immersion objective and helps to magnify the reasons so many unseasoned microscopists shy away from using oil immersion. Messy.
First, it is best to choose larger covers. The little 20 x 20 mm ones that are cheap and easily available are o.k. for general dry microscopy but for oil immersion choose the 22 x 30 mm size. I also choose # 0 slips for oil. This allows for a deeper drop of oil , which exerts less drag against the slip.
Second, try making some test slides, using various tools with which to install the sample on the slide. For each coverslip size there is an ideal volume of liquid that you want under it. In my case, I use 22 x 30mm covers and have an eyedropper with a mark on it, which collects the ideal amount. Some people use pipettes.Even a wooden stick or some sort of dipper will do. Once you get used to collecting the right volume, you are on the right track.
Third, learn to draw under the cover. Drawing under the cover, is a technique, where you can replace water lost to evaporation without disturbing the sample much, if at all in some cases. The term refers to drawing water, not with a pencil. Using a wick of tissue on one side of the cover, you administer a little fresh sample or distilled water from the other side, filling the space under the cover after some evaporation has taken place.
Fourth, if you need to stick or seal the cover somehow, vaseline or paraffin can be used but are cumbersome and usually a bit messy to clean up....no moreso than oil though, if you prefer to reuse the glass. You only need to stick the cover enough though, not permanently. For this I use pectin. Pectin is cheap, water soluble and provides enough adherence. It is available in small boxes of crystals for less than 2.00 in most stores that sell fruit. I mix 1/8 tsp. or less with slightly less water and stir it until the crystals have dissolved, with the end of a wooden skewer. I also apply it along the edge of the cover with the same skewer. The pectin won't harden, just get real thick. I usually put it on two opposing sides only, so that if need be I can draw more water or sample under the slip.
Those cheap wooden barbecue skewers are good general purpose dippers and mixers for microscopy. One drop clinging to the end of the pointed end of the ones I have, which are 2.65mm in diameter, is enough for a 20 x 20mm cover and two drops enough for a 22 x 30. The sample should take a few seconds to flow to the corners of the slip, otherwise you have too much. If you end up having too little, you can draw a little more under, rather than lift the slip.

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