when i want to try thick sectioning of my sample, I fill in the tray with water bu even with angular and axial regulations and also filling and vacuuming water , it does not wet the upper part ! and so does not approach the edge ... I do not know what is the problem...is it from the water or the glass or maybe cleanliness.... ?please advice
Long, since no simple answer! Otherwise: apologies!
Dear Seyede:
first of all: you should provide further detail on the knife/knives you are talking about...
For Ultramicrotomy usually one uses "knives" made from glass strips in a distinct fashion by scoring and breaking, then the resulting triangular pieces containing the cutting edge have to be mounted with a kind of self-adhering tape (usually a special type of golden or silver 'Scotch' TM tape, + additional 'glueing' and 'sealing' the "hole" on the back with dental wax) to get the water-trough necessary for collecting the ultrathin sections easily. As long as one does not know about your practice and making knives one cannot help you effectively...
As a 'shot into the blue': 5 possibilities:
i) the cutting edge isn't properly forming (here it would be of essence to know if you use a "knife maker" and if yes: which one...? how your cutting edge looks like?),
ii) you're using wrong glass quality,
iii) the small (approx. 4-6 mm) ridge of your glass might have gotten "oily" or "greasy" somehow... For the latter you have not much options (CAREFUL, but thorough cleaning with p.a. acetone) OR make new knives, taking care not to contaminate again the knife's ridge...,
iv) your trough("boat")(or silver-tape) isn't glued properly (upper edge must be positioned orthographically /vertically to the edge (hope you know what I mean!) and last but not least,
v) your glass has tension ( produces and adverse surface tension for wettability*) by the trough water you fill in best with a syringe .... BUT BE AWARE that unused injection needles as well as syringes taken "freshly" out of / from plastic package often are coated with silicone oil which prior to use in ultramicrotomy must be eliminated: bathingand sonicating at least thrice in p.a. acetone....
Disclaimer: having done such work hands on at the bench by myself for more than 10 years I know about those problems and how constraining and frustrating such "presumably technical" problems can be...
Last point: you might convince your PI/Lab-Head to buy at least ONE diamond knife (you know for sure a / the source)...otherwise I would be able to tell as a former satisfied user....
*) wettability issues can be improved either by buying special "wetting solutions" or (if you have no hygienic concerns) just to spit into a small volume of distilled water, fill into clean glass or plastic bottle, shaking vigorously, apply some drops into the knife's trough fluid, let stand and react for some minute [or even overnight] and then wash thoroughly with a. bidest. (e.g. jet-washing/flushing by means of a wash-bottle). Active principle: removal or at least reduction of surface tension by the composition of saliva (esp. enzymes like amylase and or lipase)....
If you own a DDK-diamond knife, their manual for working and taking care of the diamond knife describe the use of a low amount of acetone in the trough water / fluid (cf: http://www.ddk.com/PDFs/knifecareanduse.pdf ) but NOT all manufacturers of diamond knives recommend the adding of a harsh organic solvent like acetone (cf. 100% Ethanol - section E. Trouble shooting in https://www.diatomeknives.com/diamond_knives/manual.aspx ; also www.diatome.ch/Messerprospekt_01_2015_EN_LowRes.pdf; https://www.emsdiasum.com/microscopy/products/diamond/ultrathin.aspx; https://www.agarscientific.com/media/import/14-diamond_knives_pgs_270-276_date_17_06_10_web.pdf ; ).
further reading on ResearchGate perhaps: https://www.researchgate.net/post/Can_anyone_suggest_how_to_increase_wettability_of_a_glass_slide_while_making_thin_film_of_FTO_solution
Web: www3.ncc.edu/faculty/BIO/becks/Path3f/EM_Manual.pdf ;
Article Wettability of a Glass Surface in the Presence of Two Nonion... http://Pubs.acs.org/doi/abs/10.1021/la8008078
= (Notabene: add 'http://pubs. ' in front of the following address:) acs.org/doi/abs/10.1021/la8008078 ; https://books.google.at/books?isbn=0323150292 p.132: Use of 0.1-1.0% non-ionic detergent such as Triton X-100 or Freon 113 (the latter as of today obsolete and perhaps forbidden too in your country), BUT DO NOT USE common laboratory detergents like Alconox , etc. (HAYAT M.A.,1986: in 'Basic Techniques For Transmission Electron Microscopy,p.132);
or:
Article Care and Use of Diamond Knives
(HAYAT and ZHANG 1998) = (Notabene: add 'www. ' in front of the following address:) sciencedirect.com/science/article/pii/S096843289800016X
Regards, W.M.(private mail: [email protected])
W
Long, since no simple answer! Otherwise: apologies!
Dear Seyede:
first of all: you should provide further detail on the knife/knives you are talking about...
For Ultramicrotomy usually one uses "knives" made from glass strips in a distinct fashion by scoring and breaking, then the resulting triangular pieces containing the cutting edge have to be mounted with a kind of self-adhering tape (usually a special type of golden or silver 'Scotch' TM tape, + additional 'glueing' and 'sealing' the "hole" on the back with dental wax) to get the water-trough necessary for collecting the ultrathin sections easily. As long as one does not know about your practice and making knives one cannot help you effectively...
As a 'shot into the blue': 5 possibilities:
i) the cutting edge isn't properly forming (here it would be of essence to know if you use a "knife maker" and if yes: which one...? how your cutting edge looks like?),
ii) you're using wrong glass quality,
iii) the small (approx. 4-6 mm) ridge of your glass might have gotten "oily" or "greasy" somehow... For the latter you have not much options (CAREFUL, but thorough cleaning with p.a. acetone) OR make new knives, taking care not to contaminate again the knife's ridge...,
iv) your trough("boat")(or silver-tape) isn't glued properly (upper edge must be positioned orthographically /vertically to the edge (hope you know what I mean!) and last but not least,
v) your glass has tension ( produces and adverse surface tension for wettability*) by the trough water you fill in best with a syringe .... BUT BE AWARE that unused injection needles as well as syringes taken "freshly" out of / from plastic package often are coated with silicone oil which prior to use in ultramicrotomy must be eliminated: bathingand sonicating at least thrice in p.a. acetone....
Disclaimer: having done such work hands on at the bench by myself for more than 10 years I know about those problems and how constraining and frustrating such "presumably technical" problems can be...
Last point: you might convince your PI/Lab-Head to buy at least ONE diamond knife (you know for sure a / the source)...otherwise I would be able to tell as a former satisfied user....
*) wettability issues can be improved either by buying special "wetting solutions" or (if you have no hygienic concerns) just to spit into a small volume of distilled water, fill into clean glass or plastic bottle, shaking vigorously, apply some drops into the knife's trough fluid, let stand and react for some minute [or even overnight] and then wash thoroughly with a. bidest. (e.g. jet-washing/flushing by means of a wash-bottle). Active principle: removal or at least reduction of surface tension by the composition of saliva (esp. enzymes like amylase and or lipase)....
If you own a DDK-diamond knife, their manual for working and taking care of the diamond knife describe the use of a low amount of acetone in the trough water / fluid (cf: http://www.ddk.com/PDFs/knifecareanduse.pdf ) but NOT all manufacturers of diamond knives recommend the adding of a harsh organic solvent like acetone (cf. 100% Ethanol - section E. Trouble shooting in https://www.diatomeknives.com/diamond_knives/manual.aspx ; also www.diatome.ch/Messerprospekt_01_2015_EN_LowRes.pdf; https://www.emsdiasum.com/microscopy/products/diamond/ultrathin.aspx; https://www.agarscientific.com/media/import/14-diamond_knives_pgs_270-276_date_17_06_10_web.pdf ; ).
further reading on ResearchGate perhaps: https://www.researchgate.net/post/Can_anyone_suggest_how_to_increase_wettability_of_a_glass_slide_while_making_thin_film_of_FTO_solution
Web: www3.ncc.edu/faculty/BIO/becks/Path3f/EM_Manual.pdf ;
Article Wettability of a Glass Surface in the Presence of Two Nonion... http://Pubs.acs.org/doi/abs/10.1021/la8008078
= (Notabene: add 'http://pubs. ' in front of the following address:) acs.org/doi/abs/10.1021/la8008078 ; https://books.google.at/books?isbn=0323150292 p.132: Use of 0.1-1.0% non-ionic detergent such as Triton X-100 or Freon 113 (the latter as of today obsolete and perhaps forbidden too in your country), BUT DO NOT USE common laboratory detergents like Alconox , etc. (HAYAT M.A.,1986: in 'Basic Techniques For Transmission Electron Microscopy,p.132);
or:
Article Care and Use of Diamond Knives
(HAYAT and ZHANG 1998) = (Notabene: add 'www. ' in front of the following address:) sciencedirect.com/science/article/pii/S096843289800016X
Regards, W.M.(private mail: [email protected])
W
If nothing of said above helps, then may be your knife is too dirty. Try to clean it according to manufacturer procedure, for example this:
https://www.emsdiasum.com/microscopy/products/diamond/use.aspx
Soaking a knife (overnight) in a weak dishwasher detergent solution not only helps in its cleaning, it makes diamond/glass hydrophilic and solves a problem you described.
Back to simple solution. Wolfgang already mentioned (presumably) the best wetting agent for a knife - saliva. It makes wonders. My procedure is a combination of procedures described by Wolfgang and Matthew. I lick my forefinger and a thumb and drag a hair of an eyelash tool between them. Then slide the hair along knife and drag water to it's edge.
Hello,
In my experience a glass knife that has been used before is harder to wet ie get water right to the edge. So for semithin sections I persevere with Matthew's method. Once some resin has been cut water will often adhere; sometimes only where there is resin! For ultrathin sections I use an unused glass knife. One may still need to push water up to the edge. I use a glass pipette.
- Badges
- Science topic
- Similar topics
- Physical Sciences
- Materials Science
- Materials
- Inorganic Materials
- Ceramics
- Glass