I have had about 30 years experience cutting ultrathin
sections (and if I do say so, I am pretty good at what
I do). The routine thickness (a silver-grey color by
reflected light in the boat) is in the 75 to 90nm
range, and usually fairly easily obtained. Under
really good circumstances, I can cut grey sections
(about 70nm). With a lot of sweat and some choice
words I can cut down to black--generally considered to
be in the 40 to 50nm range. So, how would you know if
you had cut sections in the 30nm range? They are too
thin to provide interference color; thus, picking them
up would be "difficult". If, on the odd chance you
did cut 30nm sections and overcame the "difficult"
part, you would have to pick them up on coated
grids--which immediately obviates the advantage of
cutting them that thin to begin with. The standard in
TEM is the 70 to 90nm thickness for routine biological
work, and these can usually be picked up on uncoated
grids. Another point about the 20 to 30nm range:
contrast. Properly stained sections give a good gray
scale range. My experience with the supposed 50nm
sections (and I have done this) is that the contrast
drops off so rapidly that now the microscopist is
faced with trying to interpret (or worse, print)
images of extremely low contrast.
There are reasons for using very thin samples--but the
results of processing, embedding and staining reduce
the useful resolution within the sample to such a
point that one gains nothing by going to sections less
than 70nm thick. Well, I can think of one
possibility: if one is embedding and sectioning
protein crystals, perhaps the 50nm or less sections
could provide a "single molecular layer" slice of the
crystal. I would suggest a search through PubMed on
section thickness or ultrathin sections for some
published work in this area. Most/many of the papers
will likely be in the 1970s with a few more recently.
Try Journal of Ultrastructure Research (now J of
Structural Biology), Journal of Microscopy and
Roger Moretz, Ph.D.
Dept of Toxicology
--- Ephram Shizgal wrote:
> I have been interested in tossing a general
> microtome/sectioning question
> out to the list and this seems like a good place for
> it to go:
> If someone were to suggest that you needed to
> produce sections between 20
> and 30 nm to use on a specific TEM to get results,
> is this a thickness that
> most of you feel is attainable (albeit something of
> a challenge) or, in
> fact, too challenging to be routinely considered ?
> Thanks in advance for the feedback.
> Ephram Shizgal
> Delong America (a member of the LVEM5 Group of
> Delong Instruments)
> In USA and Canada: 1-866-DELONGUSA (1-866-335-6648)
> International: 514-904-1202
> For LVEM5 (Low Voltage Electron Microscope)
> information please visit
> -----Original Message-----
> From: Georger, Mary
> Sent: Wednesday, January 29, 2003 7:45 PM
> To: HistoNet Server; 'Timothy R. Wheelock'
> Subject: RE: MICROTOMES
> Hi Tim,
> I have dhad the Microm HM 355 S for several years
> and asolutely love it! We
> do many very tedious serial cuts of entire vessels,
> and the machine makes my
> life so much easier. It is reliable, easy to learn,
> and thus far I have had
> no problems.
> Best of luck!
> Mary Georger
> Center for Cardiovascular Research
> University of Rochester Medical Center
> KMRB Room 2-9816
> 601 Elmwood Avenue.
> PO Box 679
> Rochester, New York 14642
> > ----------
> > From: Timothy R. Wheelock
> > Sent: Wednesday, January 29, 2003 4:30 PM
> > To: HistoNet Server
> > Subject: MICROTOMES
> > Hi Everyone:
> > I am in the market for a microtome.
> > I have tested 3 fully automated machines and 3
> manual machines. The
> > electric microtomes are the Richard Allan Microm
> HM 355 S, the Shandon
> > Finesse ME, and the Leica RM 2155. I did reference
> checks on all 3
> > electric machines.
> > I eliminated the manual machines (by the same 3
> companies) mainly
> > because I found their flywheels much harder to
> turn than my present 30
> > year old AO820!
> > I am leaning toward 1 of the electric microtomes,
> but thought I would
> > ask people's opinion.
> > One of the many issues i am concerned about is
> durability, workmanship,
> > whether a machine is solidly built.
> > Thanks for your time,
> > Tim Wheelock
> > Harvard Brain Tissue Resource Center (Brain Bank)
> > McLean Hospital
> > Belmont, MA
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