From the standpoint if "why they work so well", there is a general
perception that sections, both thin and thick, have rougher surfaces than
epoxy embedments yielding more surface area, thereby exposing more antigenic
sites per unit projected area of the section, and at the same time,
increasing the sensitivity of detection.
Complete instructions are included with each kit. If you are a first time user, or just want to
know more about what has been published previously by others,
we offer a list of useful references on the use of the various Lowicryl resins in various applications.
Polymerization time at low temperature:
There are probably as many protocols for the polymerization as there are
people using Lowicryl. A typical protocol might call for a UV
polymerization -20° C for 2-3 days and then a further 1-2 days at room
temperature, which, apparently, at least in most instances, does not affect
immuno labeling sensitivity, even at the TEM level. This additional room
temperature curing does seem to make the block easier to section.
Handling precaution:
Keep in mind that these kits involve the use of methacrylic acid ester preparations
and the toxicological properties of these products are not fully known. Some users
have reported the development of allergies and/or sensitivities to these materials.
One should keep these facts in mind when handling these particular resin systems and at times,
when making the decision to even use them if you have had previous experience with sensitivities
to methacrylate resins. Good laboratory quality vinyl gloves
are recommend for the use and handling of all Lowicryl products.
Low temperature polymerization:
Keep in mind that these resins were developed for use at low temperature
using a UV cure. SPI offers an easy-to-use
low temperature UV curing chamber for this type of embedding.
Lowicryl K Type Kits
K kits are polar and therefore hydrophilic. The systems are
"partially water soluble" meaning that a complete dehydration
is not necessary. Indeed polymerization can occur quite nicely
with up to 5% water present. Therefore for immunolabeling with
these polar resin kits, one obtains both a better preservation
of structure as well as antigenicity and at the same time also
a lower level of nonspecific or background labeling. Note however,
that these kits are not completely water soluble, and at least
some dehydration is necessary. In those instances where
absolutely no alcohol dehydration can be tolerated, consider using the
SPI Chem® Low Acid GMA.
In those instances where high lipid content is present, consider using
SPI Chem® Low Acid HPMA.
An understated potential safety risk:
We are addressing now the cured block, something that to most people is about as
inert of a material as one will find. But the standard practice in many laboratories
is to use a small jeweler's (or even a small hack) saw to cut the block down to the
right size, sometimes even to shape it.
We want to address the dust that is generated and how its exposure can and should be
minimized. |
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