SPI-Chem™ HPMA Water Soluble Kit Instructions
Complete use instructions!
Precautionary Statement
Thoroughly read all instructions and warnings on container labels and also be
familiar with the MSDS sheets prior to use of this kit.
Introduction
HPMA, or 2-hydroxypropyl methacrylate embedding media is a water soluble monomer
that is finding increasing numbers of applications in the field of electron
histochemistry as well as in the materials sciences. Indeed certain types of
polymer and other samples that, because of the fragility of fine structures, such
systems absolutely can not be dried out without changing the sample and therefore
the only option is to embed without every bringing the sample to a "dried out" state.
The SPI-Chem HPMA kit contains the following components:
1. Hydroxypropyl methacrylate (HPMA)
2. Azo-bis-iso-butyronitrile
3. 2,4-Dichlorobenzoyl peroxide
Procedure - Over view
The specimen is dehydrated and impregnated (infiltrated) step-wise with
prepolymerized HPMA containing successively less water (20% and 3.0%) and 0. 1%
azo-bis-iso-butyronitrile or 2, 4-Dichlorobenzoyl peroxide as a catalyst for
subsequent polymerization with ultra-violet (UV) light or heat.
Water is added to separate aliquots of the HPMA prepolymer, to make 80% and 97%
HPMA solutions. A typical impregnation using the above prepolymers includes 2
changes, 1 hour each for both the 80% and the 97% HPMA.
About 5 ml of fluid is used for each specimen, placed in glass vials and agitated
for the duration of the impregnation. Should the HPMA turn cloudy from excess
water, replace the wet HPMA with a fresh aliquot before proceeding further. Each
change of HPMA contains 0.1% of azo-bis-iso-butyronitrile to ensure complete impregnation of samples
with the catalyst.
Embedding and Polymerization
The samples are then placed in fresh 97% prepolymer in gelatin capsules.
To obtain a softer and less brittle final polymer, up to 10% water can be used
during the final polymerization (e.g. 90% prepolymerized HPMA). The capsules
must be filled to capacity, to ensure the exclusion of all air, when their tops
are pushed down tight. This final point is very very important in terms of
determining the final outcome of the experiment.
Polymerization can be accomplished by exposing the gelatin capsules to long
wavelength UV light (315 nm), at low temperatures (1-3° C) for 12 to 24 hours.
Alternatively, for heat polymerization, the capsules can be maintained at 56-60°
C for two to three days.
Safety considerations
The preparation and mixing of the resin component must be thorough and always be
performed in a fume hood with adequate ventilation.
Further on the prepolymerization of HPMA:
According to the procedures of Leduc and Bernhard1 and Leduc and
Holt2 quantities of up to 30 ml of HPMA, containing 0.1% azonitrile
catalyst are partially polymerized by heating
to about 90° C or until it starts to show the evolution of steam. Once steam
starts to be evolved, immerse the beaker in an ice bath for rapid cooling. The
HPMA is now ready to use.
1. Leduc, E. H. and Bernard, W. V., J. Ultrastruct. Res. 19, 196 (1967).
2. Leduc, E. H., and Holt, S. J., J. Cell Biol. 26, 137 (1965).
The information given in this set of use instructions is to the best of our
knowledge, accurate, but no warranty is expressed or implied. It is the user's
responsibility to determine the suitability for his own use of the products
described herein; and since conditions of use are beyond our control, we disclaim
all liability with respect to the use of any material supplied by SPI Supplies.
Nothing contained herein shall be construed as a recommendation to practice any
patented invention nor as a recommendation to use any product or to practice any
process in violation of any law or any government regulation.
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Saturday July 05, 2008
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